Some plasma lipid values in male Syrian golden hamsters with MS (in each group n=10).
\r\n\tCertain parameters are vital for their designing viz; non-Newtonian, time-dependent, shear-thinning and viscoplastic flow behavior controls physicochemical characters of designing of nano-structures that portray substantial wide range as well specific and unique inherent rheology in order to obtain myriad utilities including photovoltaics, catalysis, optics, drug/gene/cell delivery, energetic smart materials and energy storage. Bottom-up technique supported on self- and directed-assembly of polymer-based building blocks are some few powerful means utilized to get robust fabrication in resultant efficiently manipulated target nanostructures. This book compiles a brief review and case studies of certain recent advances in assembly and reconfigurable materials of such rationally designed polymer-based nano-structures/nano-composites to highlight their role in discovering the fundamental principles of assembly science besides providing critical design tools for assembly engineering of complex nanostructures. This proposed book will strive towards providinga comprehensive overview of scientific advances in reconfigurable materials with highlighted facts and findings which could aid readers to recognize both basic and applied directions in desired in development of S&T. It will provide promising methodical approach to relevant issues/concern problems commonly faced by most researchers especially in polysaccharide, biocomposites, dendrimeric tweezers, hybrids, nanoencapsulation, nanodevice, nano-sensors and nano-emulsions. Targeted readers include both basic scientists, clinicians, pharmaceutical, biomedical, engineers, technicians, researchers and students scholars who can use it as tool to percept rationales for the development of novel prospective/myriad utility of reconfigurable structured materials/matrixes.
",isbn:"978-1-78985-514-2",printIsbn:"978-1-78985-513-5",pdfIsbn:null,doi:null,price:0,priceEur:0,priceUsd:0,slug:null,numberOfPages:0,isOpenForSubmission:!1,hash:"bc49969c3a4e2fc8f65d4722cc4d95a5",bookSignature:"Dr. Rajendra Sukhadeorao Dongre and Dr. Dilip R. Peshwe",publishedDate:null,coverURL:"https://cdn.intechopen.com/books/images_new/7960.jpg",keywords:"Metamaterials, Glass-composite, Dendrimeric tweezers, Graphite, TiO2, Graphine oxide, Self-assembly, Bio-silicification, Nanocomposite, Nanoencapsulation, Bio-polymer, Dendrimer",numberOfDownloads:null,numberOfWosCitations:0,numberOfCrossrefCitations:0,numberOfDimensionsCitations:0,numberOfTotalCitations:0,isAvailableForWebshopOrdering:!0,dateEndFirstStepPublish:"May 2nd 2019",dateEndSecondStepPublish:"May 23rd 2019",dateEndThirdStepPublish:"July 22nd 2019",dateEndFourthStepPublish:"October 10th 2019",dateEndFifthStepPublish:"December 9th 2019",remainingDaysToSecondStep:"7 months",secondStepPassed:!0,currentStepOfPublishingProcess:5,editedByType:null,kuFlag:!1,editors:[{id:"188286",title:"Dr.",name:"Rajendra",middleName:"Sukhadeorao",surname:"Dongre",slug:"rajendra-dongre",fullName:"Rajendra Dongre",profilePictureURL:"https://mts.intechopen.com/storage/users/188286/images/system/188286.jpg",biography:"Rajendra S. Dongre received his M.Sc. from the Department of\nChemistry, R.T.M., Nagpur University in 1996 (Gold Medalist)\nand his PhD in 2010. His research work includes organic synthesis, chitosan bio-composites, matrixes, and remediation of water\npollution de-fluoridation, nitrate, chromium, and phosphate\nlead (II). He has worked as a Scientists-B in CSIR-LAB, National\nEnvironmental Engineering Research Institute (NEERI) Nagpur\nM.S., India. Overall, he has 25 years of experience in research and development\nand 17 years of post-graduate teaching experience, which has resulted in fifty-five\ninternational/national research publications. He has also guided four PhD research\nstudents. He received the 6th National Award (runner-up position) for Technology\nInnovation in Petrochemicals and Downstream Plastics Processing Industry, for\nresearch in the field of polymer science and technology, handed by Hon’ble Ananth\nKumar, Petrochemical & Fertilizers Minister of Government of India in 2016. He\nreceived the 5th National Science & Technology Award for research contribution in\nthe field of developing science in 2017, by EET-CRS, Noida, India.",institutionString:"RTM Nagpur University",position:null,outsideEditionCount:0,totalCites:0,totalAuthoredChapters:"6",totalChapterViews:"0",totalEditedBooks:"1",institution:{name:"Rashtrasant Tukadoji Maharaj Nagpur University",institutionURL:null,country:{name:"India"}}}],coeditorOne:{id:"289232",title:"Dr.",name:"Dilip R.",middleName:null,surname:"Peshwe",slug:"dilip-r.-peshwe",fullName:"Dilip R. Peshwe",profilePictureURL:"https://mts.intechopen.com/storage/users/289232/images/system/289232.jpg",biography:"Prof. Dilip R. Peshwe is currently working as a Professor at the Department of Metallurgical and Materials Engineering and is also a Dean (Faculty Welfare) at Visvesvaraya National Institute of Technology, Nagpur, (M.S.), India, for more than 35 years. Till date he guided 15 PhD research scholars, and holds 5 patents. He published 156 peer reviewed international research publications in SCI and Scopus journals. Besides, he completed 25 R&D projects with funds/amounting Rs 30 Crores. He was awarded SAIL Gold Medal, Best S&T Innovation Award of Government of India, Jawaharlal Nehru memorial trust award and also achieved three Best Research Paper awards at international workshops/conferences/seminars. He also authored three books and edited three books. Apart from this, he developed ten technologies for the rural areas to improve the productivity and quality transferring all these technologies at various clusters.",institutionString:"Visvesvaraya National Institute of Technology",position:null,outsideEditionCount:0,totalCites:0,totalAuthoredChapters:"0",totalChapterViews:"0",totalEditedBooks:"0",institution:null},coeditorTwo:null,coeditorThree:null,coeditorFour:null,coeditorFive:null,topics:[{id:"14",title:"Materials Science",slug:"materials-science"}],chapters:null,productType:{id:"1",title:"Edited Volume",chapterContentType:"chapter",authoredCaption:"Edited by"},personalPublishingAssistant:{id:"177730",firstName:"Edi",lastName:"Lipovic",middleName:null,title:"Mr.",imageUrl:"https://mts.intechopen.com/storage/users/177730/images/4741_n.jpg",email:"edi@intechopen.com",biography:"As an Author Service Manager my responsibilities include monitoring and facilitating all publishing activities for authors and editors. From chapter submission and review, to approval and revision, copyediting and design, until final publication, I work closely with authors and editors to ensure a simple and easy publishing process. I maintain constant and effective communication with authors, editors and reviewers, which allows for a level of personal support that enables contributors to fully commit and concentrate on the chapters they are writing, editing, or reviewing. I assist authors in the preparation of their full chapter submissions and track important deadlines and ensure they are met. I help to coordinate internal processes such as linguistic review, and monitor the technical aspects of the process. As an ASM I am also involved in the acquisition of editors. Whether that be identifying an exceptional author and proposing an editorship collaboration, or contacting researchers who would like the opportunity to work with IntechOpen, I establish and help manage author and editor acquisition and contact."}},relatedBooks:[{type:"book",id:"6518",title:"Chitin-Chitosan",subtitle:"Myriad Functionalities in Science and Technology",isOpenForSubmission:!1,hash:"2bbe245f1821a6691cc6d07e5b3462cf",slug:"chitin-chitosan-myriad-functionalities-in-science-and-technology",bookSignature:"Rajendra Sukhadeorao Dongre",coverURL:"https://cdn.intechopen.com/books/images_new/6518.jpg",editedByType:"Edited by",editors:[{id:"188286",title:"Dr.",name:"Rajendra",surname:"Dongre",slug:"rajendra-dongre",fullName:"Rajendra Dongre"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"7360",title:"Fillers",subtitle:"Synthesis, Characterization and Industrial Application",isOpenForSubmission:!1,hash:"4cb5f0dcdfc23d6ec4c1d5f72f726ab4",slug:"fillers-synthesis-characterization-and-industrial-application",bookSignature:"Amar Patnaik",coverURL:"https://cdn.intechopen.com/books/images_new/7360.jpg",editedByType:"Edited by",editors:[{id:"43660",title:"Associate Prof.",name:"Amar",surname:"Patnaik",slug:"amar-patnaik",fullName:"Amar Patnaik"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"6805",title:"Electrical and Electronic Properties of Materials",subtitle:null,isOpenForSubmission:!1,hash:"f6b6930e7ae9d0704f68b5c180526309",slug:"electrical-and-electronic-properties-of-materials",bookSignature:"Md. Kawsar Alam",coverURL:"https://cdn.intechopen.com/books/images_new/6805.jpg",editedByType:"Edited by",editors:[{id:"199691",title:"Dr.",name:"Md. Kawsar",surname:"Alam",slug:"md.-kawsar-alam",fullName:"Md. Kawsar Alam"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"6851",title:"New Uses of Micro and Nanomaterials",subtitle:null,isOpenForSubmission:!1,hash:"49e0ab8961c52c159da40dd3ec039be0",slug:"new-uses-of-micro-and-nanomaterials",bookSignature:"Marcelo Rubén Pagnola, Jairo Useche Vivero and Andres Guillermo Marrugo",coverURL:"https://cdn.intechopen.com/books/images_new/6851.jpg",editedByType:"Edited by",editors:[{id:"112233",title:"Dr.Ing.",name:"Marcelo Rubén",surname:"Pagnola",slug:"marcelo-ruben-pagnola",fullName:"Marcelo Rubén Pagnola"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"6802",title:"Graphene Oxide",subtitle:"Applications and Opportunities",isOpenForSubmission:!1,hash:"075b313e11be74c55a1f66be5dd56b40",slug:"graphene-oxide-applications-and-opportunities",bookSignature:"Ganesh Kamble",coverURL:"https://cdn.intechopen.com/books/images_new/6802.jpg",editedByType:"Edited by",editors:[{id:"236420",title:"Dr.",name:"Ganesh Shamrao",surname:"Kamble",slug:"ganesh-shamrao-kamble",fullName:"Ganesh Shamrao Kamble"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"6517",title:"Emerging Solar Energy Materials",subtitle:null,isOpenForSubmission:!1,hash:"186936bb201bb186fb04b095aa39d9b8",slug:"emerging-solar-energy-materials",bookSignature:"Sadia Ameen, M. Shaheer Akhtar and Hyung-Shik Shin",coverURL:"https://cdn.intechopen.com/books/images_new/6517.jpg",editedByType:"Edited by",editors:[{id:"52613",title:"Dr.",name:"Sadia",surname:"Ameen",slug:"sadia-ameen",fullName:"Sadia Ameen"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"6656",title:"Phase Change Materials and Their Applications",subtitle:null,isOpenForSubmission:!1,hash:"9b257f8386280bdde4633d36124787f2",slug:"phase-change-materials-and-their-applications",bookSignature:"Mohsen Mhadhbi",coverURL:"https://cdn.intechopen.com/books/images_new/6656.jpg",editedByType:"Edited by",editors:[{id:"228366",title:"Dr.",name:"Mohsen",surname:"Mhadhbi",slug:"mohsen-mhadhbi",fullName:"Mohsen Mhadhbi"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"6320",title:"Advances in Glass Science and Technology",subtitle:null,isOpenForSubmission:!1,hash:"6d0a32a0cf9806bccd04101a8b6e1b95",slug:"advances-in-glass-science-and-technology",bookSignature:"Vincenzo M. Sglavo",coverURL:"https://cdn.intechopen.com/books/images_new/6320.jpg",editedByType:"Edited by",editors:[{id:"17426",title:"Prof.",name:"Vincenzo Maria",surname:"Sglavo",slug:"vincenzo-maria-sglavo",fullName:"Vincenzo Maria Sglavo"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"6188",title:"Solidification",subtitle:null,isOpenForSubmission:!1,hash:"0405c42586170a1def7a4b011c5f2b60",slug:"solidification",bookSignature:"Alicia Esther Ares",coverURL:"https://cdn.intechopen.com/books/images_new/6188.jpg",editedByType:"Edited by",editors:[{id:"91095",title:"Dr.",name:"Alicia Esther",surname:"Ares",slug:"alicia-esther-ares",fullName:"Alicia Esther Ares"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"3161",title:"Frontiers in Guided Wave Optics and Optoelectronics",subtitle:null,isOpenForSubmission:!1,hash:"deb44e9c99f82bbce1083abea743146c",slug:"frontiers-in-guided-wave-optics-and-optoelectronics",bookSignature:"Bishnu Pal",coverURL:"https://cdn.intechopen.com/books/images_new/3161.jpg",editedByType:"Edited by",editors:[{id:"4782",title:"Prof.",name:"Bishnu",surname:"Pal",slug:"bishnu-pal",fullName:"Bishnu Pal"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}}]},chapter:{item:{type:"chapter",id:"39537",title:"Antioxidant Complexes and Lipoprotein Metabolism – Experience of Grape Extracts Application Under Metabolic Syndrome and Neurogenic Stress",doi:"10.5772/48083",slug:"antioxidant-complexes-and-lipoprotein-metabolism-experience-of-grape-extracts-application-under-meta",body:'The oxidative hypothesis of atherosclerosis states that peroxide modification of LDL (or other lipoproteins) is important and probably required for the pathogenesis of arterial sclerotic disease; thus, there is an assumption that inhibition of LDL oxidation would increase or prevent atherosclerosis and its clinical consequences [1]. It is believed that the basis for the atherosclerotic plaque development is the foam cell formation from oxidized low-density lipoproteins (LDL) captured by monocytes and macrophages via scavenger-receptors.
Oxidation of LDL is also important for the healthy vessel functioning. High LDL concentrations can suppress the function of arteries in relation to release of nitric oxide from the endothelium, and many of such effects are mediated by the products of lipid oxidation [2]. Moreover, oxidized LDL inhibit the endothelium-dependent nitric oxide mediated relaxations in a rabbit isolated coronary arteries. Oxidized LDL induce apoptosis in the vascular cells, including macrophages, and this is prevented by nitric oxide [3].
One of the most important mechanisms of the inflammation proatherogenic effect is development of the systemic oxidative stress, and, as a consequence of proatherogenic abnormalities of the blood lipoprotein metabolism, there is appearance of antibodies to them, alterations of the main artery wall structure [4].
At the same time, on the one hand, a high atherogenicity of strongly oxidized LDL, especially tiny subfractions, has been confirmed; on the other hand, the oxidative stress is one of the causes of endothelial dysfunction.
Endothelium vascular wall cells are involved into the interaction with the pathogenic LDL[5]. While macrophages are being overloaded with esterified cholesterol, oxysterols and other biologically active substances, including powerful enzymes with a wide spectrum of action, a foam cell is formed from the macrophage. Yet so far to its apoptosis the foam cell secrets a wide complex of interleukins, enzymes, mediators. Many of them induce a local inflammatory process, destruction of the surrounding intercellular substance, damage of the fibrous structures and separate cells.
Many factors are considered as the most important factors for atherosclerosis development risk. Among such factors an important role belongs to the so-called proatherogenic states, including chronic stress and metabolic syndrome (MS) [6]. The proatherogenic character of stress is connected, first of all, with the activation of free radical oxidation and hyperlipidemia development.One of the principal statements of all contemporary conceptions of the atherosclerosis pathogenesis is thought to be the destruction of the cell membrane structure, which universal damage factor is peroxide oxidation of lipids (POL) [7].
It is well-known that free-radical processes play the leading role in atherosclerosis pathogenesis. So the antioxidants using in correction of proatherogenic states is fully explicable especially when we speak about natural antioxidants. Thus, the investigation of their biological effects under stress and metabolic syndrome is of grate interest and may be a perspective direction of research.
At the same time it is known that the enzymes associated with HDL, paraoxonase and PAF-acetyl hydrolase can hydrolyse biologically active lipids of mm-LDL, destroy monocyte aggregates and decrease the endothelial activation of mm-LDL [8]. HDL also contain a high concentration of tocopherol due to which they can be free radical scavengers as well.
Antioxidants protect LDL from peroxide oxidation and consequently from intensive uptake of LDL by macrophages decreasing the foam cell formation, theendothelium damage and possibility for lipids to infiltrate the intima. This condition supports the actuality of searching medicines for treating atherosclerosis, in which inhibition of the POL process plays an important part in the mechanism of their action [9]. Tocopherol, carotene, probucol, a number of plant medicines containing flavonoids are proposed as antioxidants.
The overwhelming majority of antioxidant substances used in pharmacotherapy are xenobiotics and so substrates of CYP system actvating ROS formation. Moreover some of them, such as probucol, leade to HDL-C decreasing.
Therefore, the substances of natural, in particular, plant origins that possess a complex activity draw attention of researchers.
Phenolic compounds are widely present in the world of plants; they are the most widespread product of the plant metabolism. Participation of polyphenols in redox processes to produce stable quinone structures by their phenolic forms reveals an antiradical direction of their action which provides their direct antioxidant activity. At present it has been proven that polyphenols as antiradical agents not only hinder the initiation of free radical oxidation, but also interrupt the chain of lipoperoxidation [10]. A great variety of studies carried out both in vitro and in vivo supports the ability of polyphenols to inactivate (“to bind”, “to scavenge”) the radicals that initiate chains of oxidation. First of all, it relates to the primary ROS - ∙O2-and ∙OH [11].
There are some data that such natural polyphenols as catechins and procyanidins exposed to the human blood plasma produce certain complexes primarily with ApoA-1, i.e. with HDL.
One of the richest sources of polyphenols is Vitisvinifera and products of its processing, in particular wine.
Phenolic substances of grapes, including flavonoids and other polyphenols of grape, wine and grape seeds, are of a great interest due to their antioxidant properties and the ability to scavenge free radicals [12].
Studiesin vitro have shown that grape, wine and grape seeds inhibit the oxidation of LDL. Theactivity of those substances as oxidation inhibitors in wine diluted 1,000 times markedly exceeded the analogous values for vitamins C and E [13]. It has been experimentally proven that red wine polyphenols slow down LDL oxidation processes and prevent platelet aggregation, thus preventing coronary heart diseases [14].
However, there is not a lot of research in this field yet. Arguments for anti-atherogenic properties of antioxidants are not enough. Results of convincing research are needed in order to decisively recommend antioxidants for treatment and prophylaxis of atherosclerosis.
Taking into account the leading role of the free-radical processes in atherosclerosis pathogenesis one can make a conclusion about expediency of using natural antioxidants in prophylaxis and correction of this disease. [15]. Consequently, the study of the antioxidant influence on the development of stress-reactions and metabolic syndrome (MS) with the purpose of prevention of harmful complications for the cardiovascular system is of undoubted interest.
A number of studies also confirm the ability of a natural antioxidant α-tocopherol to reduce the risk of cardiovascular system diseases developed in patients with MS. It has been found that administration of α-tocopherol limits oxidation and cytotoxicity of LDL in the blood plasma significantly, supports the vascular endothelial function and reduces the intensity of systemic inflammation in the conditions of MS. The inhibiting effect of this antioxidant on aggregation and adhesion of platelets, adhesion of monocytes to endothelial cells and the smooth muscle cell proliferation has also been shown. However,numbers of experimental studies confirm that the single use of α-tocopherol is not enough for prevention of cardiovascular diseases in patients with MS [16]. It has been determined that the use of α-tocopherol in combination with ascorbic acid and aspirin (as a thrombolytic drug) is more effective [8]. In the ASAP study, the combination of vitamins E plus C was also tested, and this significantly decreased the intima-to-media progression rates in human. The ATBC clinical study used a combination of vitamin E and β-carotene in human as a secondary prevention strategy; however, no benefit on major coronary events has been found. The large MRC/BHF Heart Protection Study (HPS) for secondary prevention also examined the benefit of the antioxidant combination (vitamins E and C and β-carotene).
A strong dose-dependent effect of α-tocopherol administration is one of the unwanted effects. It is known that even a slight increase of the α-tocopherol dose could affect lipoprotein oxidation, the endothelium function and the degree of systemic inflammation [12].
The results of Cambridge Heart Antioxidant Study (CHAOS) of using antioxidants in cardiology published in 1996 give the opportunity to say that in patients with true (confirmed by angiography) coronary atherosclerosis vitamin E administration (a daily dose of 544-1088 mg (400-800 МU) reduces the risk of non-fatal myocardial infarction. The overall mortality from cardiovascular diseases in this case does not decrease. A favourable effect is revealed only after one-year administration of tocopherol.
At the same time in the Heart Outcomes Prevention Evaluation (HOPE) study, which was devoted to the study of the action of both ramipril and vitamin E (400 МU/daily dose), it was found that use of this antioxidant during approximately 4.5 years did not cause any effect on either the primary (myocardial infarction, insult and death from cardiovascular diseases) or any other end points of research. In another large-scale study on the primary prophylaxis of atherosclerotic diseases in people at least with one risk factors (hypertension, hypercholesterolemia, obesity, preliminary MI of the closest relative or advanced age) vitamin E (300 МЕ/daily dose) was used during 3.6 years and did not reveal any effect on any of the end points (the incidence of cardiovascular events and death). The vitamin E effectiveness was also not confirmed for various other cases (hypercholesterolemia, the level of sportsmen training, sexual potency, retardation of aging processes, etc.).
Empirically vitamin E is used in various diseases; however, the majority of the reports about tocopherol effectiveness is based on the single clinical observation and experiment data. Nowadays there are no reliable results on the role of vitamin E in prevention of tumour diseases, though the ability to reduce formation of nitrosamines (potentially carcinogenic substances being formed in the stomach), to decrease the formation of free radicals and have antitoxic effects when using chemotherapeutic remediesis well-known. In addition, the long-term intake of vitamin E in the doses from 11 to 800 mg does not cause side effects.
In HDL Atherosclerosis Treatment Study (HATS) there was the treatment of atherosclerosis depending on the high density lipoprotein cholesterol (HDL-C) level; in 160 patients with coronary heart disease with the confirmed coronary artery stenosis and the low HDL-C level the higher (800 МU/day) dose of vitamin E than in HOPE was used. The treatment combination also included 1000 mg of vitamin C, 25 mg of β-carotene and 100 mg of selenium. The study lasted 3 years and revealed that antioxidants had no influence on the HDL-C level, but in combination with hypocholesterolemic drugs they reduced their effect on LDL-C and especially – on HDL-C.
The dominant carotenoid revealed in blood and various tissues (such as liver, kidneys, adrenal glands, ovaries and prostate) is lycopene. Due to its structure and mechanism of action lycopene belongs to the group of antioxidants; a lycopene molecule contains 13 double bonds, which can interact with free radicals. Like β-carotene lycopene can serve as a precursor of vitamin A. However, the lycopene antioxidant activity is two times stronger than that of vitamin A.
Lycopene is recommended as an adjuvant in the treatment of the following diseases: idiopatic male infertility, chronic prostatitis, preeclampsia and intrauterine growth retardation (IUGR), mastopathia, diabetes mellitus, cardiovascular diseases, leucoplakia, age-related degeneration of yellow spots and cataract [17]. As with other oxidants, lycopene is administered in immunodeficiency states against chronic infections and to reduce the harmful action of unfavourable environmental factors.
The most representative evidence of the antioxidants’ positive role in cardiovascular diseases prophylaxis was obtained in the multicultural European community multicentre study on antioxidants, myocardial infarction, and breast cancer (EURAMIC), during which the relationship between the antioxidant status and acute myocardial infarction in patients from 10 European countries was determined. The protective action was proven only for lycopene. In the Kuopio Ischemic Heart Disease Risk Factor Study (KIHD) the high level of blood plasma lycopene is associated with decreased risk of acute coronary syndrome and insult. In Erasmus Rotterdam Health Study (ERGO, also called “Rotterdam Study”) it has been proven that lycopene prevents development and progression of atherosclerosis.
The meta-analysis of 72 epidemiological studies conducted concerning the connection between the tomato intake and cancer has determined the associative feed-back between the blood plasma lycopene level and the risk of cancer in 57 studies and 35 from 57 obtained associations were statistically significant [18].
Probucol (phenbutol) is a hypolipidemic medicine and belongs to butyl phenol derivatives. Probucol is the medicine that is similar in structure to hydroxytoluene – the compound with the potent antioxidant properties.
The hypolipidemic effect of Probucol is caused by activation of non-receptor ways of LDL extraction from the blood. It is believed that the prominent antioxidant activity of probucol prevents LDL oxidation.
Probucol decreases the total cholesterol content in plasma due to intensification of the LDL catabolism at the final stage of cholesterol elimination from the organism. It also inhibits the cholesterol biosynthesis at early stages and to a small extent slows down the food cholesterol absorption. It does not influence the triacylglycerol and the VLDL content, but significantly decreases the antiatherogenic HDL level in the blood. It is believed that decrease of the HDL-C level reflects improvement of cholesterol esters transfer with HDL on acceptor lipoproteins due to increase of the cholesteryl ester transfer protein (CETP) activity.
In spite of undesired decrease in the HDL-C concentration probucol causes regression of xanthelasma; this effect is revealed best of all in patients with the most dramatic HDL-C decrease. This important observation demonstrates that the low HDL-C content is not undoubtedly a negative phenomenon. The data obtained in experiments with animals indicate that probucol due to its antioxidant properties prevents lipid peroxidation and thereby inhibits the LDL uptake by macrophages, therefore, it inhibits atherogenesis. This allows suggesting that the therapeutic effect of the medicine may not be connected with its ability to decrease the LDL level. There is no clinical evidence of this hypothesis at the moment.
The medicine is absorbed slowly when taken internally, it is readily soluble in the adipose tissue releasing gradually into the bloodstream, and so its action is kept for a long time (up to 6 months after discontinuation of the treatment).
When using probucol in MultiVitamins and Probucol (MVP) research the renewal of the endothelium function in patients with IHD, decrease of restenosis cases after coronary angioplasty (when taking it at least 4 weeks before the procedure and further treatment during 6 months) was observed. Other antioxidants (α-tocopherol in high doses (700 mg per day), β-carotene and vitamin C) turned out to be ineffective.
Combined application of the endogenous antiradical antioxidants is of particular interest. In HPS (Heart Protection Study) along with the study of the simvastatin effectiveness the prophylactic action of antioxidants was investigated. The use of the vitamin complex (600 mg of vitamin E, 250 mg of vitamin C and 20 mg of β-carotene per day) lasted in average 5.5 years and did not reveal any differences in placebo groups and groups taking vitamins. Moreover, if the tendency exists, it reflects increasing of vascular events in the antioxidant intent-to-treat group.The action of antioxidants was compared with the effect of the combined use of simvastatin and nicotinic acid (niacin). Moreover, one of the groups received simvastatine+niacin and antioxidants. Angiographic and clinical data of this study were also disappointing with respect to the use of antioxidants.
Unfortunately, a great part of the compounds synthesized, which are used for pharmacocorrection of these states, are xenobiotics, so they can activate the free-radical formation process. Synthetic antioxidants, in particular probucol, can not be recommended for patient use because they decrease the HDL-C level.
The lack of antioxidant medicines popularity and the absence of traditions of their common use in practical medicine are caused a number of reasons: unsatisfactory previous study of this issue, complexity of adequate estimation of oxidation state parameters in the organism and the absence of the effective medicines with the antioxidant activity that are able to quickly reduce the consequences of the oxidative stress.
Therefore, the main indications for using antioxidants are excessively activated free-radical oxidation processes accompanying different pathologies. The choice of specific medicines, correct indications and contraindications for their use has not been developed yet and require further research.
In our experiments we studied the indicators of lipid and lipoprotein metabolism in the blood plasma and the liver underthe experimental metabolic syndrome (MS) in Syrian hamsters of different sex and age.
In the experiments purebred male rats with 180-220 g of the body weight were used. The animals were kept in vivarium on a balanced diet. During 21 days the animals were givenlow alcoholic beverages from grapes of red and white gradesper os daily. These beverages were introduced in the maximum effective doses of 9 mg of polyphenols/100 g the body weight. Taking into account the fact that the polyphenol content in the beverages investigated was quite low,the effective dose was introduced 3 times a day by 2 ml of liquids per 100 g of the animal’s body weight. Control animals were introduced the corresponding volume of the saline solution. Ethanol was given in the corresponding dose.
