Overview of decellularization methods (Thomas W. Gilbert)
\r\n\tAntiphospholipid syndrome is likely to be involved in a small percentage of infertility. It is mainly associated with repeated miscarriages, secretion of cytokines, and growth factors that affect both ovulation, fertilization, and implantation, as well as a previous ectopic pregnancy in the fallopian tubes. Ectopic pregnancy is linked to taking drugs to induce ovulation, but the exact pathogenetic mechanism associated with hormone intake and fallopian tube damage is not yet known.
\r\n\tFertilization of the egg with the sperm under normal conditions occurs in the fibrous part of the fallopian tube. After 3-4 days the fertilized egg reaches the uterus, where the blastocyst within 2 to 4 days may be in a state of immersion in the endometrial tissue. Thus, the implantation takes place on the 20th-21st day of the 4-week menstrual cycle. The pathology of the function of the fallopian tubes is most often associated with inflammatory processes of any etiology. Non-specific infection plays a predominant role, the spread of which contributes to abortion, intrauterine contraception, diagnostic intervention and the occurrence of obstetric and perinatal complications.
\r\n\tIn recent years, the increasing incidence of chlamydial infection has been linked to the occurrence of ectopic pregnancy. Along with the inflammatory nature of the structure and function of the fallopian tubes, endometriosis also seems to play an important e localization may be ovarian, cervical or intra-abdominal, but in most cases it is tubal. The incidence of ectopic pregnancies in the USΑ has at least quadrupled in recent years with the probability nowadays standing at 20 per 1000 pregnancies. Ectopic pregnancy in the USA is reported to be the cause of death in 10% of obstetric deaths, but it is important to note that most of these deaths are bleeding-related and preventable.
\r\n\tA clear trend of increasing the frequency of ectopic pregnancies has been observed in the last ten years, which is triggered by two main reasons. On the one hand, the localization of inflammatory processes in the internal genitals is constantly increasing, while at the same time, the number of surgeries on the fallopian tubes is increasing in order to improve a woman's reproductive capacity. Τhe number of women using intrauterine and hormonal methods of contraception is also increasing and ovulation inducers are increasingly being introduced into the practice of infertility treatment.
\r\n\tOn the other hand, diagnostic capabilities have been improved in recent years by allowing detection of intolerance and even remission of ectopic pregnancy. Currently, an ectopic pregnancy occurs from 0.8-2.4% of cases that are born. In 4-10% of cases, it is repeated. Ectopic pregnancy often occurs as a result of tubal pathology.
\r\n\tRisk factors include smoking, inflammation of the pelvic organs as a result of chlamydia or gonorrhea, and endometriosis, conditions that lead to the formation of scar tissue in the fallopian tubes.role.
\r\n\tThe role of invasive procedures for the treatment of the causative agents of structural abnormalities of the fallopian tubes in the occurrence of ectopic pregnancy is becoming increasingly crucial and even to such an extent that the introduction of microsurgery does not exclude the risks.
",isbn:null,printIsbn:"979-953-307-X-X",pdfIsbn:null,doi:null,price:0,priceEur:0,priceUsd:0,slug:null,numberOfPages:0,isOpenForSubmission:!1,isSalesforceBook:!1,isNomenclature:!1,hash:"46740c30c279d7ad0334167a63819809",bookSignature:"Prof. Panagiotis Tsikouras, Prof. Nikolaos Nikolettos, Prof. Werner Rath and Prof. Georg-Friedrich Von Tempelhoff",publishedDate:null,coverURL:"https://cdn.intechopen.com/books/images_new/11282.jpg",keywords:"Differential Diagnosis, Early Diagnosis, Abortion, Intrauterine Pregnancy, Surgery, Methotrexate, Salpingostomy, New Aspects, Markers, Endometriosis, Diagnosis, Management",numberOfDownloads:null,numberOfWosCitations:0,numberOfCrossrefCitations:0,numberOfDimensionsCitations:0,numberOfTotalCitations:0,isAvailableForWebshopOrdering:!0,dateEndFirstStepPublish:"September 9th 2021",dateEndSecondStepPublish:"October 7th 2021",dateEndThirdStepPublish:"December 6th 2021",dateEndFourthStepPublish:"February 24th 2022",dateEndFifthStepPublish:"April 25th 2022",dateConfirmationOfParticipation:null,remainingDaysToSecondStep:"9 months",secondStepPassed:!0,areRegistrationsClosed:!0,currentStepOfPublishingProcess:5,editedByType:null,kuFlag:!1,biosketch:"Dr. Panagiotis Tsikouras has authored and co-authored multiple peer-reviewed scientific papers and presented works at many national and international conferences. 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From 2007 to 2011, Dr.\nNikos served as director of the In Vitro Fertilization Unit, University Regional General Hospital of Alexandroupolis, Greece. From\n2012 to 2018, he was director of the Laboratory of Reproductive\nPhysiology - Artificial Fertilization. In 2018 he became Professor of Assisted Reproduction at the Democritus University of Thrace, Greece. In 2019 he took over the\nmanagement of the University Obstetrics-Gynecology Clinic, Medical Department,\nDemocritus University of Thrace. 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Naturally derived biomaterials have been demonstrated to show several advantages compared to synthetic biomaterials. These are biocompatibility, biodegradability and remodeling. Therefore, these biomaterials are usually applied in the repair or replacement of damaged human tissues and organs. The aim of this chapter is to provide a brief knowledge of naturally derived biomaterials as well as methods of preparation and application of them.
Biomaterials can be classified into two main groups: synthetic and natural biomaterials. Synthetic biomaterials are classified as: metals, ceramics, nonbiodegradable polymers, biodegradable polymers… Some synthetic biomaterials are commercialized and applied in clinical treatment such as metal hip, Dacron, plastic intraocular lens… However, synthetic biomaterials have some disadvantages, including their structure and composition is not similar to native tissues/organs, their biocompatibility and their ability to induce tissue remodeling are low. Thus, other biomaterials have been developed that can overcome the disadvantages of synthetic biomaterials. Today, naturally derived biomaterials have been attracting scientist’s interest all over the world. Naturally derived biomaterial can be classified into many groups including protein-based biomaterials (collagen, gelatin, silk…), polysaccharide-based biomaterials (cellulose, chitin/chitosan, glucose…) and decellularized tissue-derived biomaterials (decellularized heart valves, blood vessels, liver…). Protein and polysaccharide-based biomaterials can be prepared by two distinct ways. Protein and polysaccharide from living organisms are dissolved by solvents or enzymes. Then, they are precipitated and reconstituted into fibrils. The second way to prepare protein and polysaccharide is removing other elements of living organisms by solvents or enzymes. Decellularized biomaterials are created by eliminating all cells from native tissues/organs. Physical, chemical and enzymatic approaches are combined to make the effective decellularization protocol.
Because of their advantages, naturally derived biomaterials are usually applied to replace or restore structure and function of damaged tissues/organs. They have ability to adequately support cell adhesion, migration, proliferation and differentiation. In particular, when implanted into a defective area, naturally derived biomaterials can enhance the attachment and migration of cells from the surrounding environment, therefore, induce extracellular matrix formation and promote tissue repair. Some biomaterials are used to acting as drug delivery system and medical devices such as surgical sutures. The silk fiber produced by silkworm or spider has been used as a surgical suture for a long time due to its biodegradable and non-antigenic protein. These silk fibroin nanoparticles are the globules with a fine crystallinity that may offer various possibilities for surface modification and covalent drug attachment. Furthermore, some biomaterials are used to produce environmental friendliness of packaging (such as resorbable chitosan packing) and other products. Some commercial products were made from naturally derived biomaterial such as SIS, Matrigel, Alloderm… In this chapter, we focus on a brief knowledge as well as the methods of preparation and application of naturally derived biomaterials in our researches.
Collagen is the most abundant protein of connective tissues in all animals. Now, at least sixteen types of collagen have been identified, in which 80-90% of the collagen is types I, II and III. Collagen is secreted by not only fibroblasts but also epithelial cells [1].
The basic structural unit of collagen is a triple helix. Most collagen is fibrillar because of pack of collagen molecules type I, II, III. Contrast, collagen IV forms a two dimensional network which is unique to the basement membranes [1]. Basement membranes have been performed a number of mechanical and biological functions. They provide physical support for tissue because of their tensile strength. They also influence cell proliferation, adhesion, migration, differentiation, polarization, and are thus implicated in biological processes such as development, tissue maintenance, regeneration, and repair, and in various pathological processes such as tumor growth and metastasis [2].
The basement membranes composition varies from one tissue to another. In general, the major constituents of all basement membranes are collagen IV, laminins, nidogen/entactin, and proteoglycans. The functional diversity of basement membranes arises from the molecular diversity of their components, particularly the different collagen IV and laminin isoforms [2].
Collagen can be obtained from various sources, in which amniotic membrane (AM) is an attractive source. AM is a thin membrane surrounding the fetus which is filled with amniotic fluid.
The AM consists of an epithelial monolayer, a thick basement membrane, a compact layer, a fibroblast layer and a spongy layer [3]. The innermost layer, nearest to the fetus, is monolayer of epithelial cells anchored on the basement membrane. The collagen component of basement membrane of AM includes types III, IV, V, VII, XVII which similar morphological and ultra-structural basement membrane of skin. Therefore, basement membrane of AM is often used to create skin equivalents. Besides, AM has outstanding properties such as anti-inflammatory, anti-bacterial, anti-fibrosis, anti-scaring as well as low immunogenicity and reasonable mechanical features [3].
AM can be used either with amniotic epithelium (intact) or without it (denuded), fresh or preserved. To remove the amniotic epithelium, the AM is incubated in trypsin-EDTA at 37oC in 30 min and the cells are gently scraped while maintaining the intact basement membrane. H&E staining was performed to confirm removing the amniotic epithelium. Then, the basement membrane can be preserved by drying or glycerol - cryopreservation after γ-sterilization [3, 4, 5].
Collagen is commonly used in biomedical applications. The basement membrane of the AM is a typical example. The extracellular matrix components of the basement membrane of the AM are native scaffolds for cell seeding in tissue engineering. AM has been applied in tissue engineering related to eye, skin, cartilage, nerve, especially cancer [3, 6, 7].
To use AM in creating skin equivalent, AM must be removed the epithelium. Keratinocytes are seeded onto basement membrane which is denuded AM. Briefly, keratinocytes were isolated from intact skin samples by incubation in trypsin-EDTA at 40C in 18 hours and detached mechanically. Keratinocytes were cultured in serum free medium in 7 days. Medium was chanced every two days [8]. After 3rd passage, cells were subcultured onto AM basement membrane which spreaded on bottom of the insert dishes. The cells were maintained in culture for 7 days when the cells reached confluent. Air-lifting was performed to induce cell differentiation. After 7 days, the cells formed multi-layers on the AM basement membrane. Cultured keratinocyte sheets were grafted on patients who were defected skin because of injury or burn. The result showed that, the advantages of cultured keratinocyte sheet autograft: the possibility of the grafting area multiplication (50 times after 3 weeks), the diminution of scaring, the relief of pain, the low infection risk, the same effect in compare to the split - thinness autograft.
Amniotic membrane with epithelium: intact (a) and denuded (b)
Keratinocytes formed colonies (a) and monolayer onto culture dishes (b) (200X)
Result of HE staining of the cell sheet, the cells were exposed to the air in seven days. The cells formed 4 – 5 layers onto AM basement membrane (400X).
Gelatin is obtained by controlling the hydrolysis of collagen, a fibrous insoluble protein which is widely found in nature and is the major component of skin, bone and connective tissue. Characteristic features of gelatin are the high content of the amino acids such as glycine, proline and hydroxyproline. Structually, gelatin molecules contain repeating sequences of glycine-X-Y triplets, where X and Y are frequently proline and hydroxyproline. These sequences are responsible for the triple helical structure of gelatin and its ability to form gels where helical regions form in the gelatin protein chains immobilizing water [9].
Alginate was first discovered by Edward Stanford in 1883. Since being commercialized in 1927, alginate has now expanded to about 50.000 tonnes per year worldwide; 30% of this tonnage is devoted to the food industry, the rest being used in industrial, pharmaceutical and dental applications [10]. The function of alginates in algae is primarily skeletal, with the gel located in the cell wall and intercellular matrix conferring the strength and flexibility necessary to withstand the force of water in which the seaweed grows [11].
Alginate is a hydrophilic polysaccharide extracted from marine brown algae such as Laminaria hyperborea or soil bacteria such as Azobacter vinelandii and composed of 1,4-linked β-D-mannuronic acid (M) residues and 1,4-linked α-L-guluronic acid (G) in varying proportions, displaying carboxylic acid functionality at the C5 residue. The alginates have broad distributions of molecular weights of 10-1000 kDa depending on source and processing. The relative amount and sequential distribution of homogeneous M-M segments (M-blocks), homogeneous G-G segments (G-blocks) and alternating M-G segments (MG-blocks), which represent the primary structure of alginate, depend on the producing species, and for marine sources, on seasonal and geographical variations.
Cross-linked gelatin/alginate was made in two steps. Briefly, 1wt % (w/w) aqueous solution of gelatin and sodium alginate, respectively, was dissolved in double distilled water at 500C for 3 h. Each solution with certain mixing ratios of gelatin and sodium alginate (8G:2A) was stirred for 30min at room temperature, frozen to -700C for 40 h. This soluble sponge was cross-linked with EDC by immersing the soluble sponge in 90% (w/v) aqueous acetone containing 0.3% EDC for 24 h at room temperature, while shaking slowly.
The sponges consisting of gelatin from hydrolysis of collagen and alginate-a polysaccharide from Phaeophyta were established by using EDC as a crosslinking agent. The sponges were combined with some natural substances traditional used for burn treatment such as tamanu oil (from nuts of Calophyllum inophyllum); cajeputi oil (from leaves of Meulaleuca leucadendron); madecassol (from extract of Centella asiatica); turmeric and python fat. Data obtained from testing on mice showed that the coordinated sponges have rather good ability on preventing infection and promoting wound healing compared with control. The sponges combined with mixture of cajeputi oil and madecassol have the best potential for burn treatment.
Gelatin-Alginate. Block (a), SEM (b)
Gelatin has been used in medicine as plasma expander, wound dressing, adhesive, and absorbent pad for surgical use. While collagen, also known to have wide biomedical applications, expresses antigenicity in physiological condition, gelatin is known to have no such antigenicity. Recently, gelatin has shown to exhibit activation of macrophages and high hemostatic effect. Finally, gelatin is practically more convenient than collagen because a concentrated collagen solution is extremely difficult to prepare from the native collagen, and furthermore gelatin is far more economical than the collagen.
Alginate is an effective natural disintegrant, tablet binder and offers an attractive alternative for sustained-release systems. It offers advantages over synthetic polymers as it forms hydrogels under relatively mild pH and temperature and is generally regarded as non-toxic, biocompatible, biodegradable, less expensive and abundantly available in nature; in addition, alginate meets the important requirement of being amenable to sterilization and storage. All these advantages make alginates very useful materials for biomedical applications, especially for controlled delivery of drugs and other biologically active compounds and for the encapsulation of cells. Calcium alginate is a natural haemostat, so alginate based dressings are indicated for bleeding wounds. The gel forming property of alginate helps in removing the dressing without much trauma [12].
Alginate has been much used in medical applications such as wound dressings, scaffolds for hepatocyte culture and surgical or dental impression materials. Alginates are also known to be broken down to simpler glucose type residues and can be totally absorbed.
Alginate has been successfully used as a matrix for the entrapment and/or delivery of biological agents, such as drugs and proteins. In particular proteins can be loaded and released by alginate matrices without loss of their biological activity because of the relatively mild gelation process of alginate. In pharmaceutical formulations, the alginate gel can be prepared prior to use, or it can spontaneously form in situ in physiological fluids, by low pH and/or calcium ions naturally present in the site of administration [13]. Alternatively, the gelling agent can be added either as a part of the formulation or separately administered. The microencapsulation technique has been specifically developed for the oral delivery of proteins, as they are quickly denaturated and degraded in the hostile environment of the stomach.
Among the possible applications of alginate, one of the most promising is for cell immobilization. Alginate gel allows cell suspension to be cultivated in several types of bioreactors to achieve high cell densities [14]. In cell immobilization applications, the main drawback of alginate matrix gels is represented by their high density of network, which limits the cell growth; moreover, cell anchorage, a strict requirement for survival, is limited on alginate gels, because of its hydrophilic nature. PEG copolymers are used to improve their biocompatibility [15].
Silks are generally defined as protein polymers that are spun into fibers by Lepidoptera larvae such as silkworms, spiders, scorpions, mites and flies [16]. Silks are fibrous proteins synthesized in specialized epithelial cells that line glands in these organisms. Silk fibroin polymers consist of repetitive protein sequences and provide structural roles in cocoon formation, nest building, traps, web formation, safety lines and egg protection. The most extensively used silk for various applications are those from silkworm silk; Bombyx mori and spider silk; Nephila clavipes.
The domesticated silkworm (B. mori) silk fibroin fibers are about 10–25 μm in diameter. Each fiber consists of core protein covered by a coating protein (sericin) that glues core fibers together. The core protein consists of three chains: heavy chain, light chain and a glycoprotein, P25. The light chain (26 kDa) and heavy chain (390 kDa) which are present in a 1:1 ratio and linked by a single disulfide bond. The disulfide linkage between the Cys-c20 (20th residue from the carboxyl terminus) of the heavy chain and Cys-172 of the light chain holds the fibroin together and a 25 kDa glycoprotein, named P25, is non-covalently linked to these proteins [17]. Light chain is necessary for the secretion of protein from the silk glands. Heavy chain is fiber forming protein and its structure determines properties of silk fiber [18]. Heavy chain is commonly referred as fibroin protein. These proteins are coated with a family of hydrophilic proteins called sericins (20–310 kDa). Silk proteins are particularly promising for these needs due to their unique combination of biocompatibility, biodegradability, self-assembly, mechanical stability, controllable structure and morphology.