Stress was caused by immobilization on the abdomen for 3 hours [19]. Animals were decapitated 3 hours after the immobilization. The blood was collected to get the serum. The liver was perfused by the cold extraction medium (0.25 M sucrose in 0.025 M tris-HCl, pH 7.5), homogenized in the Potter homogenizer with 2 ml of the extraction medium per1 g of the liver. All manipulations with animals were held under chloralose-urethane anaesthesia.
To distribute the plasma lipoproteins the samples were centrifuged at 65,000 rpm (342,000 g) for 4 h at 4 C in the Optima XL-100K ultracentrifuge (Beckman Coulter) set at slow acceleration and deceleration [20]. Samples were fractionated within 1 h of centrifugation.
Lipids were extracted with chloroform and methanol (1:2 v/v) twice, as described by Bligh et al [21], and the supernatant was collected for determination of TG and FFA. TG and FFA were determined by enzymatic colorimetric methods with commercial kits (Zhongsheng, Beijing, China). The total cholesterol content was detected with the help of standard enzymatic cholesteroloxidase kits of “Boehringer Mannheim GmbH diagnostica” firm (Germany). The total lipid concentration was determined with the help of a standard kit “Eagle Diagnostics” (USA) – the reaction with vanillin reagent.
Determination of the lipid peroxide product quantity was performed in heptane-isopropanol extracts [22]. The optical density was measured at the wavelength of 220 nm (for compounds with isolated double bonds), 232 nm (for diene conjugates) and 278 nm – for ketodienes and conjugate trienes.
The TBA content was determined on the spectrophotometer with the help of the reaction with thiobarbituric acid [23].
A modified version of the high performance liquid chromatography (HPLC) procedure developed by Stacewicz-Sapuntzakis et al. [24] was used to measure vitamins E in the plasma. The HPLC system included a 150 × 3.9 mm Nova-pak C18 (4 microns) column with a guard-pak pre-column (both from Waters, Milford, MA), Waters Millipore TCM column heater, Waters 490 multi-wavelength detector, Hitachi 655–61 processor, Hitachi 655A-11 liquid chromatography, and BioRad autosampler AS-100.
The serum ascorbic acid concentrations were measured as described by using HPLC [25] with salicylsalicylic acid as a deproteinizing agent, metaphosphoric acid as a stabilizer.
The serum PON1 activity was measured by the rate of generation of p-nitrophenol determined at 405 nm according to MacKness B et al. [26].
The plasma cholesterol ester transfer protein (CETP) activitywas examined using the modifications of Khosla et al. [44]. The CETP activity in duplicate10-μL aliquots of the plasma was determined after incubations with 3H-cholesterol ester (CE)-labeled HDL3 and LDL. Radioactivity transferred from 3H-HDL3 to LDL (measured in the supernatant after precipitation with heparin/MnCl2+) was used to calculate the CETP activity (expressed as the percentage of radioactivity transferred from 3H-HDL3 to LDL per 16 h of incubation).
To measure endothelium-bound LPL, the perfusion solution was changed to buffer containing 1% fatty acid–free BSA and heparin (5 units/ml). The coronary effluent was collected in timed fractions over 10 min and assayed for the LPL activity by measuring the hydrolysis of a sonicated [3H]triolein substrate emulsion [27].
The plasma LCAT activity was measured by determination of the amount of radioactivity in each spot calculating the free cholesterol/ total cholesterol ratio in each plasma sample before and after the LCAT reaction and thus estimating the esterificationrate [28]. The fractional esterification rate (%. h’) expressed as the percentage of the free cholesterol esterified in the plasma sample per hour.
The HL activity was evaluated using the glycerol-stabilized emulsion of triolein and egg phosphatidylcholine containing glycerol-tri[9,10(n)-3H] oleate by determination of the radioactivity amount during incubation [29].
Statistical analysis.All data were analyzed for statistical significance with SPSS 13.0 software. The data were presented as means ± standard deviation. Statistical analysis used one-way ANOVA. P<0.05 was considered to be statistically significant.
The results of our studies suggest the existence of significant changes in the lipid metabolism, as well as sex and age differences in the lipid and lipoprotein metabolism both in healthy animals and in animals with MS.
In male hamsters fed with a high-calorie diet atherogenic dyslipidemia develops independently of age (Table 1). As it can be seen from the data obtained, increase of the total lipid content in the animal blood plasma is caused by increasing of the ApoB-containing lipoprotein (ApoB-LP) level since the HDL content is not changed. At the same time it has been found that the plasma TAG level in young (47%) and in adult animals (30%) increased in comparison with the intact group.
Increase of the TAG blood content in conditions of MS is considered to be a key factor for development of atherogenic dyslipidemia that is typical for this pathology [30]. A strong correlation between hypertriacylglycerolemia plus the HDL-C level decrease and accumulation of LDLB in the blood plasma has been demonstrated in many experiments and clinical studies [18].
It is assumedthat atherogenic alterations occur as a result of lipoprotein disbalance in the blood plasma, i.e. because of predominance of the LDL and VLDL fractions over the antiatherogenic HDL fraction (especially when the values of the LDL+VLDL/HDL index are higher than 3.5).
Age | Group | Parameters | |||
TAG, g/L | Total cholesterol, mmol/L | ApoB-LP, g/L | HDL, g/L | ||
4 weeks | Intact | 1.06±0.07 | 2.93±0.19 | 4.72±0.23 | 1.11±0.05 |
MS | 1.56±0.09* | 3.56±0.10* | 6.68±0.15* | 0.98±0.07 | |
20 weeks | Intact | 1.57±0.22 | 2.84±0.15 | 5.66±0.34 | 1.01±0.02 |
MS | 2.00±0.13* | 3.71±0.18* | 6.68±0.21* | 0.85±0.08 | |
1 year | Intact | 1.50±0.10 | 2.730.02 | 5.21±0.06 | 1.74±0.13 |
MS | 2.27±0.13* | 3.150.08* | 7.00±0.22* | 2.32±0.13* |
Some plasma lipid values in male Syrian golden hamsters with MS (in each group n=10).
As it is known, there are 2 phenotypes of LDL: LDLA and LDLB that differ by size, density, the lipid content and the atherogencity coefficient. LDLB have less size (d 25.5-25.75) comparing to LDLA (d > 25.75) and are characterized by a lower content of polar lipids, as well as a higher content of cholesterol esters. Lipoproteins of this subfraction are slowly removed from the bloodstream that is caused by their low affinity to B/E-receptors for LDL, higher sensitivity to glycosylation and oxidative damage [31]; they also have a high affinity to scavenger-receptors of macrophages [32].
All this features explain a high atherogenicity of LDLB subfraction. Numerous clinical and epidemiological studies have confirmed that accumulation of LDLB in the blood is an independent risk factor for atherosclerosis occurrence [33].
Normally, there are predominantly LDLA in the blood plasma, and LDLB are present in a small percent of the total LDL, but in MS and insulin resistance the LDLB content increases significantly.
It is well-known that in MS the key factor for TAG and ApoB-LP accumulation in the blood is the VLDL hyperproduction by the liver [34]. According to our data, accumulation of ApoB-LP in the blood occurs parallelly with increase in the content of this lipoprotein fraction in the liver (Table 2).
These results allow us to make a suggestion that VLDL formation is activated in the liver of the animals fed with a high-calorie diet in our experiment.
The mechanisms of the VLDL hyperproduction by the liver in the conditions of FFA intensive supply to hepatocytes have remained still unclear. The stimulation of VLDL formation can occur both by using the elevated uptake of the blood FFA and via activation of fatty acid biosynthesis de novo because of hyperglycemia.
Age | Group | Parameters | ||||
Total lipids, mg/g liver | ApoB-LP, mg/g liver | HDL, mg/g liver | G6PDH, nmol/mg protein/min | Lysosomal lipase, nmol/mg protein/min | ||
Week 4 | Intact | 104.24±2.52 | 11.46±0.37 | 1.25±0.14 | 3.74±0.33 | 0.67±0.03 |
MS | 124.16±2.05* | 15.16±0.54* | 1.11±0.07 | 2.80±0.17* | 1.09±0.07* | |
Week 20 | Intact | 112.62±2.66 | 13.03±0.50 | 0.94±0.10 | 4.44±0.28 | 0.54±0.03 |
MS | 143.59±2.65* | 15.69±0.36* | 1.10±0.20 | 3.13±0.28* | 1.27±0.09* |
Some liver lipid metabolism values in male Syrian golden hamsters with MS used in the current study (in the crude tissue, in each group n=10).
It is known that in insulin resistance FFA that come to hepatocytes from the blood are primarily used for the TAG re-synthesis. It leads to increase in the intracellular TAG content and correlates with the increase of the VLDL secretion rate into the bloodstream. The VLDL morphology, which is specified predominantly at the second stage of their formation, depends significantly on the intracellular TAG content and hepatocyte sensitivity to insulin [35]. More active phospholipase D-dependent pre-VLDL lipidation takes place in the elevated intercellular TAG content and insulin resistance of hepatocytes [36]. Insulin blocks the VLDL1 formation in the liver. In the conditions of insulin resistance this effect and the elevated intercellular TAG content stimulate formation and secretion predominantly of VLDL1 by the liver [37].
The VLDL1 secretion increase leads to significant changes in the lipid and lipoprotein metabolism in the blood: the increased TAG content and accumulation of LDLB with high atherogenicity in the blood. These changes are typical for MS and considered to be separate risk factors for development of atherosclerosis.
Metabolism of ApoB-LP in the blood plasma is tightly connected with metabolism of HDL performing a reverse cholesterol transport from peripheral tissues to the liver. The leading factors in the process of transformation of VLDL into LDL in the bloodstream and determination of the LDL morphology are the rate of cholesterol esters transfer from HDL to ApoB-LP mediated by cholesteryl ester transfer protein (CETP), and the rate of TAG hydrolysis in the ApoB-LP composition mediated by lipoprotein lipase (LPL) and hepatic lipase (HL) [38].
According to data of many clinical studies, increase of the CETP activity of the HDL composition in most cases leads to decrease of the HDL-C level and accumulation of LDLB in the blood plasma. Moreover, a degree of these modifications correlates with the blood TAG level.
We observed significant changes in the cholesterol and HDL metabolism in the blood plasma in animals fed with a high-calorie diet. These changes have expressed the proatherogenic character and could be one of the causes for the LDLB accumulation in the blood.
Our results suggest that increase of the total blood cholesterol level in hamsters fed with a high-calorie diet is obviously connected with increase of the cholesterol content in the ApoB-LP composition as its level in the HDL composition decreases (Table 3).
Age (at the beginning of the experiment) | Group | Parameters | |||
HDL-C, mkmol/L | HDL-CE, mkmol/L | LCAT, mkmol/l/h | CETP, mkmol/l/h | ||
Week 4 | Intact | 174.1718.99 | 1028.3312.76 | 54.920.58 | 20.421.76 |
MS | 80.839.17* | 810.0022.78* | 49.002.50 | 33.831.56* | |
Week 20 | Intact | 138.008.00 | 770.0032.56 | 45.502.55 | 59.505.39 |
MS | 164.509.97 | 512.500.01* | 20.252.28* | 116.889.43* |
Plasma HDL-C and HDL-CE, cholesterol esterifying activity and CE transfer in Syrian golden hamsters with the experimental MS (in each group n=10).
Decrease in the HDL cholesterol level is apparently connected with increase of the transfer rate of cholesteryl esters from HDL to ApoB-LP. According to our data the rate of the cholesteryl esters transfer from HDL in the animals fed with a high-calorie diet grows to 166% and 199% compare to the values of young and adult intact animals, respectively (Table 3).
At the same time decrease in the free cholesterol and HDL esterified cholesterol levels was determined in young males, but in adult animals only the HDL esterified cholesterol content lowered. The cholesteryl ester transfer rate from HDL to ApoB-LP is activated when the TAG content increases in the blood, it is observed in the postprandial period, as well as in ApoB-LP metabolism abnormalities [39]. In both cases the cholesteryl ester transfer activation is a consequence of increasing the TAG-rich lipoproteins (TRL) in the bloodstream [40]. The latter is also confirmed by our data pertaining to the increase of the neutral lipids content in the ApoB-containing lipoproteins in hamsters with the experimental MS. These differences seem to be connected with the difference in the HDL free cholesterol esterification rate in males of various ages. This rate is primarily determined by the activity of LCAT – the enzyme associated with HDL [41].
The increase of the cholesteryl-ester transfer activity from HDL is mostly the result of the CETP activation. The increase of the CETP activity in MS was demonstrated in a great number of experiments [22]. It is known that the activation of CETP biosynthesis in the liver is primarily the cause for increasing the activity of this protein in the blood HDL composition, but mechanisms of CETP induction have been still unclear.
Thus, increase of the cholesteryl ester transfer rate from HDL on the background of hypertriacylglycerolemia, which is observed in our experiment in the animals fed with a high-calorie diet (Table 3), is atherogenic since the cholesteryl ester transfer predominantly to TAG-enriched lipoproteins leads to accumulation of CE-enriched VLDL1, which are major precursors of LDLB. Intensive TAG supply to HDL in exchange for cholesteryl esters results in accumulation of TAG-enriched HDL particles, which are the predominant substrate for hepatic lipase (HL), in the blood. So, HDL particles are rapidly removed from the bloodstream and it leads to decrease of the HDL-C content.
That is why changes in the enzymes activity, which hydrolyze lipoprotein lipids in the bloodstream, in particular – in LPL and HL activity, affect significantly the lipoprotein metabolism in MS.
TAG in the TAG-enriched lipoproteins (chylomicrons and VLDL) are the substrate for LPL. FFA, released after hydrolysis under the action of LPL, come to adipocytes and muscle cells where they are deposited as the TAG component or used as a source of energy. TAG hydrolysis in the VLDL composition increases availability of cholesterol for its transfer to HDL, therefore, in this way LPL mediates the reverse cholesterol transfer. The LPL activity is regulated by the influence on transcription, translation and enzyme transport from the cells. Insulin is known to activate LPL that results in decrease of the total blood TAG level and stimulation of cholesterol reverse transfer [35].
According to our data, the plasma LPL activity decreased in young male hamsters fed with a high-calorie diet (Table 4).
Age (at the beginning of the experiment) | Group | Parameters | |
LPL (U/ml) | HL (U/ml) | ||
Week 4 | Intact | 8±2 | 51±4 |
MS | 4±1* | 91±3* | |
Week20 | Intact | 83±2 | 3±1 |
MS | 129±3* | 2±1 |
Postheparin plasma lipase activities in Syrian golden hamsters with the experimental MS (in each group n=10).
The results obtained are in agreement with the literature data about the reduction of the LPL activity in obesity and insulin resistance [42]. The mechanisms of the LPL activity inhibition in these conditions are still unclear though a definite contribution could be made by development of insulin resistance.
The increase of the cholesteryl ester transfer rate from HDL on the background of hypertriacylglycerolemia, which was stated in our experiment both in animals fed with a high-calorie diet and in chronic stress, is an atherogenic factor for two reasons. Firstly, the cholesteryl ester transfer predominantly to the TAG-enriched lipoprotein fractions leads to accumulation of VLDL1 enriched with cholesteryl esters, which are the main LDLB precursors. Secondly, the intensive exchange of cholesteryl esters in HDL for TAGs results in accumulation of TAG-enriched HDL in the blood, which are the predominant substrates for HL, and they are rapidly removed from the bloodstream, and it, in turn, causes decrease in the HDL-C concentration. The activation of the lipoprotein secretion by the liver is also observed in the conditions of the acute chemical and emotional painful stress. This fact may be considered to be a sign of proatherogenesis since it is accompanied by hyperlipidemia development due to increase of atherogenic lipoprotein fractions.
As shown in our studies, decrease of the LPL activity in the blood plasma of young males fed with a high-calorie diet can be an additional factor for TAG accumulation in the blood and the HDL-C level reduction observed in our experiment.
HL mediates a selective transport of VLDL remnants to hepatocytes via LDL-receptors, takes part in reverse transport of cholesterol accelerating HDL coming into the liver via scavenger receptors (SRB1). Hydrolyzing TAG in the ApoB-LP composition HL plays a significant role in their re-modelling in the bloodstream. It is known that the HL activity specifies substantially the lipid composition, size and properties of LDL [43].
The HL activity is predominantly regulated at the transcriptional level under the influence of sex hormones, glucocorticoids and adipokines. The rate of the HL gene transcription is also dependent on the intercellular lipid content, primarily cholesterol in the hepatocytes [44].
In our experiment the blood plasma HL activity in male hamster fed with a high-calorie diet increased irrespective of age (Table 4), it corresponds to literature data. In a number of studies it has been shown that the HL activity increases in insulin resistance, obesity, and a high-calorie diet [45]. Moreover, it has been determined that increase of the HL mRNA content is observed when using a high-calorie diet; this is the evidence of the enzyme biosynthesis activation under these conditions. The authors associate this fact with the decrease in the blood plasma adiponectin level, which can inhibit the HL synthesis in hepatocytes.
Considering these data, as well as the data obtained in our studies about adiponectin decrease in the blood plasma in obesity (Fig. 1), we may suppose that one of the causes for the HL activity increasing when taking a high-calorie diet in our experiment is decrease of adiponectin secretion by the adipose tissue.
The HL activity increase is considered to be one of the key factors for the atherogenic dyslipidemia development in obesity and MS. In a number of works a clear correlation between the HL activity and the LDLB content in the blood plasma was demonstrated [19]. It is believed that namely HL activation results in the increased LDLB formation [33]. The latter occurs with increase of the TAG-enriched VLDL1 content in the blood and the CETP activation. Furthermore, the HL activity increase leads to decrease of the HDL cholesterol level [46]. This is associated with the fact that hydrolysis of TAG in the HDL3 composition results in their transformation into HDL2, which are rapidly removed from the bloodstream by the liver. Thus, the HDL-C level decrease that we determined in our experiment (Table 3) can be a consequence of the HL activity increase.
In the current study we have found that the blood FFA level increase is accompanied by the ApoB-LP synthesis activation in the liver of Syrian male hamsters fed with a high-calorie diet irrespective of age. This causes increase of the TAG and ApoB-LP level in the blood. Decrease of the HDL-C level is a consequence of the rate of cholesteryl ester exchange between HDL and LDL due to activation of CETP and HL. As a result of these changes the atherogenic dyslipidemia development, which is typical for MS, is observed.
Plasma adiponectine level in Syrian golden hamsters with the experimental MS development (values are mean±SD; * –р0.05 versus intact animals, in each groupn=10, * – p < 0.05 versus intact animals).
We have found the age differences of the lipid profile in the blood plasma of the normal male hamsters. So, in intact males (with age from 4 to 20 weeks) on the background of the constant content of total lipids and lipoproteins in the blood plasma there was increase of the FFA level (60% comparing to a 4-week intact), TAG (48%) and ApoB-LP (20%), and the HDL level showed a tendency to decrease. These results are the evidence of the lipidation increase with age. It has been also shown that in adult males the unesterified cholesterol and cholesteryl ester levels are lower than in young animals (20% and 25%, respectively), and the cholesteryl ester transfer rate from HDL in adult animals exceeds this index value in young animals (191%) (Table 3).
The data obtained correspond to the literature data about age-dependent changes in the lipid metabolism in males, which have the proatherogenic character [47]. It is known that with age the sex hormones level lowers in males and the glucocorticoid secretion level increases. The plasma lipid profile in males is determined, among other factors, by the secretion level of sex hormones possessing antiatherogenic properties. A lot of studies proved the presence of direct correlation between the blood testosterone plus the dehydrotestosterone level and the HDL-C content [48]. Moreover, the high level of sex hormones correlates with decrease of the TAG content and the total cholesterol in the blood. Thus, increase of the TAG level and decrease of the HDL cholesterol content in the blood plasma of males with age may be connected with reduction of the sex hormone secretion (Table 5). Changes of the lipid profile in the blood plasma of males with age may be also associated with increase of glucocorticoid secretion, which was observed in our experiment (Table 5).
Thus, with age the blood plasma lipid profile in males is subjected to unfavourable changes such as increase of the FFA and TAG content and decrease of HDL-C. The latter may be connected with decrease of the sex hormone level, and increase of the cortisol secretion. However, despite the more favourable lipid profile in the blood plasma of young males comparing with normal adult animals, atherogenic dyslipidemia in obesity and insulin resistance develops irrespective of age.
In contrast with males, in females the atherogenic dyslipidemia development is significantly dependent on age (Table 6). In particular, while in males with age there are no significant changes in the liver ApoB-LP content, in females this index raises during maturation – in intact animals – to 20%, and in animals in the experimental MS – to 31%. That indicates intensification of lipolytic processes in the liver of females during ageing, and may serve as a manifestation of the lipid metabolism activation. The analogous changes in the total lipid content also proved this tendency (Table 6).
Sex | Parameter | Group | Age (at the beginning of the experiment) | ||
4 weeks | 20 weeks | 1 year | |||
Females | Estradiol, pmol/mL | Intact | 0.550.05 | 0.640.06 | 0.540.05 |
MS | 0.630.06* | 0.750.08 | 0.360.04* | ||
Cortisol, nmol/L | Intact | 47.003.85 | 73.175.56 | 76.00±4.95 | |
MS | 74.05.49* | 87.54.45 | 117.00±2.63* | ||
Males | Estradiol, pmol/mL | Intact | 0.240.02 | 0.190.02 | 0.290.03 |
MS | 0.270.02 | 0.290.02* | 0.200.03* | ||
Testosterone, pmol/mL | Intact | 4.020.39 | 4.240.31 | 3.580.37 | |
MS | 4.450.41* | 3.510.40* | 3.030.31* | ||
Cortisol, nmol/L | Intact | 61.173.71 | 94.83.06 | 85.33±5.40 | |
MS | 84.673.62* | 132.007.88* | 148.40±9.54* |
Plasma sex hormones and cortisol levels in hamsters with the experimental MS (in each group n=16)
Age | Group | Parameters | |||
Total lipids, mg/g | ApoB-LP, mg/g liver | HDL, mg/g | Lysosomal lipase, nmol/mg protein/min | ||
4 weeks | Intact | 117.674.72 | 8.870.24 | 1.270.08 | 0.340.03 |
MS | 144.345.00* | 10.240.25* | 0.650.05* | 1.240.05* | |
10 weeks | Intact | 137.543.91 | 10.650.46 | 0.890.07 | 0.830.04 |
MS | 179.223.44* | 13.440.30* | 0.460.06* | 1.330.08* |
Lipid metabolism parameters in the liver homogenate in Syrian golden female hamsters with MS(in the crude tissue, in each group n=16)
Oxidation of LDL and VLDL (i.e. ApoB-LP) is an alternative way of the lipoprotein catabolism, which leads to their uptake by macrophages via scavenger-receptors, and may lead to the transformation of these cells into “foam” ones. That is why it is one of the factors of atherogenesis in MS.
In our experiment we also observed the composition changes in lipoproteins and in particular HDL particle enrichment with lipids (Table 7). However, the cholesterol content of these lipoproteins decreased in contrast to the ApoB-LP cholesterol content that was increased.
Parameters | Group | |
Intact | MS | |
Total lipids, % of the total HDL composition | 49.451.35 | 57.311.91* |
Total cholesterol, % of the total HDL composition | 14.970.23 | 11.210.76* |
TAG, % % of the total HDL composition | 1.750.07 | 3.080.15* |
-Tocopherol, mmol/L | 8.020.39 | 5.700.35* |
Isolated double bonds, U/ml | 8.640.59 | 7.310.17* |
Diene conjugates, mmol/L | 18.882.10 | 31.681.65* |
Ketodienes+conjugated trienes, U/ml | 1.150.08 | 1.480.06* |
Total hydroperoxides, mmol/L | 69.043.46 | 78.311.33* |
The plasma HDL composition in Syrian golden hamsters (1 year) with the experimental MS(in each group n=10).
There are several possible reasons for that phenomenon. One of them is a well-known fact that HDL contains high levels of both unsaturated fatty acids, which are rapidly utilized, and proteins, which hydrophilic properties compensate the lack of phospholipids, as well as α-tocopherol and enzymatic antioxidants, particularly paraoxonase, which protect these lipoproteins from peroxidation. There is no doubt that the changes in the cholesterol metabolism enzymes activity associated with HDL (CETP) are involved in this process (Table 3).
Nevertheless, the content decrease of compounds with isolated double bonds and accumulation of the lipoperoxidation products has been determined in the HDL fraction in MS (Table 7). Moreover, the data obtained have shown that the content of ketodienes and coupled trienes in the HDL fraction is 129% comparing to control; the content of diene conjugates – 168% and the content of the total hydroperoxides – 115%. It has been also found that there is decrease of the α-tocopherol content in HDL (41%) comparing to the control values (Table 7).
Thus, HDL can protect LDL from oxidation “providing” a cell with paraoxonase and PAF-acetyl hydrolase. However, this protective effect of HDL is reduced in response to induction of the stress acute phase in animal models [49].
As can be seen from our data (Table 8), the HDL-associated paraoxonase activity is generally decreased in experimental MS.
Sex | Age | Groups | Activity, nmol/mL/min |
Males | 4 week | Intact | 80.783.69 |
MS | 67.063.70* | ||
20 week | Intact | 62.192.63 | |
MS | 37.293.33* | ||
Females | 4 week | Intact | 104.412.95 |
MS | 75.452.21* | ||
20 week | Intact | 127.272.95 | |
MS | 121.933.05 |
The plasma HDL paraoxonase activity in Syrian golden hamsters with experimental MS(in each group n=10).
The data obtained show that application of antioxidant complexes for correction of unfavourable changes in proatherogenic states may be perspective since free radical oxidation activation is a common pathogenetic link of all those states; this link is not only involved in damage of cells and their components, but also as an alternative way of catabolism it accelerates the lipid recyclization.
At the same time, since a significant feature of proatherogenic states is the hormone status imbalance, polyphenolic antioxidants need special attention because these compounds along with the antioxidative activity also demonstrate phytoestrogen properties [50], and it may be an additional factor of the lipid metabolism regulation.
An important effect of flavonoids is scavenging of oxygen-derived free radicals. The experimental systems in vitro have also shown that flavonoids possess anti-inflammatory, antiallergic, antiviral, and anticarcinogenic properties. The so-called ‘‘Mediterranean diet’’ is thought to prevent cardiovascular diseases, as a consequence of its high content of antioxidants, which are crucial in ameliorating oxidative events implicated in many diseases. In addition to the antioxidant/antiradical activity, red wine polyphenols (RWPs) have been shown to possess many biological properties, including inhibition of platelet aggregation, the vasorelaxing activity, modulation of the lipid metabolism, and inhibition of the low-density lipoprotein oxidation.
In our research we have used wine, juice and polyphenolic extracts from grapes of different grades, and polyphenolic concentrates “Enoant” and “Polyphen” obtained from Vitis Vinifera grapes to correct the changes in the lipid metabolism in the conditions of the experimental metabolic syndrome, acute and chronical stress. All substances used in our research were developed in National Institute for Vine and Wine "Magarach" (Yalta, Ukraine).The studies carried out have specified that polyphenolic extracts and concentrates are quite active remedies that decrease negative effects in MS though the effectiveness of various substances administered are significantly different.
So, administration of any of the investigated substances has significantly decreased the total blood plasma lipoprotein content in hamsters with MS, but the use of “Cabernet” extract has the most pronounced effect (Fig. 2). The same tendency is observed in decreasing the ApoB-LP content, the total cholesterol and FFA level have also decreased, though practically no difference between the grape varieties investigated has found.
The effect of Vitis Vinifera substances on some plasma lipid metabolism values in male Syrian golden hamsters (1 year old) with the experimental MS (in each groupn= 7)
The non-enzyme antioxidant level (α-tocopherol, reduced glutathione and ascorbic acid) in the blood serum has also reached reference values under the influence of the polyphenolic extracts.This fact confirms the high antioxidant activity of the studied substances.