Spiders are look like insects and come in to the category of Arthropoda, but they belong to a completely different class of animals, called Arachnida. Spider silk is the result of 400 million years of evolution. Spiders use silk for variety of functions including reproduction as cocoon capsular structures, lines for prey capture, lifeline support (dragline), web construction and adhesion. Spider silk is a biopolymer fiber. Its composition is a mix of an amorphous polymer (which makes the fiber elastic), and chains of two of the simplest proteins (which give it toughness). Out of 20 amino acids, only Glycine and Alanine serve as a primary constituent of silk. The Dragline silk of orbweb spider seems to be most studied in the scientific research. The protein in dragline silk is fibroin (Mass of 200,000-300,000 Daltons) which is a combination of the proteins spidroin 1 (Alanine-rich) and spidroin 2 (Glycine-rich), the exact composition of these proteins depends on species. Fibroin consists of approximately 40% Glycine and 25% Alanine as the major amino acids. The remaining components are mostly glutamine, serine, leucine, valine, proline, tyrosine and arginine [19]. Nephila clavipes can produce seven types of silk from seven different silk glands as shown in depending on needs and environmental conditions [20]. The superior mechanical properties of dragline spider silks can be used as a template for developing specific structures for various biomaterial needs. Spider silks have not been commercialized in fashion as silkworm silk due to the lack of domestication and lower productivity of spiders.
Nanotechnology is becoming a key technology and capable of application in all fields of science and technology. In particular, nanoparticle delivery system significantly improved pharmaceutical treatment of many incurable diseases which require complex treatment regimens, as well as, patients must take multiple medications and need time long-term drug use. Silk Protein is the ideal material for this purpose, because they have many unique features such as highly biocompatible and biodegradable ability, self-restructuring, mechanical stability, easy control and adjustment of the object\'s structure and shape.
Nanofibroin particles
The cocoon shell of silkworm Bombyx mori was degummed in boiling solution of 0,5% Na2CO3 in 700C for 35 min. Then degummed fiber was dissolved in a mixed solution of calcium chloride, ethanol, and water (CaCl2/C2H5OH/H2O: 1:2:8 mole ratio), at 800C. After the silk fibroin–salts solution was centrifuged at 5000 rpm for 10 min, the supernatant was dialyzed continuously for 72 h against running pure water to remove CaCl2, smaller molecules, and some impurities. The resulting liquid silk fibroin was stored at 40C and used in the following experiments for the preparation of silk fibroin nanoparticles. Spider silk proteins form nanoparticles upon salting out with potassium phosphate. Milk-like silk protein particles were formed at once and suspended in the mixture comprising water and organic solvent. These protein particles were water insoluble and went down slowly due to the gathering of microparticles. The precipitates of silk protein nanoparticles were collected and purified from the mixture by repeated centrifugation at 20,000 rpm to separate these particles from the solvent. After the research, we have obtained nanoparticles (500 nm-2000 nm) from silk protein can load and delivery of proteins in vitro.
The silk bio-polymer is used in tissue regeneration for treating burn victims and as matrix of wound healing. The silk fibroin peptides are used in cosmetics due to their glossy, flexible, elastic coating power, easy spreading and adhesion characters [21]. Silk powder is touted and relieves from sunburns, due to crystalline structure it reflects UV radiation and as demulcent it acts as protective buffer between skin and environment. The lower micron silk powder is added with hair and massage oils and water dispersible finer grade silk powder is an ingredient of liquid cosmetic preparations.
The silk is used to fight edema, cystitis, impotence, adenosine augmentation therapy, epididymitis and cancer [22]. Silk protein derivative, Serratio peptidase is used as anti-inflammatory, anti-tumefacient for treating acute sinusitis, tonsiloctomy, oral surgery, tooth filling, cleaning and extractions. The silk fibroin is a useful dressing material with the property of non-cytotoxic to the tissues and also in veterinary medication.
Since long, silk fiber is being used as surgical sutures as it does not cause inflammatory reactions and is absorbed after wounds heal. Other promising medical applications are as biodegradable micro tubes for repair of blood vessels and as molded inserts for bone, cartilage and teeth reconstruction [23, 24, 25]. In biomedical and bioengineered field, the use of natural fibre mixed with biodegradable and bio-resorbable polymers can produce joints and bone fixtures to alleviate pain for patients.
Drug delivery is a rapidly developing field in biomedical research. It is interdisciplinary and requires expertise in biotechnology, pharmacology, microbiology, biochemistry, polymer chemistry and materials engineering. Advantages of using such systems include maintenance of drug levels within desired range, fewer administrations, optimal use of the drug, and better patient compliance. The material used for drug delivery should be biocompatible, chemically inert, easily processable and physically and mechanically stable. Biopolymers are of great interest for this kind of application. Silk and silk-like variants are used by some scientists as carriers for drug delivery. Their biocompatibility and ability to form hydrogels in situ makes them attractive candidates for the localized, controlled delivery of therapeutic agents. Their ability to incorporate drugs at room temperature, by simple mixing, and without the use of toxic or denaturing solvents makes them attractive for the delivery of protein or DNA-based therapies [26].
Future applications of silk biomaterials include new generation soft contact lenses that enable greater oxygen permeability, artificial corneas, skin grafts and epilepsy drug permeable devices.
The mechanism of fibrin fomation is elucidated primarily from the thrombin-mediated cleavage of fibrinogen. Fibrinogen, the principal protein of blood clotting, is a 340 kDa trinolar protein which presents at high concentration in blood plasma (2 – 4 mg/ml, 6 – 12 µM). Fibrinogen molecule consists of three different pairs of polypeptide chains (Aα, Bβ and γ) cross-linked to each other by 29 disulfide bridges (Fig. 1). The amine-termini (N-termini) of six polypeptide chains are converged in the central of fibrinogen molecule called the E domain. The carboxy-termini (C-termini) of the Bβ chain and γ chain comprise of the distal D domain. The C-termini of Aα chains which are known as globular, depart from the D domain and fold into a conformation that stretches back toward the E domain of fibrinogen [27].
Fibrinogen plays as precursor protein of fibrin in blood clotting. The conversion of fibrinogen to fibrin occurs in 3 ordered steps. In the intinial step, thrombin binds to the central E domain of fibrinogen and slipts off the fibrinopeptides A – FpA (16 amino acid residues) and B – FpB (14 amino acid residues) from N-termini of Aα and Bβ chains, respectively, whereas the γ chains remain unaltered. The cleavage of FpA and FpB results in exposure of “A” and “B” binding sites. Then, the self-assembly proccess will spontaneously occur. The “A” and “B” sites will interact with complementary sites (“a” and “b” sites located in the γ and β chain) at the D domain of other fibrinogen molecules, which results in new fibrin monomers. The fibrin monomers are bound to each other non-covalently (Fig. 2) and assemble in a half-staggered manner into two-stranded protofibrils which continue to aggregate laterally to form fibers branching into a three-dimensional network of fibrin [28, 29]. Finally, fibrin cross-linking is activated by Factor XIII (FXIII) in order to improve the strong and elastic properties, additionally, avoid fibrinolysis in solution [30, 31].
According to the usage purposes, some methods have been applied to fabricate fibrin gels. Fibrin gels can be conducted either from the separating components including thrombin, fibrinogen and CaCl2, or from serum of patients, which will be mentioned in two following methods, respectively. For manipulation of fibrin gel from separating commercial components, fibrin gels were prepared by combining fibrinogen, NaCl, thrombin, CaCl2. This complex is also supplemented with aprotinin in order to proving a stable fibrin structure and prevents postoperative bleeding. The contents are allowed to gel for 1 hr in standard culture conditions [32]. Furthermore, fibrin gel physical properties can be manipulated by adjusting the fibrinogen and CaCl2 concentration [33, 34], or using different cross linking agents such as enzymes or UV radiation [35]. In terms of autologous fibrin glue, the patient blood is havested and prepared 3 to 4 days before surgery. The plasma is separated from red blood cells by allowing the blood tube to stand vertically for at least 2 hours or centrifuged at 4000 rmp for 5 minutes. The fibrin gel preparation is created by combining plasma with commercial thrombin and calcium at appropriate concentration [36]. Autologous plasma fibrin gel not only shows an excellent hemostatic agent, but also helps eliminate the risk of viral transmission associated using donor plasma.
Numerous studies have exploited fibrin function as heamostatic plug, scaffold for cell proliferation and migration, and wound healing, which suggest fibrin potential applications in medical and tissue engineering. Fibrin glue or fibrin sealant is a formulation of fibrinogen and thrombin at very high amounts cobined with calcium and FXIII, used as an adjunct to hemostasis in patients undergoing surgery. Commercial products of fibrin sealant such as Tisseel (Immuno, Vienna, Austria), Beriplast (Behringwerke AG, Marburg/Lahn, FRG), and Biocol (CRTS, Lille, France) have been extensively used in clinical. In addition to fibrin’role in heamostatic, fibrin is also indicated as biological scaffold for cell proliferation, migration and differentiation applied in various tissue engineering. Natural fibrin matrix consists of sites for cellular binding, and has been shown to have excellent effects in cell culture and accelabrate tissue regeneration. In 2000, Ye et al. fabricated and investigated the three-dimensional fibrin scaffold in cardiovascular tissue engineering. In this research, human myofibroblasts (MFBs) from the ascending aorta were cultured in fibrin gel solution. Consequently, the cell growth, high collagen secretation and tissue development were determined. Besides, toxic degradation or inflammatory reactions was not detected in the fibrin gels [37]. In 2003, W. Bensa.ıd and colleagues conducted a research in which they use fibrin glue as a delivery system for human MSCs (HMSCs). The result confirmed a good good HMSCs spreading and proliferation in the fibrin scaffold. Besides, the HMSCs migration out of the fibrin scaffold and appearance of calcium carbonate from the differentiation of HMSCs when implanted in vivo suggest that fibrin gel is a promising delivery system for HMSCs toward bone healing application [38]. Fibrin glue also performs its role in the application of skin grafts to burned areas. Using fibrin glue instead of sutures or pressure dressings in the immediate postoperative period enhances healing, and minimizes scarring [39]. One of commercial fibrin sealant products used for burn treatment is ARTISS fibrin sealant (Baxter International Inc., USA). ARTISS fibrin sealant is indicated to adhere autologous skin grafts to surgically prepared wound beds resulting from burns, for both adults and pedipatients.