Normalization of the blood plasma phospholipid content under theinfluence of polyphenolic extracts arouses the interest. The phospholipid content returned to the intact level, which may be a result of their oxidation reduction, given that the unsaturated fatty acids in phospholipids are compounds that undergo oxidation by free radicals quickly and easily.
However, in spite of the quite favourable effect of “Isabella” extract, its administration also has negative consequences, particularly the HDL level decrease to the value observed in intact animals accompanied by the LDL content increase.
The investigated substances normalize also the blood lipoproteins composition. Thus, the total lipid and the total cholesterol content decrease in the ApoB-LP composition, moreover “Enoant” lowers the cholesterol content in this atherogenic lipoprotein fraction even below the control level.
Generally, the TAG content is also normalized under the action of all the investigated substances, but taking into account the ratio – cholesterol/triacylglycerols, “Polyphen” has the most favourable effect.
The polyphenol extracts and concentrates have significantly improved the ApoB-LP oxidative status in animals with MS. The best results have been obtained when using “Cabernet”, as well as for other indexes investigated (Table 9).
Parameter | Group | ||||
MS | MS +“Enoant” | MS +“Polyphen” | MS+“Isabella” | MS+“Cabernet” | |
Total lipids, % of the total ApoB-LP composition | 88.87 0.71* | 83.70 0.78 */** | 83.12 0.37*/** | 82.22 0.09*/** | 81.00 0.19* |
Total cholesterol, % of the total ApoB-LP composition | 8.39 0.24 | 7.87 0.04*/** | 8.13 0.04 | 8.06 0.12** | 8.17 0.08*/** |
TAG, % of the total ApoB-LP composition | 37.55 1.89*/** | 53.97 0.10*/** | 53.02 0.14*/** | 50.65 1.23*/** | 49.57 0.40*/** |
- Tocopherol, mmol/L | 2.68 0.08* | 2.98 0.05*/** | 3.06 0,04*/** | 3.17 0,02*/** | 3.19 0,05*/** |
Isolated double bonds, U/ml | 1.71 0.06* | 1.84 0.03*/** | 1.90 0.02*/** | 1.97 0.03** | 2.09 0.03** |
Diene conjugates, mmol/L | 37.25 1.50* | 30.63 0.41*/** | 29.54 0.34*/** | 28.77 0.14*/** | 26.68 1.94** |
Ketodienes+conjugated trienes, U/ml | 8.18 0.11* | 7.49 0.04*/** | 7.23 0.08*/** | 7.17 0.08*/** | 6.99 0.26** |
Total hydroperoxides, mmol/L | 108.25 1.39* | 98.52 0.55*/** | 94.97 0.15*/** | 90.65 1.15*/** | 89.30 1.06*/** |
The effect of Vitis Vinifera substances on the plasmaApoB-LP composition in male Syrian golden hamsters (1 year old) with the experimental MS (in each group n= 10)
The HDL composition in the blood is also affected by thesubstances studied. In these particles the total lipid content decreases and even reaches the level of intact animals when using “Cabernet” extract (Table 10).
The cholesterol level also changes: it decreases when using “Enoant” and increases under the action of “Isabella” and “Cabernet” extracts.
The HDL-C content decreases under the action of “Enoant” may occur due to peroxide processes inhibition since cholesterol accumulation in lipoprotein particles, as it was mentioned before, has a compensatory character in response to the phospholipid oxidation of the lipoprotein particle hydrophilic cover. The TAG content decreased under the action of all substances, and “Isabella” was the most effective substance. The TAG content decrease is probably mediated by the phytoestrogenic action of polyphenols directed to lipolysis inhibition in the adipose tissue.
Parameter | Group | ||||
MS | MS +“Enoant” | MS +“Polyphen” | MS+“Isabella” | MS+“Cabernet ” | |
Total lipids, % of the total ApoB-LP composition | 57.31 1.91* | 54.91 0.21*/** | 53.09 0.08*/** | 51.74 0.74*/** | 49.20 0.42** |
Total cholesterol, % of the total ApoB-LP composition | 11.21 0.76* | 10.54 0.30*/** | 11.14 0.04* | 12.05 0.21* | 12.45 0.34*/** |
TAG, % of the total ApoB-LP composition | 3.08 0.15* | 2.90 0.09*/** | 2.11 +0.12*/** | 1.92 0.04*/** | 1.96 0.03*/** |
- Tocopherol, mmol/L | 5.70 0.35* | 7.47 0.20*/** | 7.19 0.17*/** | 7.36 0.11*/** | 8.13 0.06** |
Isolated double bonds, U/ml | 7.31 0.17* | 7.67 0.08*/** | 7.69 0.07*/** | 7.99 0.05*/** | 8.15 0.01*/** |
Diene conjugates, mmol/L | 31.68 1.65* | 24.85 0.35*/** | 23.44 0.40*/** | 22.55 0.34*/** | 21.88 0.23** |
Ketodienes+conjugated trienes, U/ml | 1.48 0.06* | 1.24 0.03** | 1.32 0.03*/** | 1.25 0.03*/** | 1.54 0.47*/** |
Total hydroperoxides, mmol/L | 78.31 1.33* | 75.26 0.31*/** | 75.62 0.54*/** | 74.48 0.55*/** | 73.41 0.39*/** |
The effect of Vitis Vinifera substances on the plasmaHDL composition in male Syrian golden hamsters (1 year old) with the experimental MS (in each group n= 10)
The exact bimolecular mechanisms for this cardioprotection are unclear, but it is likely that actions mediated both through the estrogen receptors, such as the beneficial alteration in lipid profiles and upregulation of the low-density lipoprotein (LDL) receptor, and independently of the estrogen receptors, such as antioxidant action, contribute to the cardioprotective effects of phytoestrogens observed.
The potential role of phytoestrogens, including isoflavonoids, as cardioprotective agents has been extensively reviewed. The data obtained in our experiments showed that in male hamsters with the experimental MS the treatment with grape extracts reduced VLDL cholesterol (VLDL-C) and TG by 30 and 40 % compared with the control animals. Furthermore, golden Syrian hamsters fed with red wine phenolics had a significant decrease in the plasma apo B concentrations. Similar to our previous study, grape polyphenols may have altered hepatic secretion of TG-rich VLDL. This reduction is evident when observing the decreases in both plasma apo B and apo E concentrations. The significant decrease in apo E concentrations may have further reduced plasma TG concentrations. In general, apo E displaces apo C-II from the VLDL particle, thereby inhibiting the lipoprotein lipase (LPL) activity and overall lipolysis. Furthermore, Huang et al. [51] showed that adding apo C-II to transgenic apo-E3–enriched VLDL increased the LPL activity in a dose-dependent manner. The reductions in apo E and TG concentrations suggest less displacement by apo E, thereby promoting the grape polyphenols activity and further reducing the TG concentrations in the plasma.
Due to decreases in TG concentrations, administration of “Cabernet” extract was shown to affect the overall lipoprotein metabolism. Decreased concentrations of the plasma TG altered substrate availability in the delipidation cascade, leading to the decrease observed in LDL-C concentrations. After a 3-week treatment period the grape polyphenols treatment induced a significant decrease in the cholesteryl ester transfer protein (CETP) activity as well. Such decrease in the CETP activity may be partially a result of the substantial decrease in substrate availability, including both the plasma TG and LDL-C.
It is evident that grape polyphenols modify the packaging of VLDL through alteration in the hepatic enzyme activity and apo B secretion. These modifications seem to decrease the overall secretion of the VLDL particles and therefore, decrease plasma TG and related apo concentrations. Due to decrease of the TG substrate, further modifications in the lipoprotein metabolism may occur.
The alteration in the TG metabolism may not be the single mechanism driving the hypocholesterolemic effects of grape polyphenols. When golden Syrian hamsters were treated with dealcoholized red wine, red wine, or grape juice, similar significant reductions in both TC and LDL-C concentrations were apparent in all treatment groups compared with the control [51]. Although there was a trend for decrease in TG concentrations in all treatment groups compared with the control, the differences were not significant. That study, along with others, suggests the presence of an additional mechanism by which grape polyphenols exert the cardioprotective effect. In Hep G-2 cells, dealcoholized red wine was shown to upregulate significantly the LDL receptor activity. This significant increase in activity was similar to the increase seen when Hep G-2 cells were treated with atorvastatin. Furthermore, when Hep G-2 cells were treated with increasing doses of red wine, LDL receptor mRNA abundance was significantly increased in a dose-responsive manner. The increase of the LDL receptor activity and abundance may be a result of the homeostatic intracellular cholesterol feedback loop. In general, decrease in the intracellular cholesterol will upregulate the LDL receptor expression and activity, whereas increase in the intracellular cholesterol will downregulate the receptor [48]. Grape polyphenols were shown to decrease hepatic cholesterol concentrations; therefore, the liver compensates for this deficiency by upregulating the LDL receptor and the overall decrease in the plasma LDL concentrations occurs.
One possible explanation of the anti-atherogenic activity of grape polyphenols is the well-known HDL cholesterol-increasing effect of polyphenols in various species, including transgenic mice [52].
In our experiments it has been found that the grape extract treatment induced slight (15%) increase in HDL cholesterol concentrations is possibly related to the significant decrease in the hepatic lipase activity (Table 11). The reductions observed in both hepatic and LPL activities by grape polyphenols treatment may prevent formation of small atherogenic VLDLB particles and may also decrease their uptake by the LDL receptor -related protein.
In addition to increases in HDL cholesterol concentrations, grape extracts also change the size and quality of HDL particles [53]. Although the mechanisms by which polyphenols influence the metabolism of HDL particles are not clear, changes in LPL and cholesteryl ester transfer protein (CETP) may play an important role.
Polyphenols treatment in humans is associated with decrease in the CETP content correlated with the concomitant increase in HDL cholesterol concentrations [54]. Consistent with our findings, grape extracts caused a significant increase in the postheparin LPL activity and HDL cholesterol concentrations in patients with moderate hypercholesterolemia and in hamsters [39]. However, the HDL cholesterol-increasing action of polyphenols in animals (mouse, hamster and rat) without CETP in some cases [52] suggests that this effect is may be independent of the CETP activity.
milliunits | Control (MS) (n=50) | Grape extract “Cabernet” (n=50) |
LPL | 356.0±53.2 | 258.6±57.3* |
Hepatic lipase | 232.6±25.9 | 216.2±34.7 |
The plasma postheparin lipases activityin male hamsters with MS (in each group n=10)
The “Cabernet” extract appeared to be the most effective substance in relation to the HDL defence from peroxidation, though the other substances revealed the same but not so high activity. They decreased the content of products (diene conjugates, ketodienes+coupled trienes, total hydroperoxides) effectively and increased – substrates (compounds with isolated double bonds) of lipoperoxidation, prevented decrease of the antioxidant level (α-tocopherol).
It should be pointed out that the level of lipid peroxidation secondary products (ketodienes+coupled trienes) decreased more effectively under the influence of “Enoant” (to intact values).
Under the action of the studied substancesthe lipoprotein supply to the liver also decreases, evidenced by the decrease of the ApoB-LP content in the organ. Moreover, in the composition of these lipoproteins the TAG content normalizes, and it indicates normalization of the activity of lipases catalyzing the lipoprotein metabolism in the blood (Table 12).
The liver oxidative status is also improved: the antioxidant levels almost restore, the peroxidation products content decreases, the content of compounds with isolated double bonds increases (Tables 3, 5, 9, 12).
Testing of the “Enoant” action – one of the substances studied– in female hamsters of different age with the experimental MS proved the effectiveness of the antioxidant therapy of this pathology.
So, the total lipids, TAG and FFA contents decrease in the blood plasma of those animals under the action of “Enoant” (Table 12). In addition, in adult females “Enoant” causes decrease in the ApoB-LP and total cholesterol content, and it, in turn, reduces atherogenic changes in MS.
Parameter | Group | ||||
MS | MS +“Enoant” | MS + “Polyphen ” | MS+“Isabella” | MS+“Cabernet” | |
Total cholesterol, % of the total ApoB-LP composition. | 7.180.06* | 8.230.26*/** | 8.460.05** | 8.760.05** | 9.100.13** |
TAG, % of the total ApoB-LP composition | 42.001.29* | 44.640.52** | 44.420.43** | 45.950.50** | 45.410.73** |
Isolated double bonds, U/g | 2.130.06* | 2.390.04*/** | 2.710.03*/** | 2.990.09** | 2.770.16** |
Total hydroperoxides, mmol/g | 101.032.00* | 90.551.54*/** | 88.691.02*/** | 80.460.77*/** | 78.832.71*/** |
The effect of Vitis Vinifera substances on the liver cytosolApoB-LP composition in male Syrian golden hamsters (1 year old) with the experimental MS (in the crude tissue,in each group n= 10)
The increase of -tocopherol (the main lipid-phase antioxidant) in the blood plasma of animals that received “Enoant” proved its antioxidant activity in our experiment (Table 13).
Furthermore, the significant decrease of the body weight was observed in hamsters that received “Enoant” along with a high-calorie diet compared to the animals on a high-calorie diet alone.
Based on these findings, we may conclude that introduction of grape polyphenolic extracts and concentrates in MS can prevent the increase of the total lipid and ApoB-LP content in the blood plasma, prevent the activation of free radical processes in the plasma lipoprotein particles, and normalize the liver lipid metabolism. The ability of the investigated substances to reduce negative consequences of MS such as atherosclerosis development has been proven.
The last suggestion is confirmed by our results concerning the aorta wall lipid composition in the experimental MS. The introduction of “Enoant” for prophylaxis and treatment reduces significantly atherogenesis manifestations in the aorta, decreasing the aorta media lipidation and the neutral lipid content (Fig. 3, 4).
Thus, from our data, we can conclude that antioxidants, particularly grape polyphenolic concentrates and extracts, which have pronounced antioxidant, phytoestrogenic and stress-protector properties, should be included into a complex therapy of MS to reduce its negative effects.
The next experiment was designed to investigate the action of grape wines and polyphenolic concentrates on development of proatherogenic effects of the emotional-painful stress. In our experiments we used purebred female rats because, as it was shown in previous studies, the acute stress response in females was more expressive than in males.
Age | Group | Parameter | ||||||
Total lipids, mg/ml | ApoB-LP, mg/ml | Total cholesterol, mmol/L | TAG, mg/ml | FFA, mmol/L | Diene conjugates in ApoB-LP, nmol/ml | - Tocopherol, nmol/ml | ||
4 weeks | MS | 4.520.17 | 4.000.16 | 2.040.08 | 1.080.49 | 1.170.06 | 24.821.46 | 6.670.22 |
MS+“Enoant” | 3.540.16** | 3.470.13 | 1.880.06 | 0.910.02** | 0.950.02** | 22.270.99 | 9.790.77** | |
20 weeks | MS | 7.750.20 | 3.840.11 | 2.580.07 | 1.400.04 | 1.420.04 | 23.581.35 | 10.490.82 |
MS+“Enoant” | 6.880.14** | 3.300.08** | 2.220.05** | 1.180.03** | 1.150.03** | 21.101.14 | 12.870.36** |
The effect of polyphenol concentrate “Enoant” on some plasma lipid metabolism values in female Syrian golden hamsters with the experimental MS (in each group n= 10)
Age | Group | Parameters | |||
Total lipids, mg/g | - Tocopherol, nmol/g | Ascorbic acid, mkmol/g | TBA active substances, nmol/g | ||
4 weeks | MS | 140.75 9,15 | 21.011.47 | 3.730.14 | 1.970.06 |
MS+“Enoant” | 111.534.08** | 25.310.34** | 4.080.10 | 1.740.09 | |
20 weeks | MS | 154.182.70 | 19.590.39 | 6.100.35 | 1.950.09 |
MS+“Enoant” | 121.044.18** | 26.111.03** | 5.960.24 | 1.730.09 |
The effect of polyphenol concentrate “Enoant” on some liver lipid metabolism values in female Syrian golden hamsters with the experimental MS (in each group n= 10)
During 21 days animals were daily given per os grape wines of “Cabernet” and “Rkatsiteli” grades in the doses that corresponded to 300 ml of wine for a human of 70 kg. Other animals were given alcohol in the dose corresponding to 30 ml of alcohol for a human of 70 kg, as well as polyphenolic concentrates “Enoant” and “Polyphen” in the doses of 0.05 ml/kg of the body weight. The grape wines and polyphenolic concentrates were produced by the National Institute of Grape and Wine “Magarach”. Control animals were given the corresponding volume of the physiological solution.
The lipid content in the aorta wall in male Syrian golden hamsters with the experimental MS and “Enoant” treatment (Mm, in each group n=10).
The lipid content in the aorta wall in female Syrian golden hamsters with the experimental MS and “Enoant” treatment (Mm, in each group n=10).
It was shown that all the substances investigated: polyphenolic concentrates “Enoant” and “Polyphen”, 10% solution of ethanol, and grape wines “Cabernet” and “Rkatsiteli” possessed the stress-protective activity, which intensity was dependent on the substance used (Tables 15-17).
When introducing only “Enoant” and “Polyphen” to the animals these complexes did not cause any changes on the investigated indexes of the pro-oxidant and antioxidant status in the liver and it is an indication about safety of using these concentrates.
“Enoant” and “Polyphen” revealed the significant protective activity in the emotional-painful stress. It allows to use them as stress-protective, hepatoprotective and antiatherogenic remedies.
Parameter | Group | |||||||||
Stress + Enoant | Stress + Polyphen | Stress + ethanol | Stress + Cabernet | Stress + Rkatsiteli | ||||||
str. | non-str. | str. | non-str. | str. | non-str. | str. | non-str. | str. | non-str. | |
TL, mg/ml | 4.14 0.45* | 3.98 0.31 ** | 3.81 0.03*/** | 3.82 0.48 ** | 5.50 0.74*/** | 5.58 0.37* | 3.41 0.66 ** | 3.46 0.43 ** | 3.43 0.15 ** | 3.57 0.54 ** |
TAG, mg/ml | 0.76 0.08*/** | 0.78 0.09* | 0.51 0.08 | 0.71 0.09* | 0.66 0.12*/** | 0.91 0.05*/** | 0.50 0.05 | 0.58 0.04*/** | 0.54 0.10 | 0.54 0.05 ** |
Total cholesterol, mg/ml | 70.76 9.34* | 54.90 6.92*/** | 74.14 8.21* | 82.93 9.48 ** | 85.71 5.71 | 69.10 7.37 ** | 56.71 7.32* | 62.25 6.50 ** | 97.51 9.66 ** | 80.83 8.74 ** |
HDL, mg/ml | 0.98 0.14 | 0.89 0.08 | 0.96 0.05 | 0.88 0.08 | 1.10 0.19 | 1.00 0.12 | 1.26 0.15 | 1.04 0.08 | 1.08 0.12 | 0.96 0.05 |
АpоВ-LP, mg/ml | 1.56 0.12 ** | 1.89 0.26* | 1.59 0.16 ** | 1.54 0.22 ** | 1.85 0.19 ** | 1.67 0.18 ** | 2.07 0.29* | 1.67 0.18 ** | 1.86 0.20 ** | 1.63 0.18 ** |
Corticosterone, nmol/l | 5.50 0.99*/** | 5.87 1.08*/** | 7.10 0.80*/** | 29.33 8.58 | 15.00 1.72*/** | 38.75 5.64 ** | 32.75 5.17 ** | 26.33 4.40*/** | 16.93 2.43*/** | 40.60 6.38 ** |
The effect of grape polyphenol complexes and grape wines on the lipid metabolism and the plasma corticosterone level in rats with the neurogenic stress (in each group n=10).
Moreover, we have found out that the stress-protective activity of grape wines is equal to the polyphenolic concentrates activity given in the similar dose.
Wines of “Cabernet” and “Rkatsiteli” grades normalized the total lipid content both in a liver homogenate and in the blood plasma in stress; in addition, TAG levels also reached the control values.
Grape wine components prevented the FFA content increase noted when introducing the solution of alcohol. This fact may prove the protective action of the components mediated by inhibition of fatty infiltration of organs.
The latter is confirmed by the absence of influence of grape wine introduction on the NADPH-generating dehydrogenases activity in the liver.
The cholesterol content decrease in the blood plasma when introducing grape wines has attracted our attention, as well as a favourable redistribution of cholesterol in the LP fractions – decrease of ApoB-containing lipoprotein level with the unchanged HDL content.
Parameter | Group | ||||||||||
Stress + Enoant | Stress + Polyphen | Stress + ethanol | Stress + Cabernet | Stress + Rkatsiteli | |||||||
str. | non-str. | str. | non-str. | str. | non-str. | str. | non-str. | str. | non-str. | ||
PON, nmol/ml/min | 240.25 20.86*/** | 216.00 16.24*/** | 231.33 15.53*/** | 231.25 15.52** | 197.5014.37*/** | 150.00 19.24*/** | 239.2524.18*/** | 246.00 16.56** | 222.7513.98** | 215.25 13.78** | |
Ascorbic acid, mkmol/ml | 49.99 4.41* | 43.20 4.16* | 53.76 4.51 ** | 45.08 6.08*/** | 38.04 4.12*/** | 34.83 4.43* | 48.07 2.13* | 48.10 2.49*/** | 55.87 6.65 ** | 53.41 5.42*/** | |
-Tocopherol, nmol/ml | 10.70 1.34 ** | 10.62 0.67 ** | 10.39 1.12 ** | 10.8 1.26 ** | 6.07 0.76*/** | 5.35 0.78*/** | 9.45 1.11 | 9.99 0.48 ** | 10.26 0.48 ** | 9.32 1.04 | |
Oxidized АpоВ-LP | Isolated double bonds, U/ml | 2.76 0.36* | 2.23 0.24* | 3.14 0.46 ** | 3.51 0.42*/** | 3.13 0.37 ** | 2.35 0.24* | 3.25 0.38 ** | 2.64 0.28* | 4.13 0.43 ** | 3.11 0.41* |
Diene conjugates, mmol/ml | 22.47 3.20*/** | 25.93 2.98*/** | 21.54 1.22*/** | 25.93 1.76*/** | 30.44 2.20* | 26.28 1.24* | 23.05 4.48* | 26.99 1.38* | 19.54 0.93 ** | 21.56 0.99*/** | |
Ketodienes+conjugated trienes, U/ml | 2.31 0.14 | 2.58 0.29*/** | 2.20 0.11 ** | 2.37 0.10* | 3.01 0.50* | 2.97 0.42* | 2.29 0.16 | 2.60 0.31* | 2.22 0.11 | 2.23 0.06 ** |
The effect of grape polyphenol complexes and grape wines on the plasma oxidant/antioxidant status in rats with the neurogenic stress (in each group n=10).
Paraoxonase activity was normalized in the animals given wines and the antioxidant content both in the blood plasma and in the liver was significantly higher than in the control animals.
These effects together with much lower level of ApoB-LP oxidation in the animals given grape wines prove the high antiatherogenic potential of the wines investigated.
In addition, the grape wines have revealed a rather high level of the stress-protective activity and it is indicated by a significant decrease of the corticosterone content in the blood plasma in stressed animals given wines.
Since grape wines have shown a high level of the stress-protective activity we investigated how the ratio of wine components – polyphenols and ethanol – can influence the stress-protective activity of this complex.
As was shown in our experiments “Enoant” administration even in the combination with ethanol does not reduce the stress-protective action of it, but on the contrary – it intensifies this action preventing unfavourable effects of the alcohol. At the same time the TAG and FFA level in the liver tissue of rats given ethanol together with “Enoant” decreases even when using the lowest dose investigated (0.01 ml per 100 g) (Table 18). Since the content of TAG and FFA increases apparently due to the lipogenesis activation when using ethanol, which might lead to fatty infiltration of the liver, then reduction of this process activity could protect the liver.
Parameter | Group | |||||||||
Stress + Enoant | Stress + Polyphen | Stress + ethanol | Stress + Cabernet | Stress + Rkatsiteli | ||||||
str. | non-str. | str. | non-str. | str. | non-str. | str. | non-str. | str. | non-str. | |
Total lipids, mg/g | 149.17 33.14 | 151.84 19.86** | 130.98 23.66 | 201.93 33.71** | 193.32 25.61*/** | 220.54 23.22*/** | 171.39 28.66** | 180.31 15.98** | 164.93 16.93** | 190.03 18.21** |
TAG, mg/g | 4.65 0.78 | 3.77 0.47 | 6.00 0.32 | 5.94 1.08 ** | 7.86 0.25 ** | 6.89 0.49 ** | 4.90 1.08 | 3.88 1.08 | 5.74 0.61 | 4.93 0.68 |
АpоВ-LP, mg/g | 4.58 0.06 ** | 4.43 0.07*/** | 3.72 0.34* | 3.15 0.44* | 5.00 0.82 | 4.10 0.49*/** | 3.06 0.16*/** | 3.15 0.16*/** | 2.87 0.30*/** | 3.02 0.36* |
FFA, mg/g | 1.06 0.12 | 1.00 0.11 | 1.08 0.16 | 1.26 0.08* | 1.35 0.15* | 1.45 0.13* | 1.05 0.22 | 1.22 0.20 | 1.15 0.13 | 1.40 0.31 |
The effect of grape polyphenol complexes and grape wines on the liver lipid metabolism in rats with the neurogenic stress, in the crude tissue (in each group n=10).
Parameter | Group | |||||||||
Stress + Enoant | Stress + Polyphen | Stress + ethanol | Stress + Cabernet | Stress + Rkatsiteli | ||||||
str. | non-str. | str. | non-str. | str. | non-str. | str. | non-str. | str. | non-str. | |
GSH, mkmol/g | 6.02 0.27 */** | 3.70 0.42* | 5.12 0.46 | 5.48 0.34 | 4.40 0.34 | 2.87 0.29*/** | 4.64 0.70 | 4.38 0.35 | 4.47 0.51 | 3.59 0.50* |
-Тocopherol, nmol/g | 27.82 2.86*/** | 34.01 3.56 ** | 34.34 4.14 ** | 25.39 1.18*/** | 6.55 0.61*/** | 5.59 0.54*/** | 26.95 1.32*/** | 24.64 0.81*/** | 29.24 1.82*/** | 28.9 3.15 ** |
Ascorbic acid, mkmol/g | 1.31 0.13 ** | 1.29 0.10*/** | 1.17 0.23* | 1.25 0.15*/** | 0.59 0.07 */** | 0.81 0.06* | 1.04 0.21* | 1.21 0.11*/** | 1.24 0.20*/** | 1.24 0.17*/** |
Isolated double bonds, U/g | 18.63 1.88*/** | 15.99 1.50*/** | 20.40 1.34*/** | 18.66 1.52*/** | 16.98 0.66* | 13.89 1.73* | 24.40 2.92*/** | 16.15 1.76*/** | 26.67 1.62*/** | 18.82 1.90*/** |
Diene conjugates, mmol/g | 13.58 0.74*/** | 12.87 0.54*/** | 13.48 0.62*/** | 12.85 0.26*/** | 13.61 0.08*/** | 13.20 0.58*/** | 10.90 1.10 ** | 15.00 2.12* | 10.66 1.88 ** | 14.32 1.62* |
Ketodienes+conjugated trienes, U/g | 13.57 0.54* | 13.39 1.67 | 11.44 1.56 ** | 11.84 1.88*/** | 12.68 1.91 | 13.04 1.31 ** | 14.20 2.02* | 15.74 1.98 | 17.15 1.62* | 16.04 1.78 |
ТBA-active products, nmol/mg protein | 0.21 0.02* | 0.19 0.02 ** | 0.28 0.03* | 0.22 0.03 ** | 0.55 0.08*/** | 0.68 0.06*/** | 0.15 0.02 | 0.28 0.04* | 0.18 0.02 | 0.17 0.02 ** |
The effect of grape polyphenol complexes and grape wines on the liver tissue oxidant/antioxidant status in rats with the neurogenic stress (in the crude tissue, in each group n=10).