Cellulose is the most abundant polymer on Earth, which makes it also the most common organic compound. Annual cellulose synthesis by plants is close to 1012 tons. Plants contain approximately 33% cellulose whereas wood contains around 50% and cotton contains 90%. Most of the cellulose is utilised as a raw material in paper production. This equates to approximately 108 tons of pulp produced annually. From this, only 4 million tons are used for further chemical processing annually. It is quite clear from these values that only a very small fraction of cellulose is used for the production of commodity materials and chemicals [40]. Cellulose, a linear polysaccharide of up to 15,000 D-glucose residues linked by β-(1→4)-glycosidic bonds, is biocompatible and has excellent thermal, mechanical properties. It is considered easily biodegradable, thus less contaminating to the environment.
Cellulose is regarded as a semi-flexible polymer. The relative stiffness and rigidity of the cellulose molecule is mainly due to the intramolecular hydrogen bonding. This property is reflected in its high viscosity in solution, a high tendency to crystallise, and its ability to form fibrillar strands. The chain stiffness property is further favoured by the β-glucosidic linkage that bestows the linear form of the chain. The chair conformation of the pyranose ring also contributes to chain stiffness. This is in contrast to the α-glucosidic bonds of starch [41].
Plants are an attractive cellulose source primarily because they are abundant and there is a preexisting infrastructure in the textile industries for harvesting, retting/pulping (i.e. to treat and isolate micron sized cellulose particles), and product processing. Tunicates are the only animals known to produce cellulose microfibrils. Tunicates are a family of sea animals that have a mantle consisting of cellulose microfibrils embedded in a protein matrix. It is this thick leathery mantle in their mature phase that is used as a source of cellulose microfibrils. Most research has used a class of Tunicates that are commonly known as ‘‘sea squirts’’ (
Cellulose derivatives and composites offer an excellent biocompatibility, and are considered as promising materials for biochemical engineering for economic and scientific reasons.
Oxidized cellulose (oxycellulose) is cellulose in which some of the terminal primary alcohol groups of the glucose residues have been converted to carboxyl groups. Therefore, the product is possibly a synthetic polyanhydrocellobiuronide and that contain 25% carboxyl groups are too brittle and too readily soluble to be of use. Those products that have lower carboxyl contents are the most desirable[43].
Purified microcrystalline cellulose (MCC) is partially depolymerized cellulose prepared by treating α-cellulose, obtained as a pulp from fibrous plant material, with mineral acids. Silicified MCC (SMCC) is manufactured by codrying a suspension of MCC particles and colloidal silicon dioxide such that the dried finished product contains 2% colloidal silicon dioxide. SMCC shows higher bulk density than the common types of MCC. Also, tensile strength of compacts of SMCC is greater than that of the respective MCC and it is most probably a consequence of intersurface interactions of silicon dioxide and MCC [44].
The esterification can be considered as a typical equilibrium reaction in which an alcohol and acid react to form ester and water. Cellulose is esterified with certain acids such as acetic acid, nitric acid, sulfuric acid, and phosphoric acid. A prerequisite is that the acid used can bring about a strong swelling thus penetrating throughout the cellulose structure. Cellulose acetate phthalate is a partial acetate ester of cellulose that has been reacted with phthalic anhydride. One carboxyl of the phthalic acid is esterified with the cellulose acetate. The finished product contains about 20% acetyl groups and about 35% phthalyl groups [45].
The treatments for wood and plants involve the complete or partial removal of matrix materials (hemicellulose, lignin, etc.) and the isolation of individual complete fibers. Fortunicate the treatment involves the isolation of the mantel from the animal and the isolation of individual cellulose fibrils with the removal of the protein matrix. Treatments for algal cellulose sources typically involve culturing methods, and then purifying steps for removal of algal wall matrix material. Bacterial cellulose treatments focus on culturing methods for cellulose microfibrillar growth and then washing to remove the bacteria and other media.
The general processing of engineered BC materials can be considered to occur in four main stages: (1) BC culturing, (2) pellicle management, (3) water removal, and (4) chemical modification. For stage 1, the biosynthesis of BC occurs in culture solutions, generally in a bioreactor, in which bacteria secrete cellulose microfibrils, producing an interwoven web of fibrils that is a hydrogel. The hydrogels are composed of entangled cellulose microfibrils formed from the random motion of the bacteria, contain upwards of 99% water, and are called pellicles. For stage 2, pellicle management refers to any process imparted on the pellicle up until the point of water removal. To remove the bacteria from the pellicles, the pellicles are washed by boiling in a low concentration (2%) NaOH bath for several hours, then it is rinsed under running water for several days. Additional NaOH and NaClO treatments have also been used for further purification of the BC microfibrils. For stage 3, once the pellicle is formed and purified, a sample is cut from the gel-like sheet. Water removal either by evaporation or a combination of pressing and evaporation collapses the gel-network and produces a dense film. For stage 4, chemical modification to the BC microfibril network can be achieved at three points along the engineered BC material processing, (i) during stage 1, (ii) during stage 2, and (iii) after stage 3 (i.e. to dried BC structures or films) [42].
Cellulose is extensively used as a raw material in the paper industry in the production of paper and cardboard products. However, cellulose has shown its versatility in numerous applications.
Natural cellulose spheres are often applied in bioseparation, immobilized reaction, cell suspension culture, and as an adsorbent for sewage treatment. Spherical BC produced from dynamic method is translucent, loose, porous, and has a hydrophilic network structure. Its specific surface area increases with decreasing spherical diameter, so it could be used as a carrier to adsorb or crosslink various kinds of substances (e.g., enzyme, cell, protein, nucleic acid, and other compounds). Spherical BC may be applied in bioseparation, immobilized reaction, cell suspension culture, and as an adsorbent for sewage treatment. Compared with natural spherical cellulose, the fermentation production of BC spheres is simple, controllable and environment friendly. Moreover, BC sphere can be used repeatedly, expanding their potential applications.
Cellulosic derivatives such as cellulose acetate, cellulose propionate and cellulose acetate-butyrate, cast as membranes, have been reported as useful supports for immobilizing various enzymes such as catalase, alcohol oxidase and glucose oxidase. These supports gave better activity and storage stability for the enzymes. Cellulose ethers are widely used as important excipients for designing matrix tablets. On contact with water, the cellulose ethers start to swell and the hydrogel layer starts to grow around the dry core of the tablet. The hydrogel presents a diffusional barrier for water molecules penetrating into the polymer matrix and the drug molecules being released. Cellulose acetate butyrate microcapsules, as well as cellulose-based microspheres, have been used for the delivery of drugs [46].
Microbial cellulose synthesized by
Chitin is a white, hard, inelastic, nitrogenous polysaccharide found in the exoskeleton as well as in the internal structure of invertebrates. Chitin is a hydrophobic linear polysaccharide derived from many natural sources including the exoskeleton of arthropods and insects and is the second most abundant natural polysaccharide next to cellulose. Chitin comprises a polysaccharide consisting of (1→4)-β-N-acetyl-D-glucosamine units. Derivatives of chitin may be classified into two categories; in each case, the N-acetyl groups are removed, and the exposed amino function then reacts either with acyl chlorides or anhydrides to give the group NHCOR or is modified by reductive amination to NHCH2COOH of greatest potential importance are derivatives of both types formed by reaction with bi or polyfunctional reagents, thus carrying sites for further chemical reaction [48]. In practice, such reactions are carried out on native chitin or on incompletely deacetylated chitin, chitosan, so that the resulting polymer contains three types of monomeric units.
Chitosan is a partially deacetylated derivative of chitin and is the second most abundant biosynthesized material. Structurally, chitosan is a mixture of N-acetyl-D-glucosamine and D-glucosamine [49]. Generally, chitosan is insoluble in neutral or basic conditions, while protonation of free amino groups facilitates solubility of chitosan in dilute acids (pH < 6). In vivo degradation of chitosan is mainly attributed to the effect of lysozyme through hydrolysis of acetylated residues.
Chitosan itself chelates metal ions, especially those of transition metals, and also finds application as a matrix for immobilization of enzymes. Special attention has been given to the chemical modification of chitin, since it has the greatest potential to be fully exploited. Reactions with pure chitin have been carried out mostly in the solid state owing to the lack of solubility in ordinary solvents. A 50% deacetylated chitin has been found to be soluble in water [50]. This water soluble form of chitin is a useful starting material for its smooth modifications, through various reactions in solution phase. Some of the very recently reported chitosan derivatives are enumerated as follows:
Fully deacetylated chitosan was treated with phthalic anhydride to give N-phthaloyl-chitosan. It was readily soluble in polar organic solvents. Further reactions had been carried out using this new derivative to improve the solubility of chitosan [51].
To improve water solubility, Sashiwa
Recently, Baba
The synthesis of chitosan hydrogels was carried out by Qu
Chitin is easily obtained from crab or shrimp shells and fungal. In the first case, chitin production is associated with food industries such as shrimp canning. In the second case, the production of chitosan–glucan complexes is associated with fermentation processes, similar to those for the production of citric acid from
Chitin and chitosan are known for their excellent biological properties, among which the biocompatibility with human cells, the ordered regeneration of wounded tissues, the immunoenhancing activity, the induction of immediate hemostasis, the radical scavenging activity, and the antimicrobial activity. Recent studies indicate that chitin and chitosan are most versatile in drug and gene delivery, elaborated diagnostics, devices for selective recognition of tumor cells, and surgical aids ranging from anti-adhesion gels to coated sterile stents.
As a kind of renewable resource, unmodified chitosan has been widely used in many fields such as pharmaceutical, agriculture, food, and biomedical applications. In order to realize the full potential of chitosan and bring a breakthrough in its broader utilization, attempts have been made to modify chitosan to obtain various derivatives. For the tissue repair and regeneration applications, chitosan can be functionalized by chemical reaction, coupling with specific ligands or moieties, combining with biomacromolecules, and crosslinking in the presence or absence of crosslinkers.
Particularly, skin substitute made of chitosan or its derivatives have attracted much attention due to the outstanding characteristics of chitosan, such as biocompatibility, hemostatic activity, antibacterial property, and ability to accelerate the wound-healing process [56].
The design of artificial kidney systems has made possible repetitive hemodialysis and the sustaining life of chronic kidney failure patients. Chitosan membranes have been proposed as an artificial kidney membrane because of their suitable permeability and high tensile strength. The most important part of artificial kidney is the semipermeable membrane and so far made from commercial regenerated cellulose and cuprophane. Since the primary action of the cellulose membrane is that of a sieve, there is little selectivity in the separation of two closely related molecules. These novel membranes need to be developed for better control of transport, ease of formability and inherent blood compatibility.
A series of membranes prepared from chitin and its derivatives improved dialysis properties. One of the most serious problems of using these artificial membranes is surface induced thrombosis, where heparization of blood is needed to prevent clotting, and people who are liable to internal hemorrhage can be dialysed only at great risk. Hence, these are the most challenging problem still to be resolved in the development of membranes which are inherently blood compatible. From these point of views, chitosan is hemostatic, i.e., causes clots [57].
Chitosan has replaced the synthetic polymers in opthalmological applications. Chitosan possesses all the characteristics required for an ideal contact lens; optical clarity, mechanical stability, sufficient optical correction, gas permeability, partially towards oxygen, wettability, and immunologically compatibility. Contact lenses are made from partially depolymerized and purified squid pen chitosan by spin casting technology, and these contact lenses are clear, tough, and possess other required physical properties such as modulus, tensile strength, tear strength, elongation, water content, and oxygen permeability. Antimicrobial and wound healing properties of chitosan along with excellent film forming capability make chitosan suitable for development of ocular bandage lens [58].
The special attention on chitosan has been paid for the repair of articular cartilage. Articular cartilage is particularly vulnerable to injury trama, disease or congenital abnormalities because of its avascular, alypmhatic and aneural nature. Once damaged, it has little capacity for intrinsic repair. Although many repair techniques have been attempted over the past four decades, but none has succeeded to regenerate long-lasting hyaline cartilage tissue to replace defected or damaged cartilage. Recently, preliminary studies on chitosan-GAG composite and its biologically interaction with articular chondrocytes showed promising results. Chitosan and its derivatives are being extensively used for bone tissue engineering and central nervous system also.
The growth of
Chitosan is non-toxic and easily bioabsorbable with gel-forming ability at low pH. Moreover, chitosan has antacid and antiulcer activities which prevent or weaken drug irritation in the stomach. Also, chitosan matrix formulations appear to float and gradually swell in an acid medium. All these interesting properties of chitosan make this natural polymer an ideal candidate for controlled drug release formulations [60].
Autologous grafts are “gold standard” for implantation. However, the most disadvantage of autologous is quantity. The number of autograft does not meet needs of patients. Homograft is greater than autograft but they cannot satisfy needs of patient. Many patients must wait for a long time to take a homogenous organ. Xenograft is the greatest but they can evoke serious immune reaction. So, one method developed to process homograft and xenograft is decellularization. Every tissue/organ concludes cells and extracellular matrix. Cells are structure and functional units of tissue/organ but cells are major antigen of tissue/organ. Extracellular matrix is many protein, polysaccharide, protoglycan released by cell. ECM plays an important role in mechanical support, signal transportation, adherence of tissue/organ. Decellularization is a multi-step process to remove all cell components from tissue/organ and leave intact ECM. Many decellularization agents were researched such as physical methods, chemical methods and enzyme methods. Every decellularization agent has specific affections of cell and extracellular matrix. So, these agents are combined to make an effective decellularization process which removes all cell components and reverses maximum ECM. Decellularization effectiveness depends on type of tissue/organ. One agent can be a good detergent for decellularizing one tissue but not for another [61]. Moreover, cell derived ECM can be used as a matrix for cell culture.
\n\t\t\t\t | \n\t\t\t\n\t\t\t\t | \n\t\t\t\n\t\t\t\t | \n\t\t
Physical | \n\t\t||
Snap freezing | \n\t\t\tIntracellular ice crystals disrupt cell membrane | \n\t\t\tECM can be disrupted or fracture during rapid freezing | \n\t\t
Mechanical force | \n\t\t\tPressure can burst cells and tissue removal eliminates cells | \n\t\t\tMechanical force can cause damage to ECM | \n\t\t
Mechanical agitation | \n\t\t\tCan cause cell lysis, but more commonly used to facilitate chemical exposure and cellular material removal | \n\t\t\tAggressive agitation or sonication can disrupt ECM as the cellular material is removed | \n\t\t
Chemical | \n\t\t||
Alkaline; acid | \n\t\t\tSolubilize cytoplasmic components of cells; disrupts nucleic acids | \n\t\t\tRemoves GAGs | \n\t\t
Non-ionic detergents | \n\t\t||
Triton X-100 | \n\t\t\tDisrupts lipid–lipid and lipid–protein interactions, while leaving protein – protein interactions intact | \n\t\t\tMixed results; efficiency dependent on tissue, removes GAGs | \n\t\t
Ionic detergents | \n\t\t||
Sodium dodecyl sulfate (SDS) | \n\t\t\tSolubilize cytoplasmic and nuclear cellular membranes; tend to denature proteins | \n\t\t\tRemoves nuclear remnants and cytoplasmic proteins; tends to disrupt native tissue structure, remove GAGs and damage collagen | \n\t\t
Sodium deoxycholate | \n\t\t\t\n\t\t\t | More disruptive to tissue structure than SDS | \n\t\t
Triton X-200 | \n\t\t\t\n\t\t\t | Yielded efficient cell removal when used with zwitterionic detergents | \n\t\t
Zwitterionic detergents | \n\t\t||
CHAPS | \n\t\t\tExhibit properties of non-ionic and ionic detergents | \n\t\t\tEfficient cell removal with ECM disruption similar to that of Triton X-100 | \n\t\t
Sulfobetaine-10 and -16 (SB-10, SB-16) | \n\t\t\t\n\t\t\t | Yielded cell removal and mild ECM disruption with Triton X-200 | \n\t\t
Tri(n-butyl)phosphate | \n\t\t\tOrganic solvent that disrupts protein–protein interactions | \n\t\t\tVariable cell removal; loss of collagen content, although effect on mechanical properties was minimal | \n\t\t
Hypotonic and hypertonic solutions | \n\t\t\tCell lysis by osmotic shock | \n\t\t\tEfficient for cell lysis, but does not effectively remove the cellular remnants | \n\t\t
EDTA, EGTA | \n\t\t\tChelating agents that bind divalent metallic ions, thereby disrupting cell adhesion to ECM | \n\t\t\tNo isolated exposure, typically used with enzymatic methods (e.g., trypsin) | \n\t\t
Enzymatic | \n\t\t||
Trypsin | \n\t\t\tCleaves peptide bonds on the C-side of Arg and Lys | \n\t\t\tProlonged exposure can disrupt ECM structure, removes laminin, fibronectin, elastin, and GAGs | \n\t\t
Endonucleases | \n\t\t\tCatalyze the hydrolysis of the interior bonds of ribonucleotide and deoxyribonucleotide chains | \n\t\t\tDifficult to remove from the tissue and could invoke an immune response | \n\t\t
Exonucleases | \n\t\t\tCatalyze the hydrolysis of the terminal bonds of ribonucleotide and deoxyribonucleotide chains | \n\t\t\t\n\t\t |
Overview of decellularization methods (Thomas W. Gilbert)
Today, many decellularization grafts were applied in clinical treatment such as acellular valve, vascular and some are commercial include: SurgisSIS (porcine small intestinal submicosa), Alloderm (human dermis), ACell (porcine urinary bladder)… Small intestine contains four layers such as mucosa, submucosa, muscularis externa and serosa. Small intestine submucosa (SIS) is a submucosa tissue between mucosa and muscularis externa. SIS is isolated from small intestine by mechanically removing of internal mucosal layer and outer muscular layer. Then, SIS is processed step-by-step with 0.1% peracid acid, 0.05% gentamycin and sterilized using 2500 kRad gamma irradiation. SIS is consisted of collagen, proteoglycan, glycosaminoglycan, glycoprotein and growth factor (VEGF, FGF-2 …). Most of these components can be preserved in extracellular matrix after decellularizing. In tissue engineering, SIS is used as soft tissue grafts such as vascular, skin or used for reconstructing genitourinary, ligament tissue [62].