It should be noted that the effect of high doses of “Enoant” (0.1 and 0.15 ml/100 g) was ambiguous. Onthe one hand, it caused α-tocopherol accumulation in the liver that might be an indicator of their protective action, but on the other hand, it probably revealed some prooxidative effect initiating the increase in the content of the POL final products – thiobarbituric acid-active products, and also activating ApoB-containing lipoproteins oxidation. In this case the secondary oxidative stress developed. A tendency to decrease the lipid content in the liver and to increase it in the blood plasma testifies about it.
Such effect is typical for high doses of many antioxidants capable to reveal the prooxidant action, including α-tocopherol. These data indicate the necessity of reasonable attitude to antioxidants therapy, including “Enoant”.
Parameter | Group | ||||||
Stress +Ethanol | Stress+Ethanol+ Enoant, ml per 100 g of the body weight: | ||||||
0.01 | 0.03 | 0.05 | 0.07 | 0.1 | 0.15 | ||
Total lipids, mg/ml | 5.890.08* | 5.690.06* | 5.300.05* | 4.820.15* | 3.510.08 | 3.440.09 | 3.580.15 |
TAG, mg/ml | 0.910.07* | 0.990.04* | 0.730.04* | 0.520.03 | 0.430.03 | 0.390.02* | 0.500.02 |
Total cholesterol, mg/ml | 0.490.07 | 0.460.05 | 0.540.04 | 0.400.01* | 0.550.09 | 0.550.03 | 0.560.02 |
HDL, mg/ml | 0.930.02 | 1.000.06 | 1.020.07 | 0.910.05 | 1.810.03* | 1.230.06* | 1.320.04* |
АpоВ-LP, mg/ml | 1.730.05* | 1.840.05* | 1.480.06 | 1.440.03 | 1.180.05 | 1.410.02 | 1.640.04* |
-Тocopherol, nmol/мml | 4.110.34* | 4.880.17* | 5.840.14* | 6.680.24* | 8.200.34* | 9.230.35 | 8.800.46 |
Ascorbic acid, mkmol/L | 33.811.73* | 34.042.73* | 39.041.60* | 49.251.10* | 55.011.67 | 56.982.03 | 49.963.43 |
Diene conjugates in АpоВ-LP, mkmol/L | 28.110.34* | 28.990.14* | 29.350.80* | 28.811.30* | 23.910.51 | 20.601.43 | 28,271.35* |
ТBA-active products, mkmol/L | 2.390.55* | 2.010.30* | 1.480.16* | 1.110.04 | 1.180.34 | 0.770.21 | 1.310.20* |
Corticosterone, nmol/L | 35.254.27 | 24.003.03 | 28.254.54 | 16.000.44* | 25.500.50 | 17.502.33* | 21.606.00 |
The effect of different doses of polyphenol concentrate “Enoant” in combination with ethanol on the plasma parameters of the stress response development in rats with the neurogenic stress (in each group n=10).
At the same time small doses of “Enoant” have a relatively low biological activity; they do not reduce negative effects of ethanol intake and do not inhibit the stress response significantly.
Therefore, we can conclude that the most effective doses of “Enoant” are 0.05-0.07 ml/100 g of the body weight because with these doses “Enoant” has not only high stress-protective, antiatherogenic and hepatoprotective activities, but practically neutralizes negative effects of ethanol.
Thus, our results suggest that grape wines have a high stress-protective, antiatherogenic and hepatoprotective activity that is equal to grape polyphenolic non-alcoholic concentrates characteristics, and the wine components in the doses studied have prevented negative effects of ethanol. Introduction of ethanol to animals in the human equivalent dose – 0.43 ml/kg of the body weight increases their tolerance to stress, but is an unfavourable factor that could result in MS development, fatty infiltration of organs and other pathologies. The polyphenolic concentrates “Enoant” and “Polyphen” in the human equivalent dose – 0.3 ml/kg of the body weight reveal a significant stress-protective, hepatoprotective and anti-atherogenic activity under the action of the emotional-painful stress. Grape wines from “Cabernet” and “Rkatsiteli” grades in the human equivalent dose – 4.3 ml/kg of the body weight also reveal a high stress-protective, antiatherogenic and hepatoprotective activity equal to grape polyphenolic non-alcoholic concentrates, and the wine components in the doses used prevented the negative effect of ethanol.
The highest activity has been shown by the combination of “Enoant” and ethanol that corresponds to the ratio of components in dry red wines, as well as the absence of significant difference in the protective effects of red and white wines, in spite of the difference in the polyphenol content [55]. Based on these results, in the second series of our experiments we decided to investigate “Cabernet” and “Rkatsiteli” wine effects on the development of stress-reaction proatherogenic consequences under the action of the emotional-painful stress in different periods of introduction.
It has been shown that “Cabernet” had a higher level of the anti-atherogenic activity than “Rkatsiteli”; in relation to the stress-protective activity the wines of these grades did not differ markedly. Such effect is likely connected with accumulation of polyphenols in the organism.
To examine the last supposition it was necessary to determine how different periods of introduction of the investigated wines influenced the stress-reaction development. We have carried out the study of wine intake influence on the development of proatherogenic consequences of the emotional and painful stress in different terms after consumption.
The data obtained in the experiments showed significant improvement of the antioxidant status both in the blood plasma and the liver tissue one day after the introduction of “Cabernet” wine (tables 20, 21).
At the same time “Rkatsiteli” wine did not reveal such activity. A similar condition persisted for 2-5 days of administration.
Periods of time | Parameter | ||||||
Total lipids, mg/g | TAG, mg/g | GSH, mkmol/g | -Tocopherol, nmol/g | Diene conjugates, nmol/g | ТBA-active products, nmol/g | ||
Day 1 | C+Str | 94.945.65* | 3.800.11* | 2.120.17* | 18.880.79* | 14.930.37* | 1.680.10 |
R+Str | 103.164.63* | 5.150.61* | 2.370.28* | 15.600.39* | 15.940.39* | 2.240.32 | |
Day 2 | C+Str | 105.175.12* | 3.580.23* | 2.510.34* | 26.042.17* | 14.130.09* | 1.070.24* |
R+Str | 106.939.42* | 5.670.34* | 2.770.87* | 15.650.92* | 15.100.07* | 1.910.15 | |
Day 3 | C+Str | 115.3811.65* | 3.920.13* | 3.820.37* | 23.692.18* | 13.850.48* | 1.700.11 |
R+Str | 103.285.81* | 6.470.28* | 2.470.26* | 16.890.71* | 14.440.25* | 2.290.20* | |
Day 5 | C+Str | 139.148.06* | 5.570.31* | 2.430.37* | 26.962.12 | 13.190.34 | 1.350.15 |
R+Str | 116.663.60* | 6.880.37* | 2.430.42* | 18.641.18* | 14.220.16* | 2.060.13 | |
Day 8 | C+Str | 161.186.05* | 7.270.15 | 4.110.21 | 32.120.85 | 12.280.52 | 1.640.13 |
R+Str | 122.207.07* | 6.350.45 | 2.330.28* | 23.082.08* | 12.930.44 | 1.730.04 | |
Day 10 | C+Str | 181.829.24 | 8.040.63 | 5.150.42 | 30.622.53 | 11.860.12 | 1.300.19* |
R+Str | 153.995.30* | 8.410.56 | 3.220.48* | 26.311.26* | 11.840.48 | 1.290.14* | |
Day 12 | C+Str | 174.726.15 | 8.340.55 | 3.740.25 | 32.555.58 | 12.420.36 | 1.470.27 |
R+Str | 164.48.03* | 8.630.47 | 3.250.17* | 28.282.26 | 11.410.22 | 1.450.31 | |
Day 15 | C+Str | 162.1612.81 | 7.500.43 | 4.610.22 | 38.162.06* | 10.500.52* | 1.340.06* |
R+Str | 172.1310.42 | 7.680.69 | 4.420.34 | 41.992.42 | 10.270.63* | 1.240.17 |
The effect of prophylactic administration of grape wines of "Cabernet" (C) and "Rkatsiteli" (R) grades on the stress response development in the liver tissue in rats with theneurogenic stress in different periods of time, in the crude tissue (in each group n=10).
Periods of time | Parameter | |||||||
Total lipids, mg/ml | TAG, mg/ml | Total cholesterol, mg/ml | АpоВ-LP, mg/ml | -Тocopherol, nmol/ml | Diene conjugates, nmol/ml | Corticosterone nmol/l | ||
Day 1 | C+Str | 5.15±0.50* | 0.78±0.07* | 0.94±0.04 | 1.53±0.02* | 8.52±0.27* | 31.49±2.58* | 75.00±8.66* |
R+Str | 6.15±0.44* | 0.93±0.07* | 1.08±0.05# | 1.68±0.03* | 6.29±0.47* | 38.35±1.56* | 111.70±10.00 | |
Day 2 | C+Str | 3.87±0.23 | 0.71±0.04* | 0.89±0.03 | 1.52±0.02* | 8.90±0.72* | 31.65±1.92* | 96.6721.86* |
R+Str | 4.85±0.16* | 0.84±0.06* | 0.86±0.04 | 1.70±0.04* | 7.77±0.27 | 32.17±1.71* | 81.5416.50* | |
Day 3 | C+Str | 3.42±0.32 | 0.56±0.03 | 0.73±0.05* | 1.54±0.04* | 10.20±0.52 | 24.72±2.89 | 35.679.49 |
R+Str | 4.45±0.41 | 0.70±0.04* | 0.89±0.06 | 1.66±0.04* | 8.49±0.38* | 22.24±1.84 | 43.008.50 | |
Day 5 | C+Str | 3.54±0.27 | 0.49±0.05 | 0.72±0.02* | 1.45±0.06 | 10.28±0.65 | 28.34±1.77* | 41.5018.50 |
R+Str | 3.56±0.23 | 0.58±0.05 | 0.88±0.02 | 1.66±0.04* | 10.01±0.27 | 20.46±2.03 | 57.103.00* | |
Day 8 | C+Str | 3.47±0.39 | 0.49±0.08 | 0.77±0.02* | 1.36±0.03 | 12.02±0.35 | 20.47±2.03 | 42.5010.61 |
R+Str | 4.08±0.31 | 0.4±0.06 | 0.84±0.03 | 1.55±0.04 | 10.04±0.68 | 20.15±2.61 | 40.509.19 | |
Day 10 | C+Str | 3.73±0.24 | 0.54±0.05 | 0.69±0.02* | 1.33±0.02 | 11.03±0.89 | 22.68±2.46 | 34.0018.38 |
R+Str | 3.60±0.35 | 0.55±0.09 | 0.76±0.04* | 1.43±0.04 | 11.39±0.47 | 22.90±1.93 | 38.002.82 | |
Day 12 | C+Str | 3.71±0.35 | 0.49±0.07 | 0.65±0.03* | 1.26±0.04 | 11.18±1.01 | 21.72±1.57 | 25.502.12 |
R+Str | 3.55±0.45 | 0.054±0.06 | 0.75±0.03* | 1.35±0.03 | 11.40±0.93 | 20.08±1.45 | 27.001.31 | |
Day 15 | C+Str | 3.78±0.36 | 0.59±0.04 | 0.61±0.02* | 1.33±0.02 | 11.04±1.32 | 20.99±0.92 | 34.007.00 |
R+Str | 4.22±0.57 | 0.51±0.09 | 0.69±0.03* | 1.29±0.06 | 11.72±0.93 | 20.59±1.92 | 31.0010.82 |
The effect of prophylactic administration of grape wines of "Cabernet" (C) and "Rkatsiteli" (R) grades on the plasma parameters of the stress response development in rats with the neurogenic stress(in each group n=10).
However, on day 8 of administration the antioxidant and stress-protective effects of these wines were almost similar, and on the day 10 –they practically did not differ.
On days 12 and 15 there were also no differences as to the antioxidant and stress-protective action of the wines studied, which significantly reduced activation of the free radical oxidation under the action of stress normalizing the most of the indexes investigated.
Thus, the investigated wines are characterized by the high level of the antioxidant and stress-protective activity, and in the first days of introduction “Cabernet” wine improved more effectively the antioxidant status in the blood and the liver tissue than “Rkatsiteli” wine, but by day 10 the effects of the studied wines had no substantial difference.
Probably, these results are dependent on polyphenol cumulation in the organism because it is known that the polyphenol content of “Cabernet” is 10 times more than of “Rkatsiteli”.
Thus, the results suggest that “Cabernet” and “Rkatsiteli” wines have already revealed the high stress-protective, hepatoprotective and anti-atherogenic activity in the conditions of the emotional-painful stress on the 2-3 days after introduction, and practically normalized the oxidative status and the lipid metabolism under the action of stress in prophylactic administration within 10 days. This indicates that grape polyphenols possess a high total antioxidant activity. At the same time the last suggestion required further research.
In order to examine the effects of wine stocks and polyphenolic concentrates obtained from other grape grades on development of proatherogenic consequences of the emotional-painful stress we investigated the action of substances obtained from the grapes of hybrid grades “Krasen”, “Golubok” and “Podarok Magaracha” produced by the National Institute of Grape and Wine “Magarach”.
In the series of experiments we used purebred male rats that during 21 day were given daily, per os, table wine stocks of the grades “Podarok Magaracha”, “Krasen” and “Golubok” in the human equivalent dose corresponding to 300 ml of wine for a human with 70 kg of the body weight. Other groups of animals were given ethanol in the human equivalent dose corresponding to 30 ml of ethanol for a human with 70 kg of the body weight taking into account the species sensitivity coefficients, as well as the table wine stocks of the grades mentioned in doses equivalent to the polyphenol content of the given wines calculated by the polyphenol content in active doses (AD – 9 mg of polyphenols/100 g of the body weight).
The results have demonstrated that not only polyphenolic concentrates, but the table wine stocks also revealed a substantial stress-protective activity to a different extent (Tables 22-25).
In fact, “Krasen” table wine stock revealed the highest activity; the stress-protective activity was almost 2.4 times more the ethanol activity in the dose studied. This product effectively prevented the activation of free radical oxidation both in the blood (increased the level of compounds with isolated double bonds in the atherogenic ApoB-LP, decreased the content of peroxidation products – diene conjugates – almost 3 times comparing to the stressed animals, and 15% - comparing to the intact animals), and the liver tissue (prevented the antioxidant content decrease, particularly the content of -tocopherol and ascorbic acid returned practically to the intact level, and there was 40% decrease of the diene conjugates level). At the same time this table wine stock prevented hyperlipidemia and the shift of metabolism to the increased lipolysis, there was 60% decrease of the blood total lipid content comparing to the stressed animals, and 11% - comparing with the intact animals. At the same time the TAG content in the liver was equal to the intact level that also demonstrated the protective action of this table wine stock. Reduction of lipogenesis in the liver tissue under the action of this product is important, and it protects the organ from steatosis. It should be also mentioned that the given product normalized the cholesterol content in the blood plasma.
Group | Parameter | ||||
Total lipids, mg/g | TAG, mg/g | FFA,mmol/g | АpоВ-LP, mg/g | Lysosomal lipase, nmol/mg protein/min | |
Str.+Con.Podarok Magaracha (AD) | 149.03 2.59*,** | 5.74 0.07** | 4.46 0.04* | 4.22 0.03*,** | 0.45 0.03** |
Str.+Con. Krasen (АD) | 147.13 1.15* | 4.27 0.02*,** | 4.19 0.05*,** | 4.61 0.01*,** | 0.50 0.02 |
Srt.+WinePodarok Magaracha | 161.74 1.91*,** | 6.33 0.05** | 4.30 0.11*,** | 4.43 0.03*,** | 0.32 0.01*,** |
Srt.+WineKrasen | 155.88 1.35*,** | 6.04 0.16** | 3.24 0.04** | 4.50 0.07*,** | 0.56 0.03** |
Str.+Con.Podarok Magaracha (DW) | 141.29 1.79* | 4.95 0.15*,** | 3.83 0.09** | 3.18 0.03* | 0.65 0.02** |
Str.+Con.Krasen (DW) | 145.87 3.19* | 4.24 0.07*,** | 4.23 0.07*,** | 4.39 0.08*,** | 0.55 0.03** |
WinePodarok Magaracha | 182.4 3.08* | 6.51 0.07** | 2.70 0.08*,** | 4.80 0.10*,** | 0.37 0.01* |
Wine Krasen | 164.36 1.86 | 5.97 0.17 | 2.95 0.09* | 4.93 0.18* | 0.35 0.02* |
Con.Podarok Magaracha (AD) | 191.33 2.03* | 6.47 0.04* | 4.61 0.31* | 4.33 0.08* | 0.83 0.03* |
Con. Krasen (AD) | 170.4 2.09 | 6.15 0.14 | 3.22 0.05 | 4.97 0.11* | 0.69 0.02# |
Ethanol | 229.76 3.39* | 7.40 0.13* | 4.57 0.13* | 6.11 0.07* | 0.38 0.02* |
The effect of grape polyphenol concentrates and grape wines on the liver lipid metabolism in rats with the neurogenic stress(in the crude tissue, in each group n=10).
It is also necessary to point out that the control intake of the investigated substances (Tables 22-25) did not reveal negative effects on the organisms of theexperimental animals. Moreover, in addition to the antioxidant activity these substances revealed a significant hypocholesterolemic and anti-atherogenic action, which was more pronounced when using “Krasen” grade wine stock and the concentrate.
Group | Parameter | ||||
GSH,mkmol/g | -Тocopherol,nmol/g | Ascorbic acid, mkmol/g | Diene conjugates, nmol/g | ТBA-active products, nmol/ mg protein | |
Str.+Con.Podarok Magaracha (AD) | 3.34 0.02*,** | 24.01 0.47*,** | 1.21 0.01*,** | 14.96 0.22*,** | 0.49 0,01 |
Str.+Con. Krasen (АD) | 3.8 0.02*,** | 26.710.46*,** | 1.28 0.01*,** | 15.03 0.11*,** | 0,45 0.02 |
Srt.+WinePodarok Magaracha | 3.48 0.02*,** | 26.71 0.34*,** | 1.33 0.02*,** | 13.81 0.18#,** | 0.42 0.03** |
Srt.+WineKrasen | 3.57 0.08*,** | 28.46 0.75** | 1.41 0.03*,** | 13.36 0.40** | 0.21 0.01*,** |
Str.+Con.Podarok Magaracha (DW) | 2.00 0.07*,** | 21.72 0.51*,** | 1.02 0.03*,** | 16.05 0.11*,## | 0.47 0.01 |
Str.+Con.Krasen (DW) | 3.27 0.06* | 21.12 0.39*,** | 1.04 0.03*,** | 16.72 0.16*,## | 0.45 0.02 |
WinePodarok Magaracha | 4.68 0.10*,** | 33.59 0.60 | 2.16 0.04* | 10.42 0.53* | 0.15 0.01* |
Wine Krasen | 4.56 0.24 | 35.48 0.78* | 1.57 0.03 | 9.27 0.24* | 0.13 0.01* |
Con.Podarok Magaracha (AD) | 4.46 0.13 | 35.29 0.45* | 1.95 0.03* | 10.87 0.41 | 0.20 0.01* |
Con. Krasen (AD) | 4.82 0.14* | 27.84 0.39 | 2.00 0.04* | 10.26 0.06* | 0.15 0.01* |
Ethanol | 5.31 0.35* | 24.16 1.40* | 2.06 0.03* | 12.62 0.60 | 0.46 0.02 |
The effect of grape polyphenol concentrates and grape wines on the oxidant/antioxidant status in the liver tissue in rats with the neurogenic stress(in the crude tissue, in each group n=10).
Group | Parameter | ||||||
Total lipides, mg/ml | TAG, g/ml | FFA, mmol/L | Total cholesterol, g/ml | HDL, mg/ml | АpоВ-LP, mg/ml | Cortico-sterone, nmol/L | |
Str.+Con.Podarok Magaracha (AD) | 3.890.08** | 0.720.01*,** | 1.400.02#,** | 56.661.49*,** | 0.790.02*,** | 1.450.05*,** | 472** |
Str.+Con. Krasen (АD) | 4.120.08# | 0.670.01*,** | 1.160.03*,** | 58.701.77 | 0.820.03 | 0.,900.04*,** | 473** |
Srt.+WinePodarok Magaracha | 4.600.11* | 0.650.02*,** | 1.630.02* | 63.321.01** | 0.850.02** | 1.210.02** | 411*,** |
Srt.+WineKrasen | 3.420.09*,** | 0.520.03** | 1.400.04** | 64.691,70** | 0.860.02 | 1.080.03#,** | 421** |
Str.+Con.Podarok Magaracha (DW) | 5.250.07* | 0.740.02* | 0.740.02*,** | 56.671.15*,** | 0.870.03 | 1.750.07*,** | 602 |
Str.+Con.Krasen (DW) | 3.880.08** | 0.670.01*,** | 0.670.01*,** | 55.581.21*,** | 0.830.03** | 1.060.03*,** | 631* |
WinePodarok Magaracha | 3.720.09 | 0.380.01* | 1.140.02* | 60.781.64 | 1.40.02* | 1.110.02 | 542 |
Wine Krasen | 3.760.03 | 0.360.01* | 1.180.02* | 57.210.81# | 1.110.02* | 1.140.01 | 444 |
Con.Podarok Magaracha (AD) | 4.320.04* | 0.560.02 | 1.230.01 | 68.701.22 | 1.040.02* | 1.140.02 | 712* |
Con. Krasen (AD) | 3.780.04 | 0.390.02* | 1.430.04* | 60.891.67 | 1.160.02* | 1.200.02 | 352* |
Ethanol | 4.050.09 | 0.740.03* | 1.650.02* | 55.801.46* | 1.240.06* | 1.320.02* | 753* |
The effect of grape polyphenol concentrates and grape wines on the plasma lipid metabolism parameters and corticosterone level in rats with the neurogenic stress(in each group n=10).
Group | Parameter | ||||
PON, nmol/mlmin | Ascorbic acid, mkmol/L | -Тocopherol, nmol/ml | Isolated double bonds in ApоВ-LP | Diene conjugates in ApоВ-LP | |
Str.+Con.Podarok Magaracha (AD) | 174 2* | 37.26 1.60*,** | 7.77 0.21* | 2.07 0.09* | 27.03 1.30*,# |
Str.+Con. Krasen (АD) | 194 3*,** | 41.79 0.44*,** | 8.25 0.66*,** | 2.35 0.15* | 26.86 0.43* |
Srt.+WinePodarok Magaracha | 173 3* | 40.08 1.72*,** | 8.91 0.29** | 1.92 0.25* | 18.45 0.18*,** |
Srt.+WineKrasen | 189 2*,** | 54.30 0.97*,** | 10.42 0.16*,** | 2.47 0.03*,** | 14.50 0.35*,** |
Str.+Con.Podarok Magaracha (DW) | 159 1* | 35.46 1.10* | 7.04 0.07* | 1.52 0.06* | 25.73 0.68*,** |
Str.+Con.Krasen (DW) | 1792*,** | 48.59 0.96*,** | 8.03 0.06*,** | 2.21 0.07* | 27.68 0.60*,** |
WinePodarok Magaracha | 223 6 | 75.95 0.74* | 10.03 1.72 | 5.43 0.12* | 15.67 0.15* |
Wine Krasen | 215 3* | 74.79 0.34* | 11.52 0.24* | 5.55 0.15* | 15.81 0.15* |
Con.Podarok Magaracha (AD) | 221 4 | 67.04 1.40 | 9.49 0.24 | 4.83 0.11 | 16.87 0.21 |
Con. Krasen (AD) | 255 3 | 70.61 2.31 | 9.97 0.16 | 5.39 0.04* | 15.47 0.34* |
Ethanol | 236 3 | 79.49 2.18* | 10.22 0.36 | 5.75 0.17* | 17.82 0.24 |
The effect of grape polyphenol concentrates and grape wines on the plasma oxidant/antioxidant status in rats with the emotional-painful stress(in each group n=10).
Based on our findings, it is possible to state that antioxidant complexes, particularly polyphenol extracts and the concentrates obtained from Vitis Vinifera, which are safe and reveal the potent antioxidant and stress-protective activity, should be used for reduction of proaterogenic states consequences in the complex prophylactic and treatment of atherosclerosis as effective stress-protective remedies.
Thus, administrationof Vitis Vinifera substances can prevent the increase of the total lipoprotein and ApoB-LP content in the blood, and prevent the free radical process activation in the plasma lipoprotein particles, and, in general, normalize the lipid and lipoprotein metabolism in the liver in metabolic syndrome. These results have proven the ability of the investigated complexes to reduce such negative consequence of metabolic syndrome as development of atherosclerosis.
In addition, according to obtainedresearch data the polyphenolic concentrates possess a potent protective activity both in acute and chronical neurogenic stress.
Our studies suggest that multicomponent active substances with antioxidant properties are more effective in correction of the proatherogenic states caused by stress and metabolic syndrome negative effects in comparison with individual antioxidants (particularly, α-tocopherol). The research data suggest that the increased plasma antioxidant activity alone does not result in decreased foam cell formation, at least in the studied animal model. Moreover, in vitro studies have shown that α-tocopherol can be pro-oxidative rather than protective for lipids in isolated LDL. Similarly with vitamin E, vitamin C additives do not offer consistent benefit against atherosclerosis in animals.
The occurrence of tocopherol-mediated peroxidation and the mode of its prevention predicts that the balance of α-tocopherol and available coantioxidants, rather than α-tocopherol alone, determines whether LDL lipid peroxidation occurs in biological systems. Inhibition of the free radical process with the polyphenolic complexes administration can be associated with their ability to increase the level of antioxidants – α-tocopherol, ascorbic acid and reduced glutathione in the test animal liver tissue compared with the group of the stressed animals. The complexes obtained from Vitis Vinifera, in particular, polyphenolic concentrates “Enoant” and “Polyphen”, as well as grape wines (particularly “Cabernet”) with their moderate use revealed the potent antioxidant activity. The preliminary results also suggest that coantioxidants inhibit lipoprotein lipid peroxidation in vivo. Thus, if LDL oxidation causes atherosclerosis, the requirement for coantioxidants may explain why supplementation with individual antioxidants, particular vitamin E alone, overall has yielded inconclusive results in the controlled human and animal intervention studies.
In conclusion, our research results may be used for the development of the atherosclerosis prophylaxis strategy, and treatment of diabetes mellitus and metabolic syndrome because recent studies proved insufficient effectiveness of α-tocopherol and advantages of multicomponent antioxidant complexes administration. The high effectiveness of the polyphenolic complexes obtained from Vitis Vinifera, including polyphenolic concentrates “Enoant” (from grape of “Cabernet” grade) and “Polyphen” (from grape of “Rkatsiteli” grade) produced by the National Institute of Grape and Wine “Magarach” has been proven. Our results also confirmed the high effectiveness of the antioxidant complexes from grapes in the correction of the endothelial dysfunction, thus, including these extracts in the treatment schemes is very reasonable. As it would be expected from our observations, increasing the antioxidant oxidant defense by antioxidant supplementation has the ability to restore the endothelial vasomotor function.
An important question to be asked is whether the polyphenol antioxidants exerted their inhibitory effect on lesion progression only because of their antioxidant properties or, possibly, because of additional biological properties, in particular – the phytoestrogen activity.
However, further studies, especially in humans, are required to validate the role of these antioxidants in inhibiting LDL oxidation.
Nevertheless, there are some limitations in the use of the concentrates produced from red grade grapes because of the uric acid content changes.
Systemic sclerosis (SSc) is a multisystem, autoimmune disease characterized by excessive collagen deposition and fibrosis of the skin and internal organs. The autoimmune process may affect the lungs with the development of interstitial fibrosis, pulmonary hypertension, or both. Pulmonary hypertension (PH) may result from a pathologic process of remodeling in the pulmonary arteries, in which case it is referred to as pulmonary arterial hypertension (PAH). Pulmonary hypertension may also arise secondary to interstitial fibrosis with chronic hypoxemia or myocardial fibrosis with postcapillary pulmonary hypertension. Pulmonary arterial hypertension associated with systemic sclerosis (SSc-PAH) represents the second most common cause of PAH after the idiopathic form of the disease (iPAH). Pulmonary arterial hypertension is associated with a progressive rise in pulmonary vascular resistance that can result in right ventricular failure and death. Patients with SSc-PAH have higher mortality than the idiopathic form of PAH or PAH associated with other diseases, such as congenital heart disease. While there is a reasonable amount of information available pertaining to SSc-PAH, much of what we know about PAH in general comes from investigations of the idiopathic form of the disease. The current chapter will review current knowledge about PAH in the patient with systemic sclerosis and contrast it with information that distinguishes SSc-PAH from the idiopathic form of PAH.