Atherosclerosis is the most dangerous cardiovascular disease. Atherosclerosis is a condition in which vascular wall is harden, thicken because of fatty lipid accumulation. Atherosclerosis is responsible for millions of death all the world every year. In 2004, seventeen million people passed away because of cardiovascular disease all over the world (29% world total death). More than 7 million people are killed by coronary disease (9.6% world total death) (WHO). Heart American Association, in 2006, coronary disease was responsible for 17.6% death in America. One out of six American died for coronary disease [63].
Today, there are some methods to cure atherosclerosis including drug treatment, coronary artery bypass surgery and angioplasty. However, these methods can not cure this condition completely. More than 30% patients don’t have qualified autogenous vascular grafts. Therefore, artificial grafts are made to overcome some current drawbacks.
Artifical graft can be classified into synthetic and biologic scaffold. Synthetic scaffolds include undegradable polymers such as Dacron, ePTFE and degradable polymers such as polylactic acid (PLA) and polyglycolic acid (PGA). Some advantages of synthetic polymers are easy processing, high initial strength, however, they have some disadvantages such as slow recellularization, poor ability to support remodeling. Biologic vascular grafts are either obtained by manipulating native proteins of vascular vessels such as collagen, elastin… or by decellularizing vascular vessels. Decellularized native vascular vessels have some advantages such as slow cost, reduction of graft rejection and immune reaction [64].
In the early time, decellularization studies were tested without cell-seeding. In 1990, Lantz et al use SIS (small intestine submucosa) as small-diameter arterial graft in18 dogs. 48 weeks after surgery, 28 vascular grafts (75%) were patent and tree dogs can survive from 76 to 82 weeks after implantation. This result suggested that small intestinal submucosa can be used in small diameter vascular tissue engineering [65]. In 2001, Hodde et al demonstrated that porcine SIS ECM extract containing vascular endothelial growth factor (VEGF) with concentration of 0.77 ng VEGF/g SIS [66]. In the same year, Nemcova et al isolated and decellularized SIS from bovine and porcine. Nine decellularized SIS grafts were implanted into femoral arteries of five male mongrel dogs. After 9 weeks, eight grafts remained patent and some kinds of cell such as endothelial cells (ECs), smooth muscle cells (SMCs) invaded into the grafts. No evidence of inflammatory and aneurysmal symptom was observed during the experimental time [67].
However, the main disadvantage of vascular graft is thrombus which ussually occurs immediately in vascular graft lumen after implantation. This process can lead to graft failure and threaten patients’ survival. In order to solve this problem, ECs are used as anti-thrombus agent in vascular graft lumen, additionally, SMCs are used to improve mechanical strength of vascular graft. Broschel et al decellularized and recellularized rat iliac arterial grafts. Iliac arterial grafts were decellualrized by glycerin, SDS for 12 hours. Then, decellularized iliac grafts were recellularized with adult rat heart ECs and implanted to femoral arteries of allogeneic rats without systemic anticoagulation injection. After 4 weeks, 2 of 7 control grafts (29%) were patent (decellularized grafts without recellularization) and 8 of 9 (89%) experimental grafts (decellularized grafts with recellularization) maintained blood vessel patent [68]. Consequently, this experiment result proved excellent function of ECs in vascular tissue engineering. Some researchers on vascular tissue engineering seeded autologous vascular cells including ECs, SMCs and fibroblasts to make autologous tissue engineered vascular. At the same time, the appearance of bioreactor systems sped up development of vascular tissue engineering. Niklason developed a pulsatile perfusion bioreactor system in 2001. Bioreactor contained a stirbar, a lid for gas exchange and one (or two) silicone tubing(s). Porous vascular scaffolds were threaded over silicone tubing. Bovine aortic SMCs at the passage 2 or 3 were pipetted onto the outer surface of the scaffolds, then bioreactors were slowly rotated and removed to incubator with 10% CO2, 100% humidity, and 37oC. Each silicone tubing was linked to a pulsatile perfusion system operated at 165 beats/minute and 260/-30 mmHg. After 8 weeks culture, under nonpulsatile condition, SMC growth was on the outer surface of tubing scaffold and poorly organized. Under pulsatile condition, SMC distribute homogenously in scaffold wall from outer to inner similar to native structure [69].
Nowaday, some decellularized vascular grafts can be obtained from human. Human umbilical veins and arteries can be used in decellularization experiment. Human umbilical arteries were isolated and completely decellularized by Gui et al in 2009. Decellularized umbilical arteries preserved intact collagen matrix and mechanical properties, burst pressure had no signicant change from native form. Decellularized artery graft remained patent after 8 weeks surgery [70]. In 2005, Joel Daniel et al processed human umbilical vein by automated dissection. Human umbilical cord was inserted a mandrel into vein and frozen to -20 and -80oC, human umbilical cord was maintained for 12 hours at least at this temperature. The mandrel was lathed with rotation speed of 2900 rpm, cutting depth of 750 µm. Then, human umbilical vein was decellularized with 1% (w/v) SDS. The result showed that the treated human umbilical vein contained no cell; burst pressure results were 972.8 ± 133.8 mmHg (972/1082), compliance results were 5.7 ± 1.3% over 80 – 120 mmHg. Vascular smooth muscle cells can adhere, proliferate and migrate on the surface of dHUV [71]. Tran Le Bao Ha et al carried out a research on decellularization method for HUVs. HUVs were isolated by manual dissection. HUVs were decellularized by distilled water or NaCl 3M or SDS 1%. The result confirmed that the combination between SDS 1% (24h) and NaCl 3M (24h) showed the best effective on cellular elimination.
Human umbilical cord (a) and human umbilical vien (b)
HE staining of HUV with different decellularization methods. Control (a), distilled water (b), NaCl 3M (c), SDS 1% 24h (d), SDS 1% 36h (e), SDS 1% 24. and NaCl 3M 24h (f)
One of the most important properties of ECM is its functional diversity. ECM has been reported to support and enhance for adhesion, migration, proliferation of cells as well as to create stem cell niches
For this purpose, human foreskin fibroblasts are isolated, plated and cultured until third passage (see Figure 8). Fibroblasts are maintained in culture medium until reaching 80% confluency and stimulated to synthesize ECM by culture medium supplemented ascorbic acid. Matrices are denuded of cells and cellular remnants are removed by using Triton X-100, NH4OH and DNase.
Then, ECM coated culture surfaces are tested by staining with PI to access DNA remnant (see Figure 9), with H&E and PAS in order to characterize component of ECM. The results suggest that fibroblast-derived three-dimensional matrix was determined to be free of cellular constituents and still remain attached to the culture surface. The conducted matrices were washed and covered with PBS; and stored at 40C. Under these conditions, biological activity (for example, induction of cell attachment, proliferation…) was reported to be well-preserved for up to 6 months.
In order to evaluate the quality of the ECM, the test of cell rapid attachment ability is performed. Cell proliferation on ECM is also assessed. Results showed that the cultured cells attached and proliferated on ECM coated surfaces faster than on ECM non-coated surfaces.
Fibroblasts are in cultured surfaces (a) and are stimulated to synthesize ECM (b)
ECM stained PI before (a) and after (b) using DNase
Culture surface before (a) and after (b) is denuded cells
In modern society, many diseases have been increasing in human because of pollution, accident, lifestyle... The mutilation in human body leads to expand the needs of replacing tissues/organs. However, the available source of tissues/organs is limited. Creating artificial tissues/organs for replacing damaged, dysfunctional tissues/organs becomes a big discipline on material science. Naturally derived biomaterials have been studied and applied in clinical applications as artificial tissues/organs because they are capable of supporting cell proliferation, biodegradability and remodeling tissues. Although the current results have not completely satisfy the clinical demand, the potential applications of naturally derived biomaterials are still highly considered, therefore, research on this field have now being taken place all over the world.
COVID-19 outbreak reminds us of nationwide lockdowns, home-confinement, overburdened hospitals, country wide curfews, violent and chaotic health-facilities and domestic scenario and gradual but steady rise of stigmatizing factors like fear of isolation, racism, discrimination and marginalization cumulatively intensifying the unavoidable stress, fear, anxiety, depression, poor life quality, substance abuse resulting in enhanced mortality risk particularly among the health-compromised population [1, 2, 3] . Survival through this turmoil highlighted the elemental role of mental health organizations, individual healthcare and research, social media, awareness programs at both personal and community levels in significantly combating the world’s psychosocial distress [4, 5, 6]. Although, COVID-19 bears the brunt of rising mental illness, a clear understanding of world’s disease response would certainly aid in improving our medical emergencies and coping skills around the world.