\nThe prevalence of systemic sclerosis-associated PAH is reported to be between 5 and 15% of patients with systemic sclerosis [1, 2, 3]. There is wide variability in reported prevalence rates which range from as low as 3.7% [4] to as high as 43% [5]. This variability is in large part due to methods used to establish the diagnosis of pulmonary hypertension. While some prevalence studies base reported findings on echocardiography, others confirm diagnosis with right heart catheterization. Right heart catheterization (RHC) is the gold standard for accurate diagnosis of pulmonary hypertension and for distinguishing pulmonary arterial from postcapillary hypertension. Prevalence rates are consistently lower when diagnosis is determined by right heart catheterization [6]. In a meta-analysis, Yang et al. found 12 studies reporting the prevalence of PAH in SSc ranging from 3.6 to 32% with a pooled prevalence of 13%. Five of the 12 studies confirmed the diagnosis of PAH with right heart catheterization yielding a pooled prevalence estimate of 8.2%, while the pooled prevalence estimate from seven studies relying on echocardiography was 18% [7]. Even when pulmonary hypertension is diagnosed by right heart catheterization, some patients in cohort studies may refuse to undergo catheterization, thus affecting true prevalence [8].
\nThe prevalence of pulmonary hypertension in systemic sclerosis depends on the phenotypic form of systemic sclerosis and the pathophysiologic mechanism behind the development of PH. The Australian Scleroderma Cohort Study (ASCS) of 232 patients identified PH in 10.1% of patients with diffuse scleroderma and in 12.7% of those with the limited form of the disease [9]. Prevalence of SSc-PAH consistently exceeds interstitial lung disease-PH (ILD-PH) or postcapillary-PH (PC-PH). Evaluation of PH subtypes in the ASCS cohort revealed 83.6% with PAH, 2.2% with ILD-PH, and 7.8% with PC-PH. The DETECT study, which was designed to develop an algorithm for detection of PAH in SSc, included 145 patients all of whom underwent right heart catheterization revealing 19% with PAH, 6% with ILD-PH, and 6% with PC-PH [10]. An Italian cohort of 867 consecutive SSc patients included 69 patients confirmed to have pulmonary hypertension with point prevalence for PAH 3.7%, PH secondary to ILD 1.4%, and postcapillary-PH 1.3% [4]. The lower prevalence of PH in the Italian cohort study raised speculation that ethnic factors might influence the prevalence of PH in SSc.
\nPrevalence of SSc-PAH appears to depend on other factors, such as duration of systemic sclerosis, gender, and ethnicity. Observations in the Pulmonary Hypertension Assessment and Recognition of Outcomes in Scleroderma (PHAROS) study suggested patients who were female, Caucasian, or suffering with limited cutaneous scleroderma were more likely to have PAH [11]. Additionally, a reduction in DLCO below 55% predicted was noted in 79% of patients with SSc-PAH compared to 55% of patients with SSc alone. Other authors have observed a greater chance of developing SSc-PAH in male patients age 47 or older [12], patients with SSC more than 10 years [13], and those with DLCO <55% [14]. Iudici suggested that systemic sclerosis patients of Italian descent may be less likely to develop pulmonary hypertension based on observations that prevalence rates were substantially lower than those reported in Anglo-Saxon patients [4].
\nThe World Health Organization (WHO) has classified pulmonary hypertension into five distinct groups on the basis of the primary pathophysiologic mechanism leading to elevated pulmonary artery pressure (Table 1) [15]. In a generic sense, pulmonary hypertension is diagnosed when mean pulmonary artery pressure (mPAP) ≥ 25 mmHg is measured by pulmonary artery catheterization. Patients classified as WHO Group 1 develop an arteriopathy of the small precapillary pulmonary arteries characterized by endothelial proliferation, smooth muscle layer hypertrophy, in situ thrombosis, and formation of plexiform lesions (Figure 1). Pulmonary arterial hypertension is defined more specifically as a mPAP ≥25 mmHg and also a capillary wedge pressure (CWP) ≤ 15 mmHg and pulmonary vascular resistance (PVR) > 3 Wood units (WU) [16]. Those in WHO Group 2 have elevated pulmonary artery pressure with a postcapillary origin typically related to left heart disease or dysfunction. WHO Group 3 pulmonary hypertension is a consequence of chronic hypoxia and attendant vasoconstriction as seen in chronic lung diseases, such as pulmonary fibrosis or emphysema. The fourth WHO Group constitutes those with pulmonary vascular obstruction, most often due to chronic thromboembolic disease. Finally, WHO Group 5 is a group of patients with pulmonary hypertension of mixed etiologies that do not fit within the other categories.
\nThe World Health Organization classification of pulmonary hypertensive diseases.
Histopathology of vascular remodeling in pulmonary arterial hypertension. (A) Medial hypertrophy and cellular intimal thickening. (B) Fibrin thrombi in glomoid plexiform lesion. Reprinted by permission from Springer Nature: Ishibashi-Ueda and Ohta-Ogo [21]. Copyright 2017.
Patients who develop the characteristic vasculopathy of WHO Group 1 PAH experience a progressive rise in pulmonary vascular resistance resulting from the gradual occlusion of smaller vessels by cellular hyperproliferation, thrombosis, and plexiform lesion formation that obstruct blood flow. The resulting rise in resistance to blood flow through the pulmonary circulation causes right ventricular strain with initial compensation and hypertrophy. Eventually the rising resistance overwhelms the right ventricle resulting in its failure.
\nPulmonary hypertension as it occurs in the scleroderma spectrum of diseases can develop by virtue of one or more mechanisms and can be classified as WHO Group 1 with the characteristic features of a precapillary arteriopathy, as WHO Group 2 when scleroderma affects myocardial physiology, or as WHO Group 3 if the patient primarily suffers from interstitial fibrosis and hypoxemia. Patients with systemic sclerosis may have complex forms of pulmonary hypertension involving more than one of these mechanisms. Treatment is dependent on the mechanism or mechanisms behind rising pulmonary vascular resistance, so it is important to carefully establish the root cause, or causes, for pulmonary hypertension in this patient population. Pulmonary arterial hypertension is the most common form of pulmonary hypertension to affect patients with systemic sclerosis. Therefore, this chapter’s focus is primarily on the pathogenesis of pulmonary arterial hypertension. While other mechanisms leading to pulmonary hypertension in this group will be reviewed, the development of SSc-PAH is a devastating complication, and the greatest body of information available pertains to the WHO Group 1 type of arteriopathy.
\nCharacteristic histopathologic features of pulmonary vascular remodeling observed in the patient with WHO Group 1 PAH are well-described and involve all layers of the pulmonary arterial vessels (Figure 1A) [17, 19, 21]. It is not uncommon for a similar process to affect the postcapillary venules in systemic sclerosis. A majority of patients will have in situ vessel thrombosis [18]. Flow-limiting pathologic features involving the intimal layer of the pulmonary arteries include eccentric or concentric intimal thickening and formation of plexiform or angiomatoid lesions (Figure 1B) [17, 19, 21]. There is excessive cell proliferation and hypertrophy of the smooth muscle layer. Thickening of the adventitial layer, primarily due to collagen deposition, is also noted in these patients [17, 19, 20, 21].
\nAreas of eccentric intimal thickening may represent fibrotic organization of localized thrombi. This concept is supported by observations of myofibroblast infiltration and accumulation of mucopolysaccharides in these localized lesions along the vessel lumen [17]. Eccentric intimal lesions of this nature have been demonstrated in lung explants from patients with severe idiopathic PAH and those with the CREST (calcinosis, Raynaud phenomenon, esophageal dysmotility, sclerodactyly, and telangiectasias) variant of scleroderma [22]. Vascular resistance is also increased by concentric proliferation of the endothelial cell layer creating the well-described “onion skin” lesion that is thought to involve myofibroblasts and smooth muscle cells (SMC), as well as endothelial cells [23, 24]. Plexiform lesions consist of complex vascular networks with a myofibroblast core which distorts the vessel wall as it expands and extends into the lumen and the connective tissues surrounding the vessel [23]. A rosary of dilated channels may form an angiomatoid, or dilation, lesion and obstruct arterial flow [25]. Although specific mechanisms involved in intimal remodeling are yet to be defined with clarity, it is largely believed that the processes begin with endothelial injury and, in genetically susceptible individuals, result in endothelial proliferation, smooth muscle cell and myofibroblast migration, decreased apoptosis, and deposition of extracellular matrix [17, 18, 26].
\nNormally, the medial layer of muscularized arteries accounts for about 10–15% of the outer arterial diameter, while in iPAH, it may be 30–60% of the outside diameter [20, 27]. Thickening of the medial layer is largely due to cell hypertrophy; however, hyperplasia of the smooth muscle cells and accumulation of extracellular matrix also contribute to the shift in tunica media dimension [17, 28]. Non-muscularized arteries may become muscularized with peripheral extension of proximal smooth muscle cell segments and pericyte differentiation into smooth muscle cells [19].
\nThe adventitial layer is comprised of fibroblasts and extracellular matrix (ECM) components. While it accounts for roughly 15% of vessel diameter under normal circumstances, it may represent double that in the patient with PAH [17]. In addition to its role in providing structural support for the vessel, there is evidence that inflammatory cells and extracellular matrix components of the adventitia may serve a role in the regulation of cell activities in other layers [29]. Typical components of the pulmonary vascular ECM include elastin, collagens, fibronectin, tenascin, thrombospondin, growth factors, and matrix metalloproteinases and proteoglycans [30]. Normal vessel structural and functional integrity depend on a balance between ECM deposition and degradation. Turnover is regulated by matrix metalloproteinases, adamalysins, serine elastase, and endogenous enzyme inhibitors [31]. In PAH excessive deposition of ECM contributes to vascular remodeling and decreased vessel wall compliance. Examination of the pulmonary vascular ECM in iPAH reveals prominent deposition of collagens I and III, enhanced collagen metabolism, alterations in proteoglycans and elastin, upregulation of tenascin C which is involved in intimal hyperplasia, and modification of fibronectin contributing to SMC proliferation and migration [32, 33, 34, 35]. Scleroderma is a disease characterized by overproduction of ECM, although there have been no studies detailing ECM composition in SSc-PAH.
\nStudies comparing the pathologic features of iPAH to those with connective tissue disease- associated PAH (CTD-PAH), and specifically SSc-PAH, have highlighted both similarities and differences between the groups. In a study of lung explants from transplant recipients, Stacher et al. compared the features of vascular remodeling in patients with iPAH and CTD-PAH [36]. The investigators noted more pronounced morphologic changes in the smaller-sized and precapillary arteries in CTD-PAH. Plexiform lesions were noted with similar frequency but had a more scattered distribution in the patients with connective tissue disease. Histopathologic studies comparing these patient groups also reveal more active interstitial inflammation and fibrosis in systemic sclerosis and other connective tissue diseases [19, 36, 37]. In a study comparing tissue from 24 patients with SSc-PAH and 9 iPAH patients, Argula et al. noted fewer plexiform lesions and more interstitial cellularity and fibrosis in the SSc-PAH group, while there was little difference in intimal proliferation or arteriolar smooth muscle hypertrophy [37]. In contrast, Overbeek and colleagues found no plexiform lesions in a group of patients with limited cutaneous systemic sclerosis and PAH, while these lesions were present in 10 of 11 comparative iPAH patients [38]. Further intimal fibrosis and fibrosis of the pulmonary veins and venules were observed with significantly higher frequency in SSc-PAH. While there are similarities in the overall pattern of vascular remodeling in iPAH and SSc-PAH, differences are notable and suggest distinct pathogenetic mechanisms may be in play. Additionally, inflammation and fibrosis may have a greater role in SSc-PAH.
\nThe coordinated mechanisms leading to vascular remodeling in PAH have been the subject of intensive investigation in recent years. It has been 18 years since Gaine proposed a theoretical model of the pathogenesis of PAH [18]. This model continues to serve as a basis for our basic understanding of the pathobiology of the disease and has been a platform for the development of approved therapies for WHO Group 1 PAH in use today. The model suggests a convergence of factors including genetic susceptibility, exposure to risk factors, vascular injury, and endothelial dysfunction leading to progressive remodeling of vasculature and rising pulmonary vascular resistance.
\nEvidence of a genetic basis for PAH was first reported in 2000 with the discovery of the bone morphogenetic protein receptor II (BMPR2) gene mutation in patients with heritable PAH [39, 40]. Mutation of this gene has been identified in at least 70–80% of cases of heritable PAH and 15–25% of sporadic iPAH [41]. BMPR2 protein concentrations are decreased by 75% in lung tissue and endothelial cells from subjects with PAH [24]. Other gene mutations related to BMPR2 and its downstream signaling pathway are now known including mutations in ALK1, ENG, and genes encoding components of the SMAD downstream signaling pathway [19, 41]. Mutations unrelated to the BMPR2 signaling pathway have also been identified in a very small percentage of PAH patients and include KCNK3, which encodes a pH-sensitive potassium channel, and CAV1, which encodes a membrane protein, caveolin 1, which is essential for the formation of lipid rafts or caveolae [41]. While no link between the development of SSc-PAH and mutation of BMPR2 has been established, other unique mutations have been identified in the systemic sclerosis population with PAH. For instance, a rare functional polymorphism in the TLR2 gene, which promotes induction of interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-alpha), is associated with the development of PAH in systemic sclerosis [42]. The development of diffuse cutaneous scleroderma, fibrosing alveolitis, and PAH has been linked to polymorphisms in TNFAIP3, which regulates the NF-kB inflammatory pathway [43]. Genetic variation in the promoter region of UPAR, the urokinase-type plasminogen activator receptor, has been associated with digital ulceration and PAH in scleroderma patients [44]. Clearly, there is evidence for genetic susceptibility to develop PAH in systemic sclerosis, although genetic mechanisms appear to differ from those associated with iPAH.
\nBMPR2 is a member of the transforming growth factor-beta (TGF-beta) superfamily of genes and normally functions to limit proliferation of smooth muscle cells and enhance endothelial cell survival by inhibiting apoptosis [45]. In contrast, TGF-beta is thought to promote SMC proliferation, matrix deposition, and alterations in endothelial cell growth [46, 47]. TGF-beta is known for fibrotic effects in several disease states, among which are systemic sclerosis [48]. Evidence from investigations of heritable PAH and preclinical models of SSc suggests that endothelial injury and consequent pulmonary vasculopathy may arise from an imbalance in TGF-beta/BMP signaling pathways [49, 50, 51, 52, 53]. For example, reduced BMPR2 receptor expression in heritable PAH correlates with increased activity of TGF-beta and its downstream signaling pathways. Reduction in BMPR2 levels in patients with systemic sclerosis also correlates with enhanced activity of TGF-beta and downstream SMAD2 and MAPK signaling pathways. Based on these observations, a theory has been advanced that heightened TGF-beta activity in systemic sclerosis might suppress BMP signaling pathways that serve to protect the endothelium [19].
\nIn addition to the disruption created by structural remodeling in the pulmonary vessels, endothelial injury and dysfunction may lead to imbalances in production of mediators that affect vascular tone and platelet aggregation and further regulate cell proliferation. Immunochemical studies have demonstrated reduced levels of nitric oxide synthase and prostacyclin synthase in the pulmonary vascular endothelium [54, 55]. These enzymes are critical to the endogenous production of nitric oxide and prostacyclin, both of which have vasodilatory and antiproliferative effects. The production of thromboxane is increased leading to enhanced vasoconstriction and in situ thrombosis [56]. Vasoconstriction and cell proliferation are promoted by increased production of endothelin-1 by pulmonary endothelium [57]. Endothelin-1, survivin, and vascular endothelial growth factor (VEGF) have been found in plexiform lesions and may augment endothelial and smooth muscle cell proliferation while limiting cell apoptosis [57, 58, 59]. Levels of nitric oxide synthase, prostacyclin synthase, and tumor suppressors, such as caveolin-1, are reduced in the plexiform lesions [54, 55, 60]. The imbalances in production of vasoactive mediators have largely driven the development of treatments designed to counteract these imbalances and improve pulmonary vascular resistance.
\nThe role of serotonin in the pathogenesis of PAH has been a topic of interest and investigation for several years. Serotonin is thought to promote vasoconstriction and remodeling of pulmonary vessels by stimulating proliferation of SMCs and fibroblasts [61, 62, 63]. The induction of SMC proliferation may be affected by serotonin transporter activation of the platelet-derived growth factor-beta (PDGF-B) receptor [64]. In SSc patients, serotonin has been shown to induce ECM production by interstitial fibroblasts in a TGF-beta-dependent manner [65]. When a group of SSc-PAH patients were treated with ketanserin, a selective antagonist of S2 serotonergic receptors, a majority experienced reductions in pulmonary vascular resistance [66]. The serotonin pathway may hold promise for the development of new treatment approaches to SSc-PAH in the future.
\nEpigenetic mechanisms affecting changes in cellular function in PAH have been a focus of more recent research. Epigenetic processes alter gene expression without effecting changes in DNA sequence. Epigenetic mechanisms may involve DNA methylation, modification of histone proteins, or RNA interference via microRNAs [67]. Extensive methylation of cytosine residues in the CpG dinucleotide sequences of the BMPR2 gene promoter region suppresses BMPR2 gene expression in SSc-PAH [68]. Elevated histone deacetylase levels have been noted in the lungs of PAH patients, and the inhibition of the deacetylase reduces proliferation of vascular fibroblasts and PDGF-stimulated SMC growth [69]. A number of microRNAs have been identified that influence cellular functions in hereditary and iPAH [19]. For instance, miR424 and miR503 normally suppress expression of fibroblast growth factor-2 (FGF-2); however, they are decreased in iPAH leading to an upregulation of FGF-2 expression [70]. These are just some representative examples of the growing knowledge of the role of epigenetic factors in PAH.
\nThe discovery of gene mutations involving the TGF-beta receptor family focused attention on the role of cytokines and growth factors in vascular remodeling of PAH. Observations of inflammatory cell infiltrates associated with vascular lesions and the presence of elevated cytokine levels in PAH have further supported a role for inflammation in this disease process. Lymphocytes, macrophages, dendritic cells, and mast cells have all been demonstrated on histopathologic examination of immune cell infiltrates in vascular lesions [17, 71, 72]. Elevated levels of several cytokines have been reported in iPAH including IL-1beta, IL-2, IL-4, IL-6, IL-8, IL-10, and IL-12p70, TNF-alpha, and chemokines CXC3L1, CCL2, and CCL5 [73, 74, 75, 76, 77]. The exact role of inflammation in the pathogenesis of PAH is unclear. Inflammation may reflect a consequence of hypoxia associated with PAH, as acute and chronic inflammation are known to occur in the setting of hypoxia. Alternatively, inflammatory mechanisms may be the drivers behind vascular cell injury and dysfunction. Macrophages are known to concentrate within and around advanced vascular lesions in iPAH and are thought to play a significant role in the remodeling process [78]. IL-6 produced by activated adventitial fibroblasts has been shown to induce a macrophage phenotype with proinflammatory and profibrotic characteristics [79]. Speculation about the role of immune dysregulation is supported by an observed deficiency of regulatory T cells in the lungs from iPAH patients [80]. In contrast to a deficiency of T-cell subpopulations, circulating autoantibodies and ectopic expansion of pulmonary lymphoid tissue in PAH patients suggest there is excessive B-cell activation [81]. PDGF has been implicated in the pathogenesis of PAH. Although originally discovered as a product of platelets, isoforms of this growth factor are also known to be secreted by macrophages, endothelial cells, SMCs, and fibroblasts [82]. PDGF is a potent mitogen and chemoattractant for endothelial cells, smooth muscle cells, and fibroblasts. PDGF receptor-beta expression is more intense in small and postcapillary vessels in SSc-PAH than iPAH [83]. The PDGF receptor antagonist, imatinib, was investigated for treatment of PAH, and during the initial study phase, improvements in hemodynamics and exercise capacity were noted. However, the long-term extension study was terminated early due to severe and unexpected adverse events including intracranial hemorrhage, death, and other side effects [84].
\nAutoantibodies against endothelial cell antigens may promote pulmonary vascular remodeling, especially in SSc-PAH. The expression of anti-endothelial cell antibodies and target antigens has been confirmed in iPAH and SSc-PAH, although the role of these antibodies in the pathogenesis of PAH remains undetermined [85, 86]. Anti-endothelial cell antibody levels in serum of connective tissue disease patients with or without PAH were evaluated by Li and colleagues compared with control subjects [87]. While endothelial cell antibodies were detected at similar levels in connective tissue disease patients irrespective of whether PAH was present, one specific endothelial antibody subtype (anti-22kD) was only found in the patients with PAH. A second subtype (anti-75kD) was noted at significantly higher levels in patients with PAH. The investigators concluded that these subtypes of endothelial antibody might indicate a more specific risk for PAH in connective tissue diseases. Tamby also demonstrated the presence of serum immunoglobulin anti-fibroblast antibodies in patients with iPAH and SSc-PAH with distinct reactivity against target antigens [88]. While these observations imply immunosuppressive therapy should be a treatment option for SSc-PAH and possibly iPAH, there is no evidence to date that this approach is beneficial. A prospective, multicenter trial to evaluate the effect of rituximab on disease progression in subjects with SSc-PAH receiving concurrent standard medical therapy is currently ongoing.
\nCertain features of pulmonary vascular cell dysfunction in patients with PAH have led to the suggestion that vascular remodeling may represent a cancer-like process involving the cellular constituents of the pulmonary arteries. Investigators have reported evidence of proliferative, apoptosis-resistant, cancer-like behavior in endothelial cells, SMCs, and fibroblasts from subjects with PAH [89, 90, 91]. Specific observations leading to this concept include monoclonal expansion of endothelial cells from patients with iPAH when compared to patients with PAH associated with congenital heart disease, instability of short DNA microsatellite sequences within plexiform lesions, somatic chromosome abnormalities in the lungs of patients with PAH, persistent hyperproliferative and apoptosis-resistant state when endothelial cells are removed from their in vivo environment, and altered energy metabolism [92]. Enhanced proliferation of pulmonary vascular cells may be a consequence of excessive growth factor release from the ECM, alterations in growth factor production or receptor expression, and/or alterations in intracellular mitogenic signals [93, 94, 95]. Abnormal increases in key apoptotic factors including Bcl-xL, Bcl-2, and survivin have been reported in pulmonary vascular cells from PAH patients [58, 96, 97]. Although there is evidence of enhanced cell proliferation and resistance to apoptosis, vascular remodeling in PAH is distinguished from cancer in that there is no evidence that pulmonary vascular cells have the ability to reproduce in a clonal fashion without control.
\nEndothelial cells, SMCs, and adventitial fibroblasts from patients with PAH are not only more proliferative and apoptosis-resistant but rely more on glycolysis for energy production [89, 98, 99, 100]. Mitochondria demonstrate a metabolic shift from glucose oxidation to uncoupled aerobic glycolysis similar to that described in cancer cells [101]. The glycolytic pathway increases NADPH production which in turn enhances antioxidant defenses while producing ribonucleotides for DNA synthesis. This shift in metabolism serves as a mechanism to support rapid cell proliferation.
\nIncreased cytosolic calcium levels in SMCs of patients with PAH promote not only contraction but also hyperproliferation and apoptosis resistance [41]. Elevated cytosolic Ca+2 levels in PAH have been linked to downregulation of voltage-gated potassium channels, such as Kv1.5 [102]. Downregulation or dysfunction of voltage-gated potassium channels allows membrane depolarization and influx of calcium. Cytosolic calcium levels are further enhanced by impaired mitochondrial Ca+2 uptake. The resulting increases in intracellular calcium drive cells into the cell cycle, thus enhancing proliferation [103].
\nResearch has certainly revealed that the pathogenesis of PAH is a very complex process, and our understanding of the mechanisms involved is far from complete. Knowledge of imbalances in endogenously produced vasomotor regulators has allowed the development of therapies that have improved quality of life and survival. However, it is apparent that PAH is not merely a disease of vasomotor dysfunction, but one that involves complex genetic mechanisms, cytokines, inflammation, and metabolic derangements (Table 2). While the progressive arteriopathy of iPAH and SSc-PAH shares many features, research has disclosed distinct differences in the pathogenesis of the two entities that may lead to more effective treatments for each in the future.
\nSummary of factors involved in pathogenesis of pulmonary arterial hypertension.
The majority of patients with SSc-PAH are diagnosed with PAH when the pulmonary arteriopathy is well established, while a small percentage is diagnosed at an early, asymptomatic stage [2]. Even when symptoms are present, the symptoms of PAH are nonspecific and may be attributed to other causes. Mortality is higher in patients with SSc-PAH than iPAH or PAH associated with other disease processes, such as congenital heart disease [104, 105]. An estimated 1-year survival of 84% for patients with iPAH contrasts with a 55% rate of survival at 1 year in SSc-PAH [106]. Patients with SSc-PAH have a higher mortality rate than those with non-scleroderma connective tissue disease-associated PAH [107]. Further, mortality is higher in patients with SSc- PAH than in systemic sclerosis patients without lung involvement or with lung involvement other than PAH [108]. In recent years, PAH and lung fibrosis have replaced scleroderma renal crisis as major causes of death in systemic sclerosis [109]. Pulmonary arterial hypertension accounts for about 30% of deaths in systemic sclerosis [109, 110]. Three-year survival rates of 70, 50, and 20% have been reported in treated SSc-PAH patients with WHO FC 1, FC 2, and FC 3 symptoms, respectively [107]. Earlier discovery of PAH in the systemic sclerosis patient may have an impact on these discouraging survival statistics. In a study by Humbert et al., the 1-, 3-, 5-, and 8-year survival rates in a cohort of SSc-PAH patients managed according to routine practice were 75%, 31%, 25%, and 17%, respectively, compared to survival rates of 100%, 81%, 73%, and 64%, respectively, in a group managed in a proactive detection program [111]. These data underscore the importance of consistently screening patients with systemic sclerosis for PAH.
\nAlthough experts agree on the importance of screening for SSc-PAH in order to detect vascular involvement at an earlier stage, there is less consensus on the most effective algorithm to confirm the presence of PAH. Several risk factors have been identified that signal the potential for the development of SSc-PAH (Table 2). Patients who are older and have long-standing SSc are at greater risk of developing PAH [1, 112]. The limited cutaneous form of SSc has historically been considered a risk for PAH; however the presence of diffuse cutaneous SSc has also been reported with similar prevalence [13, 113]. Anticentromere antibodies (ACA), anti-U1-ribonucleoprotein antibodies (RNP), and a nucleolar pattern of antinuclear antibody (nucleolar-ANA) are associated with an increased risk of SSc-PAH [114, 115, 116, 117]. The absence of anti-Scl 70 has been associated with the development of PAH, while the presence of these autoantibodies is associated with the development of interstitial lung disease [14]. Symptoms that relate to PAH are nonspecific and typically relate to progressive right ventricular (RV) dysfunction. Common symptoms include shortness of breath, fatigue, weakness, chest pain, and syncope [118]. Physical findings suggesting PAH include an accentuated pulmonary component of the second heart sound, an RV third heart sound, a pansystolic murmur of tricuspid regurgitation, and a diastolic murmur of pulmonary regurgitation [16]. Jugular venous distension, hepatomegaly, ascites, edema, and cyanosis are findings in advanced disease. Certain findings on electrocardiogram, such as right axis deviation, RV hypertrophy, RV strain, and right bundle branch block, may point to a diagnosis of PAH. Electrocardiogram abnormalities are more likely to be found in severe PAH. A normal electrocardiogram does not exclude PAH. Plain chest radiography can also be helpful in diagnosing PAH if the X-ray demonstrates central pulmonary artery enlargement, pruning of the peripheral vessels, or enlargement of right heart chambers. A chest radiograph may be helpful in distinguishing other causes of PH if interstitial lung disease or pulmonary venous congestion is present. Similarly, pulmonary function tests can be very helpful in detecting airway disease or restrictive lung disease that could lead to WHO Group 3 PH. Pulmonary function testing in patients with SSc-PAH may reveal severe gas diffusion impairment. Mukerjee et al. noted that a DLCO <50% was 90% specific but only 39% sensitive in excluding a diagnosis of SSc-PAH [6]. A DLCO/VA <70% or FVC percent/DLCO percent >1.6 has been considered predictors for the development of SSc-PAH [119]. Pulmonary function testing and CXR or high-resolution CT scanning are helpful in distinguishing PAH from WHO Group 3 PH associated with ILD. Echocardiography has been considered a noninvasive alternative to RHC in determining the presence of SSc-PAH, although certain limitations are recognized. Factors affecting image quality have been noted to limit the ability to estimate pulmonary artery systolic pressure accurately in patients who were later confirmed to have PAH by RHC [120, 121]. Right heart catheterization is the gold standard for diagnosis of PAH and is required to confirm PAH. Right heart catheterization with saline volume challenge can be helpful in distinguishing WHO Group 2 PH due to abnormal left ventricular function in systemic sclerosis. Several algorithms have been proposed that rely on various combinations of symptoms, physical exam findings, biomarkers, PFTs, and findings on echocardiography to determine which patients warrant definitive study with right heart catheterization [10, 16, 120, 122, 123].
\nA screening algorithm including assessment of symptoms, Doppler echocardiography, and right heart catheterization was studied in a French prospective multicenter study by the Itinerair-Scleroderma Investigators Group that enrolled 599 patients with scleroderma [120]. The study was limited to patients without significant pulmonary function abnormalities. Patients with a velocity of tricuspid regurgitation (VTR) > 3 m/s regardless of symptoms and patients with a VTR 2.5–3 m/s with dyspnea were considered at risk for PAH and underwent right heart catheterization (RHC). Right heart catheterization confirmed mild PAH in 18 of 33 patients suspected of having PAH based on symptoms and/or Doppler echocardiography. Twelve of the 33 patients did not have PAH, and 3 patients were confirmed to have left heart dysfunction. This algorithm allowed early detection of SSc-PAH; however a substantial number of patients undergoing RHC did not have PAH.
\nAn alternative screening algorithm was suggested by the Australian Scleroderma Interest Group (ASIG) that employs N-terminal pro-brain natriuretic peptide (NT-proBNP) levels and PFT data to predict the presence of PAH [122]. Data to develop this algorithm were collected from the Australian Scleroderma Cohort Study, a multicenter study of risk and prognostic factors for cardiopulmonary outcomes in systemic sclerosis. NT-proBNP levels from SSc patients with confirmed PAH were compared with a group at risk for PAH (systolic PAP TTE > 36 mmHg, hemoglobin corrected DLCO <50% predicted, and/or FVC/DLCO percent predicted ≥1.6), a group with ILD, and a group of controls with no evidence of cardiopulmonary complications. NT-proBNP levels were positively correlated with systolic PAP by transthoracic echocardiogram, mean PAP by RHC, pulmonary vascular resistance, and mean right atrial pressure. The authors proposed a model in which patients screened positive when NT-proBNP was ≥209.8 pg./ml and/or DLCO was <70.3% with FVC/DLCO ≥1.82. They noted a sensitivity of 100% with specificity 77.8% for SSc-PAH but acknowledged a need for prospective validation of the model.
\nA third screening algorithm was employed in the DETECT study, a multinational, cross-sectional investigation of factors in SSc patients that could serve to detect PAH at an earlier stage [10]. A broad range of variables (112 in total) pertaining to standard demographic and clinical characteristics, serum tests, electrocardiography, and echocardiography were examined in patients with a diagnosis of systemic sclerosis for more than 3 years, a predicted DLCO <60%, and a predicted FVC ≥ 40%. About 466 patients underwent RHC and 87 (19%) were confirmed to have WHO Group 1 PAH. Univariate and multivariate analyses were used to select the variables with best discriminatory power for predicting PAH. These variables were incorporated in a two-step algorithm. Six non-echocardiographic variables were used in Step 1 to recommend echocardiography (FVC % predicted/DLCO % predicted, current/past telangiectasias, serum anti-centromere antibody, NT-pro BNP, serum urate, ECG with right axis deviation), and a decision to recommend RHC in Step 2 was based on right atrial area and velocity of tricuspid regurgitation (VTR). Complete Step 1 data were available for 356 patients. About 52 patients did not meet Step 1 criteria for referral to echocardiography. Of these, two patients (4%) were determined to be false PAH negative. Complete Step 2 data were available for 267 patients. About 69 patients did not meet Step 2 criteria for referral to RHC. Of these, one patient was determined to be false PAH negative. Of the 198 patients referable for RHC, 69 were true PAH positive. The overall sensitivity of this algorithm was 96% with a specificity of 48%, a 62% rate of referral for RHC, and a 4% false PAH negative rate.
\nSummary recommendations for PAH screening in systemic sclerosis from the 2015 European Society of Cardiology/European Respiratory Society (ESC/ERS) Guidelines for the Diagnosis and Treatment of Pulmonary Hypertension are summarized in Table 3. The guidelines support a combined approach incorporating biomarkers, PFTs, and echocardiography for baseline screening in patients with systemic sclerosis. Annual screening with these indicators should be considered for all patients with systemic sclerosis. Systemic sclerosis patients with a mean PAP from 21 to 24 mmHg should continue to be monitored closely for progression. Exercise echocardiography has been used for early detection of PAH in systemic sclerosis [124, 125]; however the ESC/ERS do not recommend this approach. A summary diagnostic algorithm is provided in Figure 3.
\n2015 ESC/ERS recommendations for pulmonary arterial hypertension screening in systemic sclerosis.
As understanding of the pathogenesis of PAH has evolved over the past two decades, a number of medical therapies have been developed that improve exercise capacity, hemodynamics, quality of life, and survival. Treating PAH has become a complex exercise now that there are multiple agents that can be employed alone or in various combinations. It is important to remember that PAH is a progressive disease process, and any treatment plan requires ongoing monitoring and adjustment if treatment goals are not being met. Although far fewer patients with PAH require lung transplantation in the era of targeted medical therapy, there are those who progress even on maximal medical therapy leaving transplantation as their last viable option.
\nThe treatment of pulmonary arterial hypertension involves not only selection of appropriate agents for inclusion in a treatment plan but an ongoing process of assessment of patient characteristics that should determine the selection of medications. The concept of using a risk assessment tool to aid selection of appropriate agents for treatment was introduced in 2006 based on studies showing correlation between clinical characteristics of disease and survival [104]. For instance, 6 MW, FC, and certain hemodynamic measures were shown to directly correlate with prognosis. These findings were used to develop a tool to evaluate a patient’s risk of early death. Patients could be categorized as low or high risk of rapid progression to death, and treatment agents could be selected based on the level of risk in order to modify the course of disease and extend survival. This concept further evolved with the development of a risk calculator using data from the American REVEAL Registry [126, 127]. The REVEAL Registry was a 3-year longitudinal registry of 2967 WHO Group 1 PAH patients with data collected pertaining to the clinical characteristics, evaluation, treatment, and outcomes. Data from this registry was used to develop a user-friendly algorithm to determine a patient’s risk of demise in the short term. Most recently, the ESC/ERS further refined the characteristics used to assess risk of progression in PAH and presented criteria that classified patients as low, moderate, or high risk of progression to near-term death (Figure 2). Using the ESC/ERS risk assessment algorithm, patients categorized as low risk have an estimated 1-year mortality <5%. Those within the intermediate-risk group have an estimated 1-year mortality of 5–10%, and those in the high-risk group have an estimated 1-year mortality >10%.
\nESC/ERS risk assessment in pulmonary arterial hypertension (estimated risk for 1-year mortality).
Following the accurate diagnosis of PAH (Figure 3), a careful assessment of severity of disease should be completed before deciding on specific treatment options. This assessment is critical at the outset of treatment, but it remains an important part of ongoing patient management. Given that PAH can progress rapidly, even on therapy, it is necessary to complete a reevaluation of severity of illness and risk stratification periodically several times a year. If patients show signs of deterioration in their clinical parameters, treatment plan adjustments are in order.
\nAlgorithm for the diagnosis of pulmonary arterial hypertension.
Assessment of the severity of illness begins with an understanding of symptoms and functional capacity. Patients who present symptoms, such as shortness of breath, fatigue, or edema, that have developed and worsened over a short period of time are at higher risk of early death. Further, those with overt signs of right heart failure, such as edema, ascites, cyanosis, or syncope, are in a high-risk group requiring urgent attention. The World Health Organization functional class (FC) is a valuable indicator of severity of illness and has been shown to correlate with survival [128, 129]. Patients are classified in four groups (FC 1–4) based on degree of functional impairment (Table 4). Patients who have FC 3 or 4 functional impairment are considered high risk; a goal of any treatment plan is to achieve FC 1 or 2 functional capacity [16]. Although functional class has been shown to correlate well with survival prognosis, it is a subjective measure of symptoms which is subject to interpretation by the healthcare provider [130]. Another important indicator of illness severity is the 6 min walk (6 MW) test [131]. The 6 MW test is a submaximal exercise test that is easy to perform in the outpatient or inpatient setting. The 6 MW test has been shown to correlate with survival and has served as a primary endpoint in the majority of clinical investigations leading to today’s therapeutic options [132, 133]. Six-min walk distance has been shown to correlate with pulmonary pressures and represent a direct predictor of mortality in SSc-PAH [131]. The ESC/ERS Guidelines suggest that patients who can ambulate >440 m have better survival prognosis and are an appropriate goal to target when making treatment decisions [16]. The 6 MW test does have limitations with its reliability being challenged by factors such as age, gender, weight, comorbid conditions, and the individual’s motivation. Cardiopulmonary exercise testing (CPET) is often employed to evaluate exercise capacity in this population. CPET can provide important information about general exercise capacity, as well as more detailed information about gas exchange, ventilation, and cardiac function during exercise. Patients with PAH will demonstrate exercise limitation characterized by low end-tidal partial pressure of carbon dioxide (pCO2), high ventilation equivalents for carbon dioxide (VE/VCO2), low oxygen pulse (VO2/HR), and low peak oxygen uptake (peak VO2) [134, 135]. These parameters have been included in the ESC/ERS risk assessment algorithm. Peak VO2 > 15 ml/min/kg or greater than 65% predicted and a VE/VCO2 slope < 36 portend a more favorable prognosis and represent goals of targeted therapy [136].
\nThe World Health Organization (WHO) functional class.
Echocardiography is not only a valuable screening tool for detecting the presence of pulmonary hypertension, but it plays a role in assessing severity of illness and response to treatment. The measurement of pulmonary artery pressure (PAP) by echocardiography is not a reliable prognostic indicator, nor does it accurately reflect progression or improvement in pulmonary vascular resistance [121, 128]. The absence of tricuspid regurgitation and/or poor image quality limits the ability to determine systolic PAP by echocardiography in 20–39% of patients [121]. The value of echocardiography in assessing severity of illness lies in measurement of chamber sizes, assessment of right ventricular (RV) function, and the presence or absence of pericardial effusion which is considered a reflection of RV failure. A complete echocardiographic assessment in the PAH patient would include description of right atrial (RA) and RV dimensions, measurement of tricuspid regurgitant velocity, left ventricular (LV) eccentricity index, and RV contractility [137, 138]. RV contractility can be determined from RV longitudinal systolic strain/strain rate, RV fractional area change, Tei index, or tricuspid annular plane systolic excursion (TAPSE) [137, 139]. Echocardiography with exercise may provide useful information about RV function. An increase in estimated PAP by >30 mmHg during exercise indicates better RV reserve associated with better long-term outcome and is considered an independent marker of prognosis in PAH [140]. A right atrial area < 18 cm3 with no evident pericardial effusion are indicators for favorable prognosis or treatment outcome [16]. Additional information about prognosis or treatment effect can be gained from right heart catheterization and hemodynamic measurements. As in the case of echocardiography, PA pressure is of little value. Cardiac index, RA pressure, and mixed venous oxygen saturation have been shown to have the greatest prognostic significance [128, 129]. Goals of therapy that are associated with favorable prognosis include CI ≥ 2.5 L/min/m2, RA pressure < 8 mmHg, and SvO2 > 65% [16].
\nThere are a number of biomarkers of vascular dysfunction, inflammation, cardiac function, and tissue hypoxia that have been investigated as a specific marker for pulmonary vascular remodeling [141, 142, 143, 144]. Of these, brain natriuretic peptide (BNP) and NT-proBNP are used in clinical practice and research [145, 146, 147]. These biomarkers reflect ventricular wall stress, as seen in volume overload and ventricular contractile dysfunction, and serve as surrogate indicators of myocardial dysfunction [122, 148]. Either marker is considered an acceptable choice for assessing severity of illness. BNP has slightly better correlation with pulmonary vascular hemodynamics and is less likely to be affected by renal function, while NT-proBNP appears to be a stronger predictor of prognosis [149]. Compared to BNP, NT-proBNP is more sensitive to early increases in systolic PAP as measured by echocardiography [150]. BNP levels below 50 ng/L or NT-proBNP levels below 300 ng/L are associated with a more favorable prognosis [16].
\nImplementation of the prognostic indicators outlined in Figure 2 is variable among centers providing expert care for patients with PAH. After a treatment plan is established, patients will typically be reevaluated every 3–4 months depending on stability of their disease. During early phases of treatment or times when therapy targets indicate a need to alter the treatment plan, patients are often seen more frequently. It is not practical to perform all of the measures listed in Figure 3 at every visit. In the outpatient setting, clinicians tend to rely on assessment of symptoms, physical exam findings, FC, 6 MW distance, and BNP or NT-proBNP levels to determine severity of disease at any given point in time. This information may be supplemented periodically with echocardiography or cardiopulmonary exercise testing. Right heart catheterization is performed initially at diagnosis and in some centers yearly thereafter or in the event condition deteriorates in those patients with high-risk features. In other centers repeat hemodynamic measurements are obtained less frequently and typically if there is an indication the patient’s condition is progressing.
\nOnce the diagnosis is established and severity of illness is defined, decisions about disease-targeted therapy can be made. Available targeted therapies exert clinical benefit via the nitric oxide, endothelin, or prostacyclin pathways which were discussed earlier in this chapter. There are several options that affect each of these pathways. The choice of therapy for any given patient will depend on severity of illness and may be further influenced by side effects, safety issues, and in some cases economic and social support factors.
\nThe phosphodiesterase 5 (PDE-5) inhibitors effect smooth muscle relaxation and inhibit proliferation and inflammatory mechanisms by augmenting cyclic guanosine monophosphate (cGMP) activity in pulmonary vascular smooth muscle. As noted earlier in this chapter, patients with PAH have been shown to have deficient nitric oxide synthase activity in the pulmonary vasculature leading to a deficiency of nitric oxide production [54]. Nitric oxide is produced by the pulmonary endothelium and catalyzes the production of cGMP in nearby smooth muscle cells. PDE-5 degrades cGMP, thus limiting its effect on smooth muscle cells. The phosphodiesterase 5 inhibitors sildenafil and tadalafil block the degradation of cGMP, thus permitting beneficial cGMP effects to continue.
\nSildenafil has been shown to improve symptoms, exercise capacity, and hemodynamics in patients with PAH including those with connective tissue disease (CTD) [151, 152]. It is available as an oral agent prescribed at 20 mg tid. This drug is generally well tolerated with most common side effects including headache, flushing, nausea, and nasal congestion, all resulting from vasodilation. The use of nitroglycerin is contraindicated in patients taking sildenafil due to a risk of severe hypotension when these drugs are used in combination. Safety during human pregnancy has not been studied; however no fetal harm has been noted in animal studies.
\nTadalafil has also proven to have beneficial effects on symptoms, exercise capacity, hemodynamics, and time to clinical worsening in a large randomized clinical trial involving 405 PAH patients including 95 with connective tissue disease [153]. Tadalafil’s greatest benefit was realized at a dose of 40 mg daily. The drug is well tolerated with side effects and precautions similar to sildenafil.
\nVardenafil is a third agent within the PDE-5 inhibitor class that demonstrated significant advantage when comparing 6 MW distance, cardiac index, mean PA pressure, and pulmonary vascular resistance in patients treated with vardenafil 5 mg twice daily or placebo [154]. The long-term effects of vardenafil in PAH have not been evaluated, and the drug has not been approved for the treatment of PAH in the United States.
\nWhile the phosphodiesterase 5 inhibitors promote vasodilation and limit proliferation by preventing the degradation of cGMP, the soluble guanylate cyclase (sGC) stimulator riociguat interacts directly with guanylate cyclase to stimulate production of cGMP [54].
\nRiociguat is the only member of this family in use to date. Riociguat was studied in two randomized clinical trials, one focused on patients with PAH (PATENT 1) and included those with CTD [155] and the other patients with chronic thromboembolic pulmonary hypertension (CTEPH) [156]. Significant improvements were observed in exercise capacity, FC, time to clinical worsening, and hemodynamics. Subgroup analysis of the PATENT 1 trial specifically evaluating benefit in CTD-PAH revealed improvements in 6 MW, FC, pulmonary vascular resistance, and cardiac index [157]. Riociguat is an oral therapy with maximum daily use of 2.5 mg tid. Side effects are similar to those seen with the PDE-5 inhibitors. In addition, riociguat can induce systemic hypotension and has been linked to an increased risk of bleeding. It is teratogenic and contraindicated in pregnancy. Females of childbearing age are required to participate in a Risk Evaluation and Mitigation Strategy (REMS) program and undergo monthly pregnancy testing in addition to practicing careful contraceptive measures. This drug should not be used with nitroglycerin or PDE-5 inhibitors due to the risk of severe hypotension.
\nExcessive levels of endothelin 1 produced by pulmonary vascular endothelial cells have been implicated in the vasoconstriction and cell proliferation seen in PAH [57, 158, 159]. Endothelin binds with two G protein-coupled receptors, type A and B, located on the smooth muscle cell surface and thereby promotes its physiologic effects. Type A receptors mediate vasoconstriction, cell growth, and inflammation, while type B receptors mediate opposing effects including vasodilation and natriuresis while inhibiting proliferation and inflammation.
\nBosentan was the first targeted oral therapy developed to treat PAH and is prescribed with a bid dosing schedule. Bosentan has been investigated in patients with iPAH, PAH associated with CTD, and Eisenmenger syndrome [160, 161, 162]. The drug interacts with both type A and B receptors to effect improvements in exercise capacity, FC, time to clinical worsening, hemodynamics, and echocardiographic variables [163]. About 10% of patients treated with bosentan in clinical trials developed reversible elevations in liver transaminases. Monthly monitoring of liver function tests is required for patients using bosentan. Other side effects that can be seen are fluid retention and anemia. Further, this drug is teratogenic and contraindicated during pregnancy. Females of childbearing age who use bosentan must enroll in a Risk Evaluation and Mitigation Strategy (REMS) program and are required to undergo monthly pregnancy testing. They should be counseled to avoid pregnancy with careful contraceptive practices if sexually active. It is important to note that hormonal contraceptive effectiveness is reduced by bosentan. It is also important to note that cyclosporine and glyburide may increase bosentan levels and increase the risk of liver toxicity.
\nAmbrisentan is a selective endothelin type A receptor blocker which is available as an oral therapy prescribed for once daily use. This drug has been studied in one pilot and two randomized clinical trials demonstrating improvements in symptoms, exercise capacity, time to clinical worsening, and hemodynamics in patients with iPAH, CTD-PAH, and HIV-associated PAH [164, 165]. The risk of liver function abnormalities is minimal, and monthly liver function testing is not required for patients using ambrisentan; however its use is not recommended in patients with moderate to severe liver dysfunction. Ambrisentan use can be complicated by the development of edema and anemia. Like bosentan, ambrisentan is teratogenic and contraindicated during pregnancy. All of the precautions relating to use in females of childbearing age noted for bosentan are also true for ambrisentan.
\nMacitentan is the most recent endothelin 1 antagonist available to PAH patients as a once daily oral therapy. Like bosentan, macitentan is a dual endothelin receptor blocker. In contrast to bosentan and ambrisentan, the benefits of macitentan were realized in a large event-driven investigation involving 742 patients treated for an average of 100 weeks [166]. Macitentan significantly reduced time from initiation to a composite endpoint of worsening PAH, initiation of intravenous or subcutaneous prostanoid therapy, atrial septostomy, lung transplantation, or death. The study population included a significant proportion of patients on background therapy who also experienced significant benefit. Macitentan is well tolerated and, as with other endothelin antagonists, may be associated with fluid retention or anemia. Again, this drug is teratogenic and contraindicated during pregnancy. Patients using this drug should follow the same risk reduction recommendations as noted with bosentan and ambrisentan.
\nA deficiency of prostacyclin activity characterizes the dysfunction of the third major pathway involved in the development of PAH. Prostacyclin is produced by the pulmonary endothelium, and its bioactive effects include vasodilation of the pulmonary vascular bed, inhibition of platelet aggregation, and cell proliferation [167]. A reduction of prostacyclin synthase expression has been recognized in pulmonary arteries from patients with PAH and is thought to be the central focus of dysfunction in this pathway [55]. The prostacyclin analogues are available as oral, inhaled, or systemically administered disease-targeted therapies.
\nEpoprostenol is available for use as a continuous intravenous (IV) infusion. Epoprostenol has a short half-life of 3–5 min. The original formulation was unstable at room temperature after about 8 h and required considerable effort to maintain at cooler temperatures. A newer formulation of the drug is now available that has extended room temperature stability. Treatment is initiated at a dose of 2–4 ng/kg/min and titrated upward to reach clinical therapy targets. Patients experience tachyphylaxis with the continuous infusion, therefore requiring intermittent dose escalation over time. The maximum beneficial dose of epoprostenol is typically 40 ng/kg/min, although titration may go beyond this point. Epoprostenol has been shown to improve symptoms, exercise capacity, and hemodynamics in FC 3 and 4 patients with iPAH and SSc-PAH [168, 169, 170]. Side effects with epoprostenol can be pronounced and may include jaw pain, nausea, diarrhea, flushing, and headache. There is a risk of catheter-related complications including infection and thrombosis. Epoprostenol can cause hypotension when used with other antihypertensives and may increase risk of bleeding when used in patients taking anticoagulants or antiplatelet agents. Epoprostenol has been used during pregnancy without evidence of fetal harm to date. Given the short half-life of epoprostenol, an infusion of this drug should not be discontinued abruptly due to the risk of rebound pulmonary vasoconstriction and death.
\nTreprostinil is an analogue of epoprostenol available in systemic, inhaled, and oral formulations. The systemically infused form of treprostinil is stable at room temperature, has a half-life of 3–4 h, and can be administered by continuous subcutaneous (SC) or IV infusions. Dosing typically begins with 1–2 ng/kg/min with gradual dose escalation to achieve clinical target goals. Side effects are similar to epoprostenol. Additionally those patients using the subcutaneous formulation may experience significant infusion site pain. Several topical analgesic preparations are available that can successfully control local infusion site pain. As is the case with epoprostenol, patients develop tachyphylaxis requiring dose escalation to maintain clinical benefit. The usual effective dose is 20–80 ng/kg/min, although dosing can extend well beyond this range. Treprostinil was first studied in its continuous SC formulation. A randomized clinical trial of 470 patients treated with SC treprostinil, including 17% CTD patients, revealed improvements in exercise tolerance and hemodynamics [171, 172]. Dose titration was limited by side effects, including infusion site pain, and as such benefits were noted in those patients achieving higher doses >13.8 ng/kg/min. Later treprostinil was approved for use as a continuous IV infusion. Treprostinil can be administered in an inhaled formulation with a specialized nebulizer four times daily. This formulation is very well tolerated with most commonly reported effects including mouth soreness, cough, and headache. Tachyphylaxis does not develop due to intermittent dosing. Some patients may notice recurrence of PAH symptoms as effect wanes between treatments. In a randomized clinical trial of inhaled treprostinil added to background therapy with bosentan or sildenafil, there were improvements in 6 MW, NT-proBNP levels, and quality of life measures [173]. More recently, treprostinil has been offered in an oral formulation that is taken by either bid or tid scheduled dosing. Although intermittent dosing is employed in the treatment of patients with oral treprostinil, dose escalation over time helps achieve and maintain clinical target goals. The use of oral treprostinil can be complicated by significant gastrointestinal side effects, such as nausea, anorexia, and diarrhea. In a randomized clinical trial of treatment-naïve PAH patients, oral treprostinil use was associated with improvement in 6 MW distance [174]. Treprostinil can cause hypotension when used with other antihypertensives and may increase risk of bleeding when used in patients taking anticoagulants or antiplatelet agents. Parenteral and inhaled treprostinil safety during pregnancy has not been studied in humans but did not lead to fetal harm in animals, and as such they have Category B designations. Oral treprostinil has been associated with adverse fetal effects in animal studies and is designated Category C. Continuous IV therapy carries a risk of catheter-related complications including infection and thrombosis. Oral treprostinil use is contraindicated in patients with Child-Pugh Class 3 hepatic impairment. Treprostinil should not be discontinued abruptly due to the risk of rebound pulmonary vasoconstriction and death.
\nIloprost is a stable analogue of prostacyclin that is also available in IV, inhaled, or oral formulations. Oral iloprost has not been evaluated for use; however, both the IV and inhaled forms have been used in Europe, and the inhaled form has been approved for use in the United States. The inhaled formulation is administered with a specifically designed handheld and portable nebulizer device. This form of iloprost is used by nebulization 6–9 times a day at a dose of 2.5–5 ug/inhalation. The effect lasts from 30 to 90 min. The intermittent dosing eliminates the development of tachyphylaxis. Improvements in symptoms, exercise capacity, and pulmonary vascular resistance were observed in a clinical trial in which iloprost was compared with placebo in patients with PAH and CTEPH [175]. The effect of IV iloprost was noted to be similar to epoprostenol in a small group of patients with PAH and CTEPH [176]. The inhaled drug is well tolerated with most frequent side effects being cough, flushing, and jaw pain. Inhaled iloprost can cause hypotension and should be avoided or used cautiously in patients with relative hypotension.
\nThe development of the prostacyclin receptor agonist class of disease-targeted therapies represents a new approach to treating PAH. Although the prostacyclin receptor agonism of this new class is similar to that of prostacyclins, the receptor interaction is selective for the IP receptor. The established prostanoid receptors in the human pulmonary artery are the IP, EP3, and TP receptors. The IP receptor mediates vasodilation and inhibits proliferation, while the EP3 and TP receptors may promote vasoconstriction and cell proliferation [177, 178, 179].
\nSelexipag is a selective IP receptor agonist that is structurally distinct from prostacyclin with an active metabolite that is 37-fold more potent. Selexipag is prescribed for oral BID dosing beginning with 200 mcg bid and titrating to a maximal dose of up to 1600 mcg bid. The target treatment dose for individual patients is determined by the development of side effects limiting further dose escalation. Selexipag reduced the risk of reaching a composite morbidity and mortality (worsening PAH resulting in need for atrial septostomy or lung transplantation, initiation of parenteral prostanoid therapy or chronic oxygen therapy, hospitalization for PAH, other indication of disease progression, or death) by 40% in a large placebo-controlled, event-driven trial including 1156 patients [180, 181]. At baseline, 80% of patients were being treated with stable doses of an endothelin blocker, a PDE-5 inhibitor, or both. A subgroup analysis of 334 patients with connective tissue disease-associated PAH (170 SSc, 82 systemic lupus, 82 mixed or other) revealed a similar 41% reduction in risk of the composite morbidity and mortality events [182]. Further the treatment effect was consistent regardless of background PAH treatment or connective tissue disease subtype. Commonly reported side effects include headache, nausea, diarrhea, flushing, myalgia, and arthralgia.