Psychosocial impact of COVID-19 has been widely discussed in literature but what remains untouched as with all other parts of psychiatry is that the cause of impact has not been discussed [2, 3, 4, 7]. It is often easy to label each and every disorder that a psychiatrist is seeing in COVID-19 patients or in patients or healthy beings having the effects somehow related to the pandemic to label as evidence. It thus becomes imperative to discuss the impact that a society is having through a unique lens that widens the perspective relating to the cause and consequence of each and every response that happens to the disease process. To help with this perspective, we shall use the model described by Kunal in his paper on hypothesis spanning the diversities of psychiatry and normalcy, in the section on pathophysiology [8]. But before we shall delve into this new perspective, we shall see what evidence has to say for the psychosocial impact of the pandemic.
To analyze the psychosocial impact of COVID19.
To delineate the process of impact, by looking into its pathophysiology.
Literature was reviewed from PubMed and Google Scholar to find out articles describing the psychosocial impact of COVID19. A search was also carried out to find the explanations pertaining to different behavior of an individual to a given stress in order to justify the impact and explore management options.
The compilation of effects was studied and a way was formulated to better understand the psychosocial impact (Figure 1).
Algorithm to show mapping of psychosocial impact of COVID19 based on literature.
When talking about social impact one needs to understand how a society is formed and functions. Society is like a big web of individuals who are inter-related because of friendship, families or workplace. There are visible and some invisible connections where a person relates to another person or set of person based on ideologies, principles or practice like in name of color, caste, creed or nationality. Invisible connections that often turn into sentiments of positivity or negativity. Stigmatization based on religion, social status and nationality has been a core of social discrimination in this pandemic [2, 9].
A revolution this pandemic brought was the strengthening of visible connections. Spending more time with families had both positive and negative impact. The bonding of families increased however spending too much of time together also had reportedly increased rates of divorce among couples [2, 10]. Veliyannoor coined this as ‘return of the repressed’ where old conflicts re-emerged during the lockdown due to prolonged forced togetherness between couples [10].
Joshi noticed in her review of psycho-social impact of COVID19 in India that adolescents and young people found it restricting and frustrating to be monitored by parents. Also, she pointed out how the conventional gender role expectations in an Indian society resulted in their overburdening leading to frustration and anger bursts. Family dynamics were also influenced by forced togetherness or separation [2].
Based on age, Dubey
Education builds society into a framework as we see and want to see. Conventional education has long been debated but has never been challenged at this scale. Pandemic showed the world a new perspective towards education in terms of online education system. The cellphones which were till an year back not allowed in many schools have become an integral part of education system and has also given food for thought as whether our thought process of keeping these at bay mattered? Long term effects are yet to be seen and short term reports have shown that not going to school may have psychologically affected children but again what impact it shall have on overall growth of a child is yet to be found out. Educational impact can be understood in three different ways- one on parents, one on children and one on teachers. While parents are still doubtful of new education system, it may be difficult for teachers too as maintaining the sanctity and discipline of class could not be strictly possible in addition to the fact that it is not possible to look after every student in classroom now. Also, teachers may face a constant struggle for validation as this technique of teaching is new for many and they might be under supervision of hundreds of parents who might be listening to what they speak. For students the learning experience has changed, as the way they interact with teachers and fellows have been changed. But not only the schools, even the crucial education deliveries have changed like that of medical training program even though examination patterns have remained nearly same. How this will change the social dynamics is yet to be seen in future [12, 13, 14].
The increasing role of social media cannot be overlooked in context of COVID19. The major role it played in pandemic has both positive and negative sides. Positive side includes spreading awareness, helping out friends and locals in times of need like shortage of supplies, finding hospitals etc. seeking help in times of distress (which inculcated a sense of hope in lockdown and quarantines) and enhance bonding among people. But nothing comes without a cost, like enhanced bonding promoted stigmatization of particular community or nationality, outbursts of racism and therapeutic misadventures tried by lay people misleading the general public about treatment or prophylaxis. Coronavirus ‘infodemic’ is another concern that was created on social media in which panic was created by laying out unchecked mind-boggling rumors [15]. Also it helped to promote the disobedience attitude among people who failed to recognize the disease as pandemic and labeled it as hoax created by world governments to divert the commoners from daily problems.
Economic profiles can lead to formation of two groups in society- one whose livelihood has been greatly affected like those of laborers or small businessmen and another those whose work were less affected like those of HCPs, government servants, big businessmen and those working in private sector companies that still ran work-from home. The latter group had lesser financial implications, while former group went nearly bankrupt. Financial implications along with separation from families led migrant workers into great despair. Daily wage earners could not get enough wages to support family. Movement in groups and living in poor conditions predisposed them to infection and stigmatization of being carriers. The ill planned distribution of health services and expense of medical treatment led to hoarding of supplies, black marketing and other health crimes [9].
This section shall be incomplete without the mention of social and political revolutions. Revolution does not see the face of pandemic to break out. A very crucial aspect of any society is opposition to the laid rules. Protest is common in democracy as problems do not cease to exist in democracy. These people are marginalized and unheard, complicated by existence of pandemic, media often ignore either their plea or glorify their ignorance towards safety guidelines like wearing of masks, grouping together or following proper guidelines. The protestors may also have a feeling of vulnerability and learned helplessness as described for those people who have patients admitted in hospitals and must break social protocols to be able to provide proper care.
Social inequities like belonging to marginalized sections with poor health facilities often expose to chronic diseases leading to higher mortality when contracting COVID19 which has been reported by Osofsky
Based on health care the society was clearly divided into two strata during the pandemic- one providing health service and the other at receiving end. Patients developed anxiety, depression, panic, irritability, while the attendants developed a sense of uncertainty, helplessness, worthlessness and a fear of infection [1, 7, 17]. The kin of those who died developed prolonged grief, depression, substance abuse, and stigma related to spread of infection [7]. Inability to do a proper funeral further accentuated the problem. The visuals of ill treatment of dead bodies or mass funerals can further develop a sense of despair in individuals who could not say a proper good-bye to family and same could be for the families too. Another perspective could be for those persons who have their patients admitted in hospitals. They might develop fear of contracting the infection on one hand and on other might feel helpless in not being able to maintain social restrictions thus depicting learned helplessness [7].
Another special set of patients that needs special mention is psychiatric patients. Violence, addiction and disturbed routine follow-up were common issues faced by them. Xiao proposed novel approach of structure letter therapy while Saladino
On the other hand, psychological impact on HCWs could be very different. They felt overworked, unable to attend to family duties and guilt of not being able to save the patients. Those not involved in direct care of patients could have felt worthlessness for not being able to help [7]. Overwork can lead to burnouts, depression and anxiety [5]. Lack of facilities can also produce secondary traumatic stress disorder where HCWs have to decide who can access and who cannot access the facilities [20]. Lai
Trainee doctors and medical graduates were also affected in their training. Reduced patient contact led to decrement in development of skills [22, 23]. Surgeons faced redeployment and decreased operation theater postings which in turn affected their surgical skill development [24]. Adaptation to online learning and problems with submission of dissertation was also a challenge and all these stress added to psychological impact on them [24, 25, 26]. A positive statement was that with online classes and posting in backup teams, these trainees got time to let the steam off and de-stress them. The training of not only health care workers but also every sector suffered tremendously thus also creating a question mark for not only training modules in past but also in future of the trainees trained during COVID19. Although, detailed discussion is out of scope of this chapter.
Romm
So what caused this impact? This is not a direct impact of the virus. Let us consider COVID-19 as a stress- a newly found one (Figure 2), according the “third-eye model’ [8]. It is a new experience and in some way have affected every person even the dead who could not get proper funerals and good-byes. And indeed that too has affected the living. Figure 3 shows normal response to stress. Whenever a stress strikes, it tends to push the brain cycle into NREM phase. A person always tries to end it and mount a REM response based on conditionings in brain. Conditionings are the coping skills that a person had learned during lifetime, so technically children will have lesser conditioning than adults and young will have lesser conditioning than elders. Thus problems with children will focus on what they do as in difficulty in concentrating, boredom, irritability, restlessness, nervousness, sense of loneliness, uneasiness, and worries, while adults would have anxiety, depression, burnouts, loneliness, panic, and substance use disorders.
Algorithm showing basic response to any stress (further algorithm is given in
Algorithm showing response when a patient can control the thoughts provoking mental stress and possible responses.
But not all people suffer from psychological impact at a given time. If one believes that the stress shall be controlled and it is indeed controlled then the behavior is more towards normalcy like with those whose income was not affected- pensioners, HCPs and government employees. Even though many times they worked under the fear of getting infected sometimes this fear was casual but at times where situation was not under control like working with limited supplies or exposure to large number of COVID patients or working long shifts, this could turn into anxiety or panic.
Whenever the stress is identified as stress like a person believes that abnormal response to COVID19 is validated, a justification happens for stress. If this justification is for a process that just a mirror image of what a person usually saw this could lead to adjustment disorders giving sense of irritability or loneliness to the person but if it backed by emotional turmoil or magnification where COVID19 is seen to impact the lives of people like never before killing thousands, breaking families, blowing the finances of families or simply loss of very close one who had no disease otherwise can lead to PTSD. It is often the effect of COVID19 that tends to cause adjustment disorder and PTSD than itself. Multiplied by isolation and long stays at home it is not difficult to imagine that bereavement reaction can also be prolonged.
Now all of this may comprise a normal response to a pandemic because the person still has some control over thoughts of stress and often the problem would lay on impact of stress as in death of a person, inability to help others, financial breakdowns and social cut-offs rather than COVID19 itself. Once the stress is incorporated in psyche of an individual, the principle of inertia decides as to whether the psyche can overcome the persistence of thoughts. If it tries to overcome it, it searches for a manipulation (remember here the thoughts have persisted and cannot be simply overthrown by mounting a response as explained in above text). From Figure 4, we can see if the manipulation is specific to stress it can be lead to anxiety if the manipulation is feasible for psyche but patient cannot do that. Another common way to mount a response is to mount a response which is very feasible to psyche like washing hands repeatedly as in OCD or shifting to over smoking or over drinking for stress relief. Sometimes this substance abuse can be an excuse for stress where the patient may be trying to gain sympathy or needs a leave from workplace and do not want to malinger which indeed can be very common due to non-specific and objective complaints of COVID-19 which can be practically anything from myalgia to loose stools. If the patient is already not ill can present with a spectrum of somatization and this could be for same reason except for the fact that malingering or claiming false symptoms is not feasible to psyche, i.e. the person does not want to malinger or do substance abuse which can be due to ethics, emotions or rationality.
Algorithm showing response when a patient cannot control the thoughts provoking mental stress and still tries to find possible ways to control its inception in thought process.
Although, the above pathologies are not uncommon but another common response pattern is where the NREM state induced by stress is only controlled to some extent or cannot be controlled at all (Figures 2 and 5). In any of these cases, giving up to brain cycle is difficult as these people would have otherwise stronger will to control the brain. COVID19 is a pandemic and has affected everyone in some extent and thus does not target any specific group and is a boon for others. This feeling helps the person to control rather than be controlled by brain. Dysthymia has a definition to fulfill and can be underreported or can be diagnosed as depression by many. Low mood is not uncommon as it does not require any specific action from the person, it is like ‘going with the flow’.
Algorithm showing response when a patient cannot control the thoughts provoking mental stress and cannot find possible ways to control its inception in thought process.
Each and every response can be mapped and the therapy oriented at that, however it is beyond the scope of this chapter to discuss every disease and intervention in detail [8].
Studying the impact can help us delineate the stress parameters and try to streamline treatment modalities. Saladino
The visuals of people dying despite best health facilities puts the question forward for us as to what are we in front of nature? How much have we learned and how much we are we prepared for catastrophe? The rapid response and adaptability of general population cannot be overlooked. But it has left a deep impact with long term unforeseen effects. We shall not only study the effects in its past form but also anticipate further damage it can cause in late future. Other than teaching us the fact that we need a lot of medical preparedness, it also taught us that healthy individuals can also become transiently stressed and show symptoms which may not be deemed as psychologically fit. This blemish had to be erased to bring people to talk about psychological issues with even non-medico or non-psychiatrist friends and colleagues. Ignorance had been a bliss for ignorant but not for those who suffered in silence. It is the awareness that has brought us together in the face of crisis.
COVID19 came as wave and swayed the humanity. Different states of individual showed different responses to the same stress creating a gamut of responses which led to different socio-economic, socio-political, professional and psychosocial impact. Identification of stress, mapping of response and a better preparedness from experience can all help us in preventing and controlling it next time.
The authors declare no conflict of interest.
This is a brief overview of the main steps involved in publishing with IntechOpen Compacts, Monographs and Edited Books. Once you submit your proposal you will be appointed a Author Service Manager who will be your single point of contact and lead you through all the described steps below.
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Professor Derbel has published many articles in journals and collaborates intensively with IntechOpen Access Publisher as an editor.",institutionString:"Clinique les Oliviers",institution:null},{id:"300144",title:"Dr.",name:"Meriem",middleName:null,surname:"Braiki",slug:"meriem-braiki",fullName:"Meriem Braiki",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/300144/images/system/300144.jpg",biography:"Dr. Meriem Braiki is a specialist in pediatric surgeon from Tunisia. She was born in 1985. She received her medical degree from the University of Medicine at Sousse, Tunisia. She achieved her surgical residency training periods in Pediatric Surgery departments at University Hospitals in Monastir, Tunis and France.\r\nShe is currently working at the Pediatric surgery department, Sidi Bouzid Hospital, Tunisia. Her hospital activities are mostly concerned with laparoscopic, parietal, urological and digestive surgery. She has published several articles in diffrent journals.",institutionString:"Sidi Bouzid Regional Hospital",institution:null},{id:"229481",title:"Dr.",name:"Erika M.",middleName:"Martins",surname:"de Carvalho",slug:"erika-m.-de-carvalho",fullName:"Erika M. de Carvalho",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/229481/images/6397_n.jpg",biography:null,institutionString:null,institution:{name:"Oswaldo Cruz Foundation",country:{name:"Brazil"}}},{id:"186537",title:"Prof.",name:"Tonay",middleName:null,surname:"Inceboz",slug:"tonay-inceboz",fullName:"Tonay Inceboz",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/186537/images/system/186537.jfif",biography:"I was graduated from Ege University of Medical Faculty (Turkey) in 1988 and completed his Med. PhD degree in Medical Parasitology at the same university. I became an Associate Professor in 2008 and Professor in 2014. I am currently working as a Professor at the Department of Medical Parasitology at Dokuz Eylul University, Izmir, Turkey.\n\nI have given many lectures, presentations in different academic meetings. I have more than 60 articles in peer-reviewed journals, 18 book chapters, 1 book editorship.\n\nMy research interests are Echinococcus granulosus, Echinococcus multilocularis (diagnosis, life cycle, in vitro and in vivo cultivation), and Trichomonas vaginalis (diagnosis, PCR, and in vitro cultivation).",institutionString:"Dokuz Eylül University",institution:{name:"Dokuz Eylül University",country:{name:"Turkey"}}},{id:"71812",title:"Prof.",name:"Hanem Fathy",middleName:"Fathy",surname:"Khater",slug:"hanem-fathy-khater",fullName:"Hanem Fathy Khater",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/71812/images/1167_n.jpg",biography:"Prof. Khater is a Professor of Parasitology at Benha University, Egypt. She studied for her doctoral degree, at the Department of Entomology, College of Agriculture, Food and Natural Resources, University of Missouri, Columbia, USA. She has completed her Ph.D. degrees in Parasitology in Egypt, from where she got the award for “the best scientific Ph.D. dissertation”. She worked at the School of Biological Sciences, Bristol, England, the UK in controlling insects of medical and veterinary importance as a grant from Newton Mosharafa, the British Council. Her research is focused on searching of pesticides against mosquitoes, house flies, lice, green bottle fly, camel nasal botfly, soft and hard ticks, mites, and the diamondback moth as well as control of several parasites using safe and natural materials to avoid drug resistances and environmental contamination.",institutionString:null,institution:{name:"Banha University",country:{name:"Egypt"}}},{id:"99780",title:"Prof.",name:"Omolade",middleName:"Olayinka",surname:"Okwa",slug:"omolade-okwa",fullName:"Omolade Okwa",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/99780/images/system/99780.jpg",biography:"Omolade Olayinka Okwa is presently a Professor of Parasitology at Lagos State University, Nigeria. She has a PhD in Parasitology (1997), an MSc in Cellular Parasitology (1992), and a BSc (Hons) Zoology (1990) all from the University of Ibadan, Nigeria. She teaches parasitology at the undergraduate and postgraduate levels. She was a recipient of a Commonwealth fellowship supported by British Council tenable at the Centre for Entomology and Parasitology (CAEP), Keele University, United Kingdom between 2004 and 2005. She was awarded an Honorary Visiting Research Fellow at the same university from 2005 to 2007. \nShe has been an external examiner to the Department of Veterinary Microbiology and Parasitology, University of Ibadan, MSc programme between 2010 and 2012. She is a member of the Nigerian Society of Experimental Biology (NISEB), Parasitology and Public Health Society of Nigeria (PPSN), Science Association of Nigeria (SAN), Zoological Society of Nigeria (ZSN), and is Vice Chairperson of the Organisation of Women in Science (OWSG), LASU chapter. She served as Head of Department of Zoology and Environmental Biology, Lagos State University from 2007 to 2010 and 2014 to 2016. She is a reviewer for several local and international journals such as Unilag Journal of Science, Libyan Journal of Medicine, Journal of Medicine and Medical Sciences, and Annual Research and Review in Science. \nShe has authored 45 scientific research publications in local and international journals, 8 scientific reviews, 4 books, and 3 book chapters, which includes the books “Malaria Parasites” and “Malaria” which are IntechOpen access publications.",institutionString:"Lagos State University",institution:{name:"Lagos State University",country:{name:"Nigeria"}}},{id:"273100",title:"Dr.",name:"