\nDespite observations from clinical trials that individual therapeutic agents can improve exercise capacity, time to clinical worsening, and hemodynamics, pulmonary arterial hypertension remains a progressive disease that is difficult to control. The progressive nature of this disease process in patients treated with monotherapy has fostered the practice of combining agents to limit progression. One approach has been the sequential addition of agents affecting the three known pathophysiologic pathways. In this approach an agent affecting one of the pathways is chosen to begin monotherapy and if clinical response is inadequate, one or more agents affecting the other pathways are added until desired clinical benefit is achieved. Upfront combination therapy has become a more contemporary approach to managing pulmonary arterial hypertension. This approach to treating PAH was conceived from experience with the treatment of other disease states, such as cancer or congestive heart failure, with agents affecting multiple mechanisms of disease upfront. The upfront combination approach gained momentum with the AMBITION trial which demonstrated a 50% reduction in composite morbidity/mortality events in patients treated with an upfront combination of tadalafil and ambrisentan compared to either agent as monotherapy [183]. This benefit was also recognized in a subgroup analysis of patients with CTD-PAH and SSc-PAH [184]. Hemodynamics, RV structure and function, and overall functional status were significantly improved in SSc-PAH patient treated with the upfront combination [185]. Investigations of several newer treatments for PAH, such as riociguat, macitentan, and selexipag, have included significant proportions of patients on background therapies and have demonstrated added improvements in exercise capacity, functional class, and time to clinical worsening [155, 166, 181]. These studies have fueled the impetus to include recommendations for combination therapy in contemporary treatment guidelines [16, 186].
\nMedical therapy can improve activity tolerance, hemodynamics, and quality of life and can even improve survival prognosis; however, in some cases PAH will progress even with aggressive medical therapy. Nonmedical options may include balloon atrial septostomy and/or lung transplantation. Atrial septostomy may be beneficial in FC 4 patients with right heart failure or severe syncopal symptoms who are progressing on maximal medical therapy [187]. Atrial septostomy is also a consideration as a bridge to lung transplantation when medical therapy fails. An interatrial right-to-left shunt may decompress the right heart chambers and ultimately improve oxygen transport despite an observed oxyhemoglobin desaturation [188]. Atrial septostomy is not recommended in end-stage patients with mean RAP >20 mmHg and a resting room air saturation below 85% [187, 188]. Lung transplantation is also an option for patients with end-stage SSc-PAH failing medical therapy. In some centers, patients with SSc-PAH may not be offered lung transplantation due to the risk of aspiration pneumonia related to esophageal disease. However, studies have shown that survival after lung transplantation is similar in patients with SSc-PAH and other transplant indications [189]. There has been increasing interest in stem cell therapy as a treatment option for PAH. Although animal models have shown some promise, stem cell therapy is not currently a viable option for treatment of human PAH [190].
\nThe poor survival prognosis associated with SSc-PAH and the availability of multiple disease-targeted treatment options have fostered the development of algorithms to guide the treatment decision process. Both the American College of Chest Physicians [186] and the Joint Task Force for the Diagnosis and Treatment of Pulmonary Hypertension of the ESC/ERS [16] have published guidelines to aid clinicians in the treatment of patients with PAH.
\nIn addition to a careful risk assessment, choice of appropriate disease-targeted agent, and close monitoring of treatment effect, there are severe supportive measures that apply in the management of the patient with SSc-PAH. Patients are not restricted from physical activity. Physical activity and supervised rehabilitation have been shown to improve exercise tolerance, reduce fatigue, and improve quality of life [191, 192]. The ESC/ERS Guidelines suggest that patients who are stable clinically should consider participation in a rehabilitation program at a center experienced with the management of PAH if possible [16]. Patients with SSc-PAH should be vaccinated against influenza and pneumococcal pneumonia. Pregnancy in patients with PAH is associated with a high mortality risk and should be avoided. If patients with PAH become pregnant, the high risk of complications and pregnancy termination should be discussed. Some PAH treatments cause fetal harm, and patients are required to undergo monthly pregnancy monitoring when using such therapies. Riociguat and the endothelin antagonists are teratogenic. Patients using these targeted therapies should be carefully counseled about the risk of fetal harm and instructed to use at least two barrier methods of contraception while using these agents. In the event patients do become pregnant, they may continue PAH therapies that are not considered fetal toxic, such as the prostanoids, plan an elective delivery, and work closely with a high-risk obstetrical team and experienced pulmonary hypertension specialist throughout the pregnancy [193]. Patients with PAH are often overwhelmed by the physical limitations, financial burden, and social impact associated with PAH [194]. Screening for depression is helpful in identifying patients who could benefit from referral to appropriate services in the community where help is available to ease the psychosocial burden of this disease. Genetic counseling may be appropriate for select patients [195]. It is often helpful for the affected patient and at-risk family members to understand their mutation status in order to plan for the future. Genetic testing and counseling should involve a multidisciplinary team including pulmonary hypertension specialists, genetic counselors, geneticists, psychologists, and nurses. Elective surgery is not contraindicated but does carry an increased risk to the PAH patient. Patients with significantly impaired RV function are at highest risk and should undergo careful preoperative assessment [196, 197, 198]. Epidural anesthesia may be better tolerated [199]. Patients using oral therapies may require transition to an intravenous or inhaled form of therapy until able to take oral medications postoperatively.
\nPulmonary arterial hypertension is a leading cause of death in patients with systemic sclerosis. While the pathogenesis of PAH in the patient with systemic sclerosis bears resemblance to that of idiopathic PAH, there are distinct differences in genetic predisposition, role of inflammation and autoantibodies, and pathologic manifestations of disease. Early detection is essential in preventing early demise from SSc-PAH. Several algorithms have been suggested for screening SSc patients for PAH. In general, it is recommended that annual screening with biomarkers, PFTs, and echocardiography be considered in any patient with systemic sclerosis, even if they are asymptomatic. There are a number of medical therapies available which have demonstrated benefit in SSc-PAH, as well as iPAH. The importance of regular monitoring and repeat risk assessment cannot be underemphasized. Lung transplantation may be an option for those patients who progress on maximal medical therapy. While the prognosis for SSc-PAH has certainly improved over the past two decades, continued research into the mechanisms of disease and development of new treatments will ensure further improvements in quality of life and survival in the future.
\nAuthors are listed below with their open access chapters linked via author name:
",metaTitle:"IntechOpen authors on the Global Highly Cited Researchers 2018 list",metaDescription:null,metaKeywords:null,canonicalURL:null,contentRaw:'[{"type":"htmlEditorComponent","content":"New for 2018 (alphabetically by surname).
\\n\\n\\n\\n\\n\\n\\n\\n\\n\\nJocelyn Chanussot (chapter to be published soon...)
\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\nYuekun Lai
\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\n\\nPrevious years (alphabetically by surname)
\\n\\nAbdul Latif Ahmad 2016-18
\\n\\nKhalil Amine 2017, 2018
\\n\\nEwan Birney 2015-18
\\n\\nFrede Blaabjerg 2015-18
\\n\\nGang Chen 2016-18
\\n\\nJunhong Chen 2017, 2018
\\n\\nZhigang Chen 2016, 2018
\\n\\nMyung-Haing Cho 2016, 2018
\\n\\nMark Connors 2015-18
\\n\\nCyrus Cooper 2017, 2018
\\n\\nLiming Dai 2015-18
\\n\\nWeihua Deng 2017, 2018
\\n\\nVincenzo Fogliano 2017, 2018
\\n\\nRon de Graaf 2014-18
\\n\\nHarald Haas 2017, 2018
\\n\\nFrancisco Herrera 2017, 2018
\\n\\nJaakko Kangasjärvi 2015-18
\\n\\nHamid Reza Karimi 2016-18
\\n\\nJunji Kido 2014-18
\\n\\nJose Luiszamorano 2015-18
\\n\\nYiqi Luo 2016-18
\\n\\nJoachim Maier 2014-18
\\n\\nAndrea Natale 2017, 2018
\\n\\nAlberto Mantovani 2014-18
\\n\\nMarjan Mernik 2017, 2018
\\n\\nSandra Orchard 2014, 2016-18
\\n\\nMohamed Oukka 2016-18
\\n\\nBiswajeet Pradhan 2016-18
\\n\\nDirk Raes 2017, 2018
\\n\\nUlrike Ravens-Sieberer 2016-18
\\n\\nYexiang Tong 2017, 2018
\\n\\nJim Van Os 2015-18
\\n\\nLong Wang 2017, 2018
\\n\\nFei Wei 2016-18
\\n\\nIoannis Xenarios 2017, 2018
\\n\\nQi Xie 2016-18
\\n\\nXin-She Yang 2017, 2018
\\n\\nYulong Yin 2015, 2017, 2018
\\n"}]'},components:[{type:"htmlEditorComponent",content:'New for 2018 (alphabetically by surname).
\n\n\n\n\n\n\n\n\n\nJocelyn Chanussot (chapter to be published soon...)
\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\nYuekun Lai
\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\nPrevious years (alphabetically by surname)
\n\nAbdul Latif Ahmad 2016-18
\n\nKhalil Amine 2017, 2018
\n\nEwan Birney 2015-18
\n\nFrede Blaabjerg 2015-18
\n\nGang Chen 2016-18
\n\nJunhong Chen 2017, 2018
\n\nZhigang Chen 2016, 2018
\n\nMyung-Haing Cho 2016, 2018
\n\nMark Connors 2015-18
\n\nCyrus Cooper 2017, 2018
\n\nLiming Dai 2015-18
\n\nWeihua Deng 2017, 2018
\n\nVincenzo Fogliano 2017, 2018
\n\nRon de Graaf 2014-18
\n\nHarald Haas 2017, 2018
\n\nFrancisco Herrera 2017, 2018
\n\nJaakko Kangasjärvi 2015-18
\n\nHamid Reza Karimi 2016-18
\n\nJunji Kido 2014-18
\n\nJose Luiszamorano 2015-18
\n\nYiqi Luo 2016-18
\n\nJoachim Maier 2014-18
\n\nAndrea Natale 2017, 2018
\n\nAlberto Mantovani 2014-18
\n\nMarjan Mernik 2017, 2018
\n\nSandra Orchard 2014, 2016-18
\n\nMohamed Oukka 2016-18
\n\nBiswajeet Pradhan 2016-18
\n\nDirk Raes 2017, 2018
\n\nUlrike Ravens-Sieberer 2016-18
\n\nYexiang Tong 2017, 2018
\n\nJim Van Os 2015-18
\n\nLong Wang 2017, 2018
\n\nFei Wei 2016-18
\n\nIoannis Xenarios 2017, 2018
\n\nQi Xie 2016-18
\n\nXin-She Yang 2017, 2018
\n\nYulong Yin 2015, 2017, 2018
\n'}]},successStories:{items:[]},authorsAndEditors:{filterParams:{sort:"featured,name"},profiles:[{id:"6700",title:"Dr.",name:"Abbass A.",middleName:null,surname:"Hashim",slug:"abbass-a.-hashim",fullName:"Abbass A. Hashim",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/6700/images/1864_n.jpg",biography:"Currently I am carrying out research in several areas of interest, mainly covering work on chemical and bio-sensors, semiconductor thin film device fabrication and characterisation.\nAt the moment I have very strong interest in radiation environmental pollution and bacteriology treatment. The teams of researchers are working very hard to bring novel results in this field. I am also a member of the team in charge for the supervision of Ph.D. students in the fields of development of silicon based planar waveguide sensor devices, study of inelastic electron tunnelling in planar tunnelling nanostructures for sensing applications and development of organotellurium(IV) compounds for semiconductor applications. I am a specialist in data analysis techniques and nanosurface structure. I have served as the editor for many books, been a member of the editorial board in science journals, have published many papers and hold many patents.",institutionString:null,institution:{name:"Sheffield Hallam University",country:{name:"United Kingdom"}}},{id:"54525",title:"Prof.",name:"Abdul Latif",middleName:null,surname:"Ahmad",slug:"abdul-latif-ahmad",fullName:"Abdul Latif Ahmad",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:null},{id:"20567",title:"Prof.",name:"Ado",middleName:null,surname:"Jorio",slug:"ado-jorio",fullName:"Ado Jorio",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Universidade Federal de Minas Gerais",country:{name:"Brazil"}}},{id:"47940",title:"Dr.",name:"Alberto",middleName:null,surname:"Mantovani",slug:"alberto-mantovani",fullName:"Alberto Mantovani",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:null},{id:"12392",title:"Mr.",name:"Alex",middleName:null,surname:"Lazinica",slug:"alex-lazinica",fullName:"Alex Lazinica",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/12392/images/7282_n.png",biography:"Alex Lazinica is the founder and CEO of IntechOpen. After obtaining a Master's degree in Mechanical Engineering, he continued his PhD studies in Robotics at the Vienna University of Technology. Here he worked as a robotic researcher with the university's Intelligent Manufacturing Systems Group as well as a guest researcher at various European universities, including the Swiss Federal Institute of Technology Lausanne (EPFL). During this time he published more than 20 scientific papers, gave presentations, served as a reviewer for major robotic journals and conferences and most importantly he co-founded and built the International Journal of Advanced Robotic Systems- world's first Open Access journal in the field of robotics. Starting this journal was a pivotal point in his career, since it was a pathway to founding IntechOpen - Open Access publisher focused on addressing academic researchers needs. Alex is a personification of IntechOpen key values being trusted, open and entrepreneurial. Today his focus is on defining the growth and development strategy for the company.",institutionString:null,institution:{name:"TU Wien",country:{name:"Austria"}}},{id:"19816",title:"Prof.",name:"Alexander",middleName:null,surname:"Kokorin",slug:"alexander-kokorin",fullName:"Alexander Kokorin",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/19816/images/1607_n.jpg",biography:"Alexander I. Kokorin: born: 1947, Moscow; DSc., PhD; Principal Research Fellow (Research Professor) of Department of Kinetics and Catalysis, N. Semenov Institute of Chemical Physics, Russian Academy of Sciences, Moscow.\nArea of research interests: physical chemistry of complex-organized molecular and nanosized systems, including polymer-metal complexes; the surface of doped oxide semiconductors. He is an expert in structural, absorptive, catalytic and photocatalytic properties, in structural organization and dynamic features of ionic liquids, in magnetic interactions between paramagnetic centers. The author or co-author of 3 books, over 200 articles and reviews in scientific journals and books. He is an actual member of the International EPR/ESR Society, European Society on Quantum Solar Energy Conversion, Moscow House of Scientists, of the Board of Moscow Physical Society.",institutionString:null,institution:null},{id:"62389",title:"PhD.",name:"Ali Demir",middleName:null,surname:"Sezer",slug:"ali-demir-sezer",fullName:"Ali Demir Sezer",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/62389/images/3413_n.jpg",biography:"Dr. Ali Demir Sezer has a Ph.D. from Pharmaceutical Biotechnology at the Faculty of Pharmacy, University of Marmara (Turkey). He is the member of many Pharmaceutical Associations and acts as a reviewer of scientific journals and European projects under different research areas such as: drug delivery systems, nanotechnology and pharmaceutical biotechnology. Dr. Sezer is the author of many scientific publications in peer-reviewed journals and poster communications. Focus of his research activity is drug delivery, physico-chemical characterization and biological evaluation of biopolymers micro and nanoparticles as modified drug delivery system, and colloidal drug carriers (liposomes, nanoparticles etc.).",institutionString:null,institution:{name:"Marmara University",country:{name:"Turkey"}}},{id:"61051",title:"Prof.",name:"Andrea",middleName:null,surname:"Natale",slug:"andrea-natale",fullName:"Andrea Natale",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:null},{id:"100762",title:"Prof.",name:"Andrea",middleName:null,surname:"Natale",slug:"andrea-natale",fullName:"Andrea Natale",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"St David's Medical Center",country:{name:"United States of America"}}},{id:"107416",title:"Dr.",name:"Andrea",middleName:null,surname:"Natale",slug:"andrea-natale",fullName:"Andrea Natale",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Texas Cardiac Arrhythmia",country:{name:"United States of America"}}},{id:"64434",title:"Dr.",name:"Angkoon",middleName:null,surname:"Phinyomark",slug:"angkoon-phinyomark",fullName:"Angkoon Phinyomark",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/64434/images/2619_n.jpg",biography:"My name is Angkoon Phinyomark. I received a B.Eng. degree in Computer Engineering with First Class Honors in 2008 from Prince of Songkla University, Songkhla, Thailand, where I received a Ph.D. degree in Electrical Engineering. My research interests are primarily in the area of biomedical signal processing and classification notably EMG (electromyography signal), EOG (electrooculography signal), and EEG (electroencephalography signal), image analysis notably breast cancer analysis and optical coherence tomography, and rehabilitation engineering. I became a student member of IEEE in 2008. During October 2011-March 2012, I had worked at School of Computer Science and Electronic Engineering, University of Essex, Colchester, Essex, United Kingdom. In addition, during a B.Eng. I had been a visiting research student at Faculty of Computer Science, University of Murcia, Murcia, Spain for three months.\n\nI have published over 40 papers during 5 years in refereed journals, books, and conference proceedings in the areas of electro-physiological signals processing and classification, notably EMG and EOG signals, fractal analysis, wavelet analysis, texture analysis, feature extraction and machine learning algorithms, and assistive and rehabilitative devices. I have several computer programming language certificates, i.e. Sun Certified Programmer for the Java 2 Platform 1.4 (SCJP), Microsoft Certified Professional Developer, Web Developer (MCPD), Microsoft Certified Technology Specialist, .NET Framework 2.0 Web (MCTS). I am a Reviewer for several refereed journals and international conferences, such as IEEE Transactions on Biomedical Engineering, IEEE Transactions on Industrial Electronics, Optic Letters, Measurement Science Review, and also a member of the International Advisory Committee for 2012 IEEE Business Engineering and Industrial Applications and 2012 IEEE Symposium on Business, Engineering and Industrial Applications.",institutionString:null,institution:{name:"Joseph Fourier University",country:{name:"France"}}},{id:"55578",title:"Dr.",name:"Antonio",middleName:null,surname:"Jurado-Navas",slug:"antonio-jurado-navas",fullName:"Antonio Jurado-Navas",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/55578/images/4574_n.png",biography:"Antonio Jurado-Navas received the M.S. degree (2002) and the Ph.D. degree (2009) in Telecommunication Engineering, both from the University of Málaga (Spain). He first worked as a consultant at Vodafone-Spain. From 2004 to 2011, he was a Research Assistant with the Communications Engineering Department at the University of Málaga. In 2011, he became an Assistant Professor in the same department. From 2012 to 2015, he was with Ericsson Spain, where he was working on geo-location\ntools for third generation mobile networks. Since 2015, he is a Marie-Curie fellow at the Denmark Technical University. His current research interests include the areas of mobile communication systems and channel modeling in addition to atmospheric optical communications, adaptive optics and statistics",institutionString:null,institution:{name:"University of Malaga",country:{name:"Spain"}}}],filtersByRegion:[{group:"region",caption:"North America",value:1,count:5319},{group:"region",caption:"Middle and South America",value:2,count:4828},{group:"region",caption:"Africa",value:3,count:1471},{group:"region",caption:"Asia",value:4,count:9370},{group:"region",caption:"Australia and Oceania",value:5,count:837},{group:"region",caption:"Europe",value:6,count:14788}],offset:12,limit:12,total:108345},chapterEmbeded:{data:{}},editorApplication:{success:null,errors:{}},ofsBooks:{filterParams:{topicId:"521"},books:[{type:"book",id:"8321",title:"Digital Forensic Science",subtitle:null,isOpenForSubmission:!0,hash:"15540cde4d2e598046f7a520f6c4b107",slug:null,bookSignature:"Dr. B Suresh Shetty, Dr. Pavanchand Shetty and Dr. Adithi Shetty",coverURL:"https://cdn.intechopen.com/books/images_new/8321.jpg",editedByType:null,editors:[{id:"70242",title:"Dr.",name:"B Suresh",surname:"Shetty",slug:"b-suresh-shetty",fullName:"B Suresh Shetty"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}}],filtersByTopic:[{group:"topic",caption:"Agricultural and Biological Sciences",value:5,count:34},{group:"topic",caption:"Biochemistry, Genetics and Molecular Biology",value:6,count:33},{group:"topic",caption:"Business, Management and Economics",value:7,count:10},{group:"topic",caption:"Chemistry",value:8,count:30},{group:"topic",caption:"Computer and Information Science",value:9,count:25},{group:"topic",caption:"Earth and Planetary Sciences",value:10,count:15},{group:"topic",caption:"Engineering",value:11,count:71},{group:"topic",caption:"Environmental Sciences",value:12,count:13},{group:"topic",caption:"Immunology and Microbiology",value:13,count:3},{group:"topic",caption:"Materials Science",value:14,count:38},{group:"topic",caption:"Mathematics",value:15,count:14},{group:"topic",caption:"Medicine",value:16,count:136},{group:"topic",caption:"Nanotechnology and Nanomaterials",value:17,count:6},{group:"topic",caption:"Neuroscience",value:18,count:6},{group:"topic",caption:"Pharmacology, Toxicology and Pharmaceutical Science",value:19,count:9},{group:"topic",caption:"Physics",value:20,count:20},{group:"topic",caption:"Psychology",value:21,count:2},{group:"topic",caption:"Robotics",value:22,count:6},{group:"topic",caption:"Social Sciences",value:23,count:13},{group:"topic",caption:"Technology",value:24,count:10},{group:"topic",caption:"Veterinary Medicine and Science",value:25,count:3},{group:"topic",caption:"Genesiology",value:300,count:1},{group:"topic",caption:"Machine Learning and Data Mining",value:521,count:1},{group:"topic",caption:"Intelligent System",value:535,count:1}],offset:12,limit:12,total:1},popularBooks:{featuredBooks:[{type:"book",id:"7878",title:"Advances in Extracorporeal Membrane Oxygenation",subtitle:"Volume 3",isOpenForSubmission:!1,hash:"f95bf990273d08098a00f9a1c2403cbe",slug:"advances-in-extracorporeal-membrane-oxygenation-volume-3",bookSignature:"Michael S. Firstenberg",coverURL:"https://cdn.intechopen.com/books/images_new/7878.jpg",editors:[{id:"64343",title:null,name:"Michael S.",middleName:"S",surname:"Firstenberg",slug:"michael-s.-firstenberg",fullName:"Michael S. Firstenberg"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"8299",title:"Timber Buildings and Sustainability",subtitle:null,isOpenForSubmission:!1,hash:"bccf2891cec38ed041724131aa34c25a",slug:"timber-buildings-and-sustainability",bookSignature:"Giovanna Concu",coverURL:"https://cdn.intechopen.com/books/images_new/8299.jpg",editors:[{id:"108709",title:"Dr.",name:"Giovanna",middleName:null,surname:"Concu",slug:"giovanna-concu",fullName:"Giovanna Concu"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7614",title:"Fourier Transforms",subtitle:"Century of Digitalization and Increasing Expectations",isOpenForSubmission:!1,hash:"ff3501657ae983a3b42fef1f7058ac91",slug:"fourier-transforms-century-of-digitalization-and-increasing-expectations",bookSignature:"Goran S. Nikoli? and Dragana Z. Markovi?-Nikoli?",coverURL:"https://cdn.intechopen.com/books/images_new/7614.jpg",editors:[{id:"23261",title:"Prof.",name:"Goran",middleName:"S.",surname:"Nikolic",slug:"goran-nikolic",fullName:"Goran Nikolic"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7062",title:"Rhinosinusitis",subtitle:null,isOpenForSubmission:!1,hash:"14ed95e155b1e57a61827ca30b579d09",slug:"rhinosinusitis",bookSignature:"Balwant Singh Gendeh and Mirjana Turkalj",coverURL:"https://cdn.intechopen.com/books/images_new/7062.jpg",editors:[{id:"67669",title:"Prof.",name:"Balwant Singh",middleName:null,surname:"Gendeh",slug:"balwant-singh-gendeh",fullName:"Balwant Singh Gendeh"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7087",title:"Tendons",subtitle:null,isOpenForSubmission:!1,hash:"786abac0445c102d1399a1e727a2db7f",slug:"tendons",bookSignature:"Hasan Sözen",coverURL:"https://cdn.intechopen.com/books/images_new/7087.jpg",editors:[{id:"161402",title:"Dr.",name:"Hasan",middleName:null,surname:"Sözen",slug:"hasan-sozen",fullName:"Hasan Sözen"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7955",title:"Advances in Hematologic Malignancies",subtitle:null,isOpenForSubmission:!1,hash:"59ca1b09447fab4717a93e099f646d28",slug:"advances-in-hematologic-malignancies",bookSignature:"Gamal Abdul Hamid",coverURL:"https://cdn.intechopen.com/books/images_new/7955.jpg",editors:[{id:"36487",title:"Prof.",name:"Gamal",middleName:null,surname:"Abdul Hamid",slug:"gamal-abdul-hamid",fullName:"Gamal Abdul Hamid"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7701",title:"Assistive and Rehabilitation Engineering",subtitle:null,isOpenForSubmission:!1,hash:"4191b744b8af3b17d9a80026dcb0617f",slug:"assistive-and-rehabilitation-engineering",bookSignature:"Yves Rybarczyk",coverURL:"https://cdn.intechopen.com/books/images_new/7701.jpg",editors:[{id:"72920",title:"Prof.",name:"Yves",middleName:"Philippe",surname:"Rybarczyk",slug:"yves-rybarczyk",fullName:"Yves Rybarczyk"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7837",title:"Geriatric Medicine and Gerontology",subtitle:null,isOpenForSubmission:!1,hash:"e277d005b23536bcd9f8550046101979",slug:"geriatric-medicine-and-gerontology",bookSignature:"Edward T. Zawada Jr.",coverURL:"https://cdn.intechopen.com/books/images_new/7837.jpg",editors:[{id:"16344",title:"Dr.",name:"Edward T.",middleName:null,surname:"Zawada Jr.",slug:"edward-t.-zawada-jr.",fullName:"Edward T. Zawada Jr."}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7123",title:"Current Topics in Neglected Tropical Diseases",subtitle:null,isOpenForSubmission:!1,hash:"61c627da05b2ace83056d11357bdf361",slug:"current-topics-in-neglected-tropical-diseases",bookSignature:"Alfonso J. Rodriguez-Morales",coverURL:"https://cdn.intechopen.com/books/images_new/7123.jpg",editors:[{id:"131400",title:"Dr.",name:"Alfonso J.",middleName:null,surname:"Rodriguez-Morales",slug:"alfonso-j.-rodriguez-morales",fullName:"Alfonso J. Rodriguez-Morales"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7610",title:"Renewable and Sustainable Composites",subtitle:null,isOpenForSubmission:!1,hash:"c2de26c3d329c54f093dc3f05417500a",slug:"renewable-and-sustainable-composites",bookSignature:"António B. Pereira and Fábio A. O. Fernandes",coverURL:"https://cdn.intechopen.com/books/images_new/7610.jpg",editors:[{id:"211131",title:"Prof.",name:"António",middleName:"Bastos",surname:"Pereira",slug:"antonio-pereira",fullName:"António Pereira"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"8416",title:"Non-Equilibrium Particle Dynamics",subtitle:null,isOpenForSubmission:!1,hash:"2c3add7639dcd1cb442cb4313ea64e3a",slug:"non-equilibrium-particle-dynamics",bookSignature:"Albert S. Kim",coverURL:"https://cdn.intechopen.com/books/images_new/8416.jpg",editors:[{id:"21045",title:"Prof.",name:"Albert S.",middleName:null,surname:"Kim",slug:"albert-s.-kim",fullName:"Albert S. Kim"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"8008",title:"Antioxidants",subtitle:null,isOpenForSubmission:!1,hash:"76361b4061e830906267933c1c670027",slug:"antioxidants",bookSignature:"Emad Shalaby",coverURL:"https://cdn.intechopen.com/books/images_new/8008.jpg",editors:[{id:"63600",title:"Prof.",name:"Emad",middleName:null,surname:"Shalaby",slug:"emad-shalaby",fullName:"Emad Shalaby"}],productType:{id:"1",chapterContentType:"chapter"}}],offset:12,limit:12,total:4398},hotBookTopics:{hotBooks:[],offset:0,limit:12,total:null},publish:{},publishingProposal:{success:null,errors:{}},books:{featuredBooks:[{type:"book",id:"7878",title:"Advances in Extracorporeal Membrane Oxygenation",subtitle:"Volume 3",isOpenForSubmission:!1,hash:"f95bf990273d08098a00f9a1c2403cbe",slug:"advances-in-extracorporeal-membrane-oxygenation-volume-3",bookSignature:"Michael S. Firstenberg",coverURL:"https://cdn.intechopen.com/books/images_new/7878.jpg",editors:[{id:"64343",title:null,name:"Michael S.",middleName:"S",surname:"Firstenberg",slug:"michael-s.-firstenberg",fullName:"Michael S. Firstenberg"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"8299",title:"Timber Buildings and Sustainability",subtitle:null,isOpenForSubmission:!1,hash:"bccf2891cec38ed041724131aa34c25a",slug:"timber-buildings-and-sustainability",bookSignature:"Giovanna Concu",coverURL:"https://cdn.intechopen.com/books/images_new/8299.jpg",editors:[{id:"108709",title:"Dr.",name:"Giovanna",middleName:null,surname:"Concu",slug:"giovanna-concu",fullName:"Giovanna Concu"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7614",title:"Fourier Transforms",subtitle:"Century of Digitalization and Increasing Expectations",isOpenForSubmission:!1,hash:"ff3501657ae983a3b42fef1f7058ac91",slug:"fourier-transforms-century-of-digitalization-and-increasing-expectations",bookSignature:"Goran S. Nikoli? and Dragana Z. Markovi?-Nikoli?",coverURL:"https://cdn.intechopen.com/books/images_new/7614.jpg",editors:[{id:"23261",title:"Prof.",name:"Goran",middleName:"S.",surname:"Nikolic",slug:"goran-nikolic",fullName:"Goran Nikolic"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7062",title:"Rhinosinusitis",subtitle:null,isOpenForSubmission:!1,hash:"14ed95e155b1e57a61827ca30b579d09",slug:"rhinosinusitis",bookSignature:"Balwant Singh Gendeh and Mirjana Turkalj",coverURL:"https://cdn.intechopen.com/books/images_new/7062.jpg",editors:[{id:"67669",title:"Prof.",name:"Balwant Singh",middleName:null,surname:"Gendeh",slug:"balwant-singh-gendeh",fullName:"Balwant Singh Gendeh"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7087",title:"Tendons",subtitle:null,isOpenForSubmission:!1,hash:"786abac0445c102d1399a1e727a2db7f",slug:"tendons",bookSignature:"Hasan Sözen",coverURL:"https://cdn.intechopen.com/books/images_new/7087.jpg",editors:[{id:"161402",title:"Dr.",name:"Hasan",middleName:null,surname:"Sözen",slug:"hasan-sozen",fullName:"Hasan Sözen"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7955",title:"Advances in Hematologic Malignancies",subtitle:null,isOpenForSubmission:!1,hash:"59ca1b09447fab4717a93e099f646d28",slug:"advances-in-hematologic-malignancies",bookSignature:"Gamal Abdul Hamid",coverURL:"https://cdn.intechopen.com/books/images_new/7955.jpg",editors:[{id:"36487",title:"Prof.",name:"Gamal",middleName:null,surname:"Abdul Hamid",slug:"gamal-abdul-hamid",fullName:"Gamal Abdul Hamid"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7701",title:"Assistive and Rehabilitation Engineering",subtitle:null,isOpenForSubmission:!1,hash:"4191b744b8af3b17d9a80026dcb0617f",slug:"assistive-and-rehabilitation-engineering",bookSignature:"Yves Rybarczyk",coverURL:"https://cdn.intechopen.com/books/images_new/7701.jpg",editors:[{id:"72920",title:"Prof.",name:"Yves",middleName:"Philippe",surname:"Rybarczyk",slug:"yves-rybarczyk",fullName:"Yves Rybarczyk"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7837",title:"Geriatric Medicine and Gerontology",subtitle:null,isOpenForSubmission:!1,hash:"e277d005b23536bcd9f8550046101979",slug:"geriatric-medicine-and-gerontology",bookSignature:"Edward T. Zawada Jr.",coverURL:"https://cdn.intechopen.com/books/images_new/7837.jpg",editors:[{id:"16344",title:"Dr.",name:"Edward T.",middleName:null,surname:"Zawada Jr.",slug:"edward-t.-zawada-jr.",fullName:"Edward T. Zawada Jr."}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7123",title:"Current Topics in Neglected Tropical Diseases",subtitle:null,isOpenForSubmission:!1,hash:"61c627da05b2ace83056d11357bdf361",slug:"current-topics-in-neglected-tropical-diseases",bookSignature:"Alfonso J. Rodriguez-Morales",coverURL:"https://cdn.intechopen.com/books/images_new/7123.jpg",editors:[{id:"131400",title:"Dr.",name:"Alfonso J.",middleName:null,surname:"Rodriguez-Morales",slug:"alfonso-j.-rodriguez-morales",fullName:"Alfonso J. Rodriguez-Morales"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7610",title:"Renewable and Sustainable Composites",subtitle:null,isOpenForSubmission:!1,hash:"c2de26c3d329c54f093dc3f05417500a",slug:"renewable-and-sustainable-composites",bookSignature:"António B. Pereira and Fábio A. O. Fernandes",coverURL:"https://cdn.intechopen.com/books/images_new/7610.jpg",editors:[{id:"211131",title:"Prof.",name:"António",middleName:"Bastos",surname:"Pereira",slug:"antonio-pereira",fullName:"António Pereira"}],productType:{id:"1",chapterContentType:"chapter"}}],latestBooks:[{type:"book",id:"7698",title:"Educational Psychology",subtitle:"Between Certitudes and Uncertainties",isOpenForSubmission:!1,hash:"740943e2d029253e777150e98ebe2f0d",slug:"educational-psychology-between-certitudes-and-uncertainties",bookSignature:"Victori?a Trif",coverURL:"https://cdn.intechopen.com/books/images_new/7698.jpg",editedByType:"Edited by",editors:[{id:"201656",title:"Ph.D.",name:"Victorița",middleName:null,surname:"Trif",slug:"victorita-trif",fullName:"Victorița Trif"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"8747",title:"Asphalt and Asphalt Mixtures",subtitle:null,isOpenForSubmission:!1,hash:"6083f7c9881029f1e033a1e512af7e20",slug:"asphalt-and-asphalt-mixtures",bookSignature:"Haitao Zhang",coverURL:"https://cdn.intechopen.com/books/images_new/8747.jpg",editedByType:"Edited by",editors:[{id:"260604",title:"Prof.",name:"Haitao",middleName:null,surname:"Zhang",slug:"haitao-zhang",fullName:"Haitao Zhang"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"8516",title:"Metacognition in Learning",subtitle:null,isOpenForSubmission:!1,hash:"5fa6eaad7b509b8b7ec5124d79e5f605",slug:"metacognition-in-learning",bookSignature:"Nosisi Feza",coverURL:"https://cdn.intechopen.com/books/images_new/8516.jpg",editedByType:"Edited by",editors:[{id:"261665",title:"Prof.",name:"Nosisi",middleName:"N.",surname:"Feza",slug:"nosisi-feza",fullName:"Nosisi Feza"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"7000",title:"Legume Crops",subtitle:"Characterization and Breeding for Improved Food Security",isOpenForSubmission:!1,hash:"4d0f73bf883bbb984cc2feef1259a9a7",slug:"legume-crops-characterization-and-breeding-for-improved-food-security",bookSignature:"Mohamed Ahmed El-Esawi",coverURL:"https://cdn.intechopen.com/books/images_new/7000.jpg",editedByType:"Edited by",editors:[{id:"191770",title:"Dr.",name:"Mohamed A.",middleName:null,surname:"El-Esawi",slug:"mohamed-a.-el-esawi",fullName:"Mohamed A. El-Esawi"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"8292",title:"Oral Health by Using Probiotic Products",subtitle:null,isOpenForSubmission:!1,hash:"327e750e83634800ace02fe62607c21e",slug:"oral-health-by-using-probiotic-products",bookSignature:"Razzagh Mahmoudi",coverURL:"https://cdn.intechopen.com/books/images_new/8292.jpg",editedByType:"Edited by",editors:[{id:"245925",title:"Dr.",name:"Razzagh",middleName:null,surname:"Mahmoudi",slug:"razzagh-mahmoudi",fullName:"Razzagh Mahmoudi"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"8323",title:"Traditional and Complementary Medicine",subtitle:null,isOpenForSubmission:!1,hash:"60eadb1783d9bba245687adf284d4871",slug:"traditional-and-complementary-medicine",bookSignature:"Cengiz Mordeniz",coverURL:"https://cdn.intechopen.com/books/images_new/8323.jpg",editedByType:"Edited by",editors:[{id:"214664",title:"Associate Prof.",name:"Cengiz",middleName:null,surname:"Mordeniz",slug:"cengiz-mordeniz",fullName:"Cengiz Mordeniz"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"8347",title:"Computer Architecture in Industrial, Biomechanical and Biomedical Engineering",subtitle:null,isOpenForSubmission:!1,hash:"3d7024a8d7d8afed093c9c79ec31f15a",slug:"computer-architecture-in-industrial-biomechanical-and-biomedical-engineering",bookSignature:"Lulu Wang and Liandong Yu",coverURL:"https://cdn.intechopen.com/books/images_new/8347.jpg",editedByType:"Edited by",editors:[{id:"257388",title:"Dr.",name:"Lulu",middleName:null,surname:"Wang",slug:"lulu-wang",fullName:"Lulu Wang"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"7583",title:"Advanced Computational Fluid Dynamics for Emerging Engineering Processes",subtitle:"Eulerian vs. Lagrangian",isOpenForSubmission:!1,hash:"896509fa2e7e659811bffd0f9779ca9d",slug:"advanced-computational-fluid-dynamics-for-emerging-engineering-processes-eulerian-vs-lagrangian",bookSignature:"Albert S. Kim",coverURL:"https://cdn.intechopen.com/books/images_new/7583.jpg",editedByType:"Edited by",editors:[{id:"21045",title:"Prof.",name:"Albert S.",middleName:null,surname:"Kim",slug:"albert-s.-kim",fullName:"Albert S. Kim"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"7839",title:"Malaria",subtitle:null,isOpenForSubmission:!1,hash:"91cde4582ead884cb0f355a19b67cd56",slug:"malaria",bookSignature:"Fyson H. Kasenga",coverURL:"https://cdn.intechopen.com/books/images_new/7839.jpg",editedByType:"Edited by",editors:[{id:"86725",title:"Dr.",name:"Fyson",middleName:"Hanania",surname:"Kasenga",slug:"fyson-kasenga",fullName:"Fyson Kasenga"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"7093",title:"Pneumothorax",subtitle:null,isOpenForSubmission:!1,hash:"0b1fdb8bb0448f48c2f234753898f3f8",slug:"pneumothorax",bookSignature:"Khalid Amer",coverURL:"https://cdn.intechopen.com/books/images_new/7093.jpg",editedByType:"Edited by",editors:[{id:"63412",title:"Dr.",name:"Khalid",middleName:null,surname:"Amer",slug:"khalid-amer",fullName:"Khalid Amer"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}}]},subject:{topic:{id:"463",title:"International Trade",slug:"international-trade",parent:{title:"International Economics",slug:"international-economics"},numberOfBooks:3,numberOfAuthorsAndEditors:44,numberOfWosCitations:8,numberOfCrossrefCitations:12,numberOfDimensionsCitations:25,videoUrl:null,fallbackUrl:null,description:null},booksByTopicFilter:{topicSlug:"international-trade",sort:"-publishedDate",limit:12,offset:0},booksByTopicCollection:[{type:"book",id:"6487",title:"Trade and Global Market",subtitle:null,isOpenForSubmission:!1,hash:"7f1afebc7552003672f0c62b354538be",slug:"trade-and-global-market",bookSignature:"Vito Bobek",coverURL:"https://cdn.intechopen.com/books/images_new/6487.jpg",editedByType:"Edited by",editors:[{id:"128342",title:"Prof.",name:"Vito",middleName:null,surname:"Bobek",slug:"vito-bobek",fullName:"Vito Bobek"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"5492",title:"International Trade",subtitle:"On the Brink of Change",isOpenForSubmission:!1,hash:"f64d7edf6aef7b32784cc01a18836699",slug:"international-trade-on-the-brink-of-change",bookSignature:"Anita Macek",coverURL:"https://cdn.intechopen.com/books/images_new/5492.jpg",editedByType:"Edited by",editors:[{id:"142587",title:"Dr.",name:"Anita",middleName:null,surname:"Maček",slug:"anita-macek",fullName:"Anita Maček"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"2355",title:"International Trade from Economic and Policy Perspective",subtitle:null,isOpenForSubmission:!1,hash:"8fe6804794ddc1a7f4202db20aed5985",slug:"international-trade-from-economic-and-policy-perspective",bookSignature:"Vito Bobek",coverURL:"https://cdn.intechopen.com/books/images_new/2355.jpg",editedByType:"Edited by",editors:[{id:"128342",title:"Prof.",name:"Vito",middleName:null,surname:"Bobek",slug:"vito-bobek",fullName:"Vito Bobek"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}}],booksByTopicTotal:3,mostCitedChapters:[{id:"38487",doi:"10.5772/48103",title:"A Perspective on Remanufacturing Business: Issues and Opportunities",slug:"a-perspective-on-remanufacturing-business-issues-and-opportunities",totalDownloads:3247,totalCrossrefCites:3,totalDimensionsCites:8,book:{slug:"international-trade-from-economic-and-policy-perspective",title:"International Trade from Economic and Policy Perspective",fullTitle:"International Trade from Economic and Policy Perspective"},signatures:"Mosè Gallo, Elpidio Romano and Liberatina Carmela Santillo",authors:[{id:"11826",title:"Prof.",name:"Elpidio",middleName:null,surname:"Romano",slug:"elpidio-romano",fullName:"Elpidio Romano"},{id:"11827",title:"Prof.",name:"Liberatina",middleName:null,surname:"Santillo",slug:"liberatina-santillo",fullName:"Liberatina Santillo"},{id:"138996",title:"PhD.",name:"Mosè",middleName:null,surname:"Gallo",slug:"mose-gallo",fullName:"Mosè Gallo"}]},{id:"38482",doi:"10.5772/48342",title:"A Comparative Analysis of the Economic Effects of Cross-Border Mergers and Acquisitions in European Countries",slug:"a-comparative-analysis-of-the-economic-effects-of-cross-border-mergers-and-acquisitions-in-european-",totalDownloads:2529,totalCrossrefCites:2,totalDimensionsCites:3,book:{slug:"international-trade-from-economic-and-policy-perspective",title:"International Trade from Economic and Policy Perspective",fullTitle:"International Trade from Economic and Policy Perspective"},signatures:"Anita Maček",authors:[{id:"142587",title:"Dr.",name:"Anita",middleName:null,surname:"Maček",slug:"anita-macek",fullName:"Anita Maček"}]},{id:"38485",doi:"10.5772/48151",title:"Transboundary Animal Diseases and International Trade",slug:"transboundary-animal-diseases-and-international-trade",totalDownloads:3112,totalCrossrefCites:1,totalDimensionsCites:3,book:{slug:"international-trade-from-economic-and-policy-perspective",title:"International Trade from Economic and Policy Perspective",fullTitle:"International Trade from Economic and Policy Perspective"},signatures:"Andrés Cartín-Rojas",authors:[{id:"139628",title:"Ms.",name:"Andrés",middleName:null,surname:"Cartín-Rojas",slug:"andres-cartin-rojas",fullName:"Andrés Cartín-Rojas"}]}],mostDownloadedChaptersLast30Days:[{id:"58969",title:"Corruption, Causes and Consequences",slug:"corruption-causes-and-consequences",totalDownloads:19070,totalCrossrefCites:1,totalDimensionsCites:1,book:{slug:"trade-and-global-market",title:"Trade and Global Market",fullTitle:"Trade and Global Market"},signatures:"Štefan Šumah",authors:[{id:"228073",title:"Mr.",name:"Stefan",middleName:null,surname:"Sumah",slug:"stefan-sumah",fullName:"Stefan Sumah"}]},{id:"61175",title:"Trade Openness and Economic Growth: Empirical Evidence from Transition Economies",slug:"trade-openness-and-economic-growth-empirical-evidence-from-transition-economies",totalDownloads:1556,totalCrossrefCites:0,totalDimensionsCites:1,book:{slug:"trade-and-global-market",title:"Trade and Global Market",fullTitle:"Trade and Global Market"},signatures:"Sabina Silajdzic and Eldin Mehic",authors:[{id:"233162",title:"Associate Prof.",name:"Sabina",middleName:null,surname:"Silajdzic",slug:"sabina-silajdzic",fullName:"Sabina Silajdzic"},{id:"233367",title:"Prof.",name:"Eldin",middleName:null,surname:"Mehic",slug:"eldin-mehic",fullName:"Eldin Mehic"}]},{id:"38488",title:"The Impact and Consequences of Tax Revenues' Components on Economic Indicators: Evidence from Panel Groups Data",slug:"the-impact-and-consequences-of-tax-revenues-components-on-economic-indicators-evidence-from-panel-gr",totalDownloads:6112,totalCrossrefCites:0,totalDimensionsCites:1,book:{slug:"international-trade-from-economic-and-policy-perspective",title:"International Trade from Economic and Policy Perspective",fullTitle:"International Trade from Economic and Policy Perspective"},signatures:"Taufik Abdul Hakim and Imbarine Bujang",authors:[{id:"143713",title:"Dr.",name:"Imbarine",middleName:null,surname:"Bujang",slug:"imbarine-bujang",fullName:"Imbarine Bujang"},{id:"143844",title:"Mr.",name:"Taufik",middleName:null,surname:"Abdul Hakim",slug:"taufik-abdul-hakim",fullName:"Taufik Abdul Hakim"}]},{id:"38485",title:"Transboundary Animal Diseases and International Trade",slug:"transboundary-animal-diseases-and-international-trade",totalDownloads:3112,totalCrossrefCites:1,totalDimensionsCites:3,book:{slug:"international-trade-from-economic-and-policy-perspective",title:"International Trade from Economic and Policy Perspective",fullTitle:"International Trade from Economic and Policy Perspective"},signatures:"Andrés Cartín-Rojas",authors:[{id:"139628",title:"Ms.",name:"Andrés",middleName:null,surname:"Cartín-Rojas",slug:"andres-cartin-rojas",fullName:"Andrés Cartín-Rojas"}]},{id:"53827",title:"Malaysia and China: The Trade Balances, Foreign Exchanges and Crises Impacts",slug:"malaysia-and-china-the-trade-balances-foreign-exchanges-and-crises-impacts",totalDownloads:1047,totalCrossrefCites:0,totalDimensionsCites:0,book:{slug:"international-trade-on-the-brink-of-change",title:"International Trade",fullTitle:"International Trade - On the Brink of Change"},signatures:"Tze-Haw Chan",authors:[{id:"191390",title:"Dr.",name:"Chan",middleName:null,surname:"Tze-Haw",slug:"chan-tze-haw",fullName:"Chan Tze-Haw"}]},{id:"59485",title:"Impacts of Exchange Rate Volatility on Macroeconomic and Financial Variables: Empirical Evidence from PVAR Modeling",slug:"impacts-of-exchange-rate-volatility-on-macroeconomic-and-financial-variables-empirical-evidence-from",totalDownloads:460,totalCrossrefCites:0,totalDimensionsCites:0,book:{slug:"trade-and-global-market",title:"Trade and Global Market",fullTitle:"Trade and Global Market"},signatures:"Oguzhan Ozcelebi",authors:[{id:"226325",title:"Prof.",name:"Oguzhan",middleName:null,surname:"Ozcelebi",slug:"oguzhan-ozcelebi",fullName:"Oguzhan Ozcelebi"}]},{id:"59359",title:"Analysis of the Role of Exchange Rate Volatility in Monetary Policy Conduction in OECD Countries: Empirical Evidence from Panel-VAR Models",slug:"analysis-of-the-role-of-exchange-rate-volatility-in-monetary-policy-conduction-in-oecd-countries-emp",totalDownloads:514,totalCrossrefCites:0,totalDimensionsCites:0,book:{slug:"trade-and-global-market",title:"Trade and Global Market",fullTitle:"Trade and Global Market"},signatures:"Oguzhan Ozcelebi",authors:[{id:"226325",title:"Prof.",name:"Oguzhan",middleName:null,surname:"Ozcelebi",slug:"oguzhan-ozcelebi",fullName:"Oguzhan Ozcelebi"}]},{id:"53456",title:"Globalization, Governance, Democratization and Fair Trade",slug:"globalization-governance-democratization-and-fair-trade",totalDownloads:928,totalCrossrefCites:0,totalDimensionsCites:0,book:{slug:"international-trade-on-the-brink-of-change",title:"International Trade",fullTitle:"International Trade - On the Brink of Change"},signatures:"Rasto Ovin and Pedja Ašanin Gole",authors:[{id:"175615",title:"Prof.",name:"Rasto",middleName:null,surname:"Ovin",slug:"rasto-ovin",fullName:"Rasto Ovin"}]},{id:"53513",title:"Risks Associated with International Trading of Medicines by Non-Licensed Entities and Non-Professionals",slug:"risks-associated-with-international-trading-of-medicines-by-non-licensed-entities-and-non-profession",totalDownloads:894,totalCrossrefCites:2,totalDimensionsCites:2,book:{slug:"international-trade-on-the-brink-of-change",title:"International Trade",fullTitle:"International Trade - On the Brink of Change"},signatures:"Ntambwe Malangu",authors:[{id:"84773",title:"Prof.",name:"Ntambwe",middleName:null,surname:"Malangu",slug:"ntambwe-malangu",fullName:"Ntambwe Malangu"}]},{id:"38487",title:"A Perspective on Remanufacturing Business: Issues and Opportunities",slug:"a-perspective-on-remanufacturing-business-issues-and-opportunities",totalDownloads:3247,totalCrossrefCites:3,totalDimensionsCites:8,book:{slug:"international-trade-from-economic-and-policy-perspective",title:"International Trade from Economic and Policy Perspective",fullTitle:"International Trade from Economic and Policy Perspective"},signatures:"Mosè Gallo, Elpidio Romano and Liberatina Carmela Santillo",authors:[{id:"11826",title:"Prof.",name:"Elpidio",middleName:null,surname:"Romano",slug:"elpidio-romano",fullName:"Elpidio Romano"},{id:"11827",title:"Prof.",name:"Liberatina",middleName:null,surname:"Santillo",slug:"liberatina-santillo",fullName:"Liberatina Santillo"},{id:"138996",title:"PhD.",name:"Mosè",middleName:null,surname:"Gallo",slug:"mose-gallo",fullName:"Mosè Gallo"}]}],onlineFirstChaptersFilter:{topicSlug:"international-trade",limit:3,offset:0},onlineFirstChaptersCollection:[],onlineFirstChaptersTotal:0},preDownload:{success:null,errors:{}},aboutIntechopen:{},privacyPolicy:{},peerReviewing:{},howOpenAccessPublishingWithIntechopenWorks:{},sponsorshipBooks:{sponsorshipBooks:[{type:"book",id:"10080",title:"Vortex Dynamics",subtitle:null,isOpenForSubmission:!0,hash:"ea97962e99b3e0ebc9b46b48ba5bea14",slug:null,bookSignature:"Dr. Zambri Harun",coverURL:"https://cdn.intechopen.com/books/images_new/10080.jpg",editedByType:null,editors:[{id:"243152",title:"Dr.",name:"Zambri",middleName:null,surname:"Harun",slug:"zambri-harun",fullName:"Zambri Harun"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"8903",title:"Carbon Based Material for Environmental Protection and Remediation",subtitle:null,isOpenForSubmission:!0,hash:"19da699b370f320eca63ef2ba02f745d",slug:null,bookSignature:"Dr. Mattia Bartoli and Dr. Marco Frediani",coverURL:"https://cdn.intechopen.com/books/images_new/8903.jpg",editedByType:null,editors:[{id:"188999",title:"Dr.",name:"Mattia",middleName:null,surname:"Bartoli",slug:"mattia-bartoli",fullName:"Mattia Bartoli"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"8771",title:"Raman Scattering",subtitle:null,isOpenForSubmission:!0,hash:"1354b3097eaa5b27d9d4bd29d3150b27",slug:null,bookSignature:"Dr. Samir Kumar and Dr. Prabhat Kumar",coverURL:"https://cdn.intechopen.com/books/images_new/8771.jpg",editedByType:null,editors:[{id:"296661",title:"Dr.",name:"Samir",middleName:null,surname:"Kumar",slug:"samir-kumar",fullName:"Samir Kumar"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"10073",title:"Recent Advances in Nanophotonics-Fundamentals and Applications",subtitle:null,isOpenForSubmission:!0,hash:"aceca7dfc807140870a89d42c5537d7c",slug:null,bookSignature:"Dr. Mojtaba Kahrizi and Ms. Parsoua Abedini Sohi",coverURL:"https://cdn.intechopen.com/books/images_new/10073.jpg",editedByType:null,editors:[{id:"113045",title:"Dr.",name:"Mojtaba",middleName:null,surname:"Kahrizi",slug:"mojtaba-kahrizi",fullName:"Mojtaba Kahrizi"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"10132",title:"Applied Computational Near-surface Geophysics - From Integral and Derivative Formulas to MATLAB Codes",subtitle:null,isOpenForSubmission:!0,hash:"38cdbbb671df620b36ee96af1d9a3a90",slug:null,bookSignature:"Dr. Afshin Aghayan",coverURL:"https://cdn.intechopen.com/books/images_new/10132.jpg",editedByType:null,editors:[{id:"311030",title:"Dr.",name:"Afshin",middleName:null,surname:"Aghayan",slug:"afshin-aghayan",fullName:"Afshin Aghayan"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"10110",title:"Advances and Technologies in Building Construction and Structural Analysis",subtitle:null,isOpenForSubmission:!0,hash:"df2ad14bc5588577e8bf0b7ebcdafd9d",slug:null,bookSignature:"Dr. Ali Kaboli and Dr. Sara Shirowzhan",coverURL:"https://cdn.intechopen.com/books/images_new/10110.jpg",editedByType:null,editors:[{id:"309192",title:"Dr.",name:"Ali",middleName:null,surname:"Kaboli",slug:"ali-kaboli",fullName:"Ali Kaboli"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"10175",title:"Ethics in Emerging Technologies",subtitle:null,isOpenForSubmission:!0,hash:"9c92da249676e35e2f7476182aa94e84",slug:null,bookSignature:"Prof. Ali Hessami",coverURL:"https://cdn.intechopen.com/books/images_new/10175.jpg",editedByType:null,editors:[{id:"108303",title:"Prof.",name:"Ali",middleName:null,surname:"Hessami",slug:"ali-hessami",fullName:"Ali Hessami"}],productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"9284",title:"Computational Aeroacoustics",subtitle:null,isOpenForSubmission:!0,hash:"7019c5e5985faef7dc384c87dca5c8ef",slug:null,bookSignature:"Prof. Ramesh K. Agarwal",coverURL:"https://cdn.intechopen.com/books/images_new/9284.jpg",editedByType:null,editors:[{id:"38519",title:"Prof.",name:"Ramesh K.",middleName:null,surname:"Agarwal",slug:"ramesh-k.-agarwal",fullName:"Ramesh K. Agarwal"}],productType:{id:"1",chapterContentType:"chapter"}}],offset:8,limit:8,total:16},humansInSpaceProgram:{},teamHumansInSpaceProgram:{},route:{name:"profile.detail",path:"/profiles/66978/dirk-roymans",hash:"",query:{},params:{id:"66978",slug:"dirk-roymans"},fullPath:"/profiles/66978/dirk-roymans",meta:{},from:{name:null,path:"/",hash:"",query:{},params:{},fullPath:"/",meta:{}}}},function(){var e;(e=document.currentScript||document.scripts[document.scripts.length-1]).parentNode.removeChild(e)}()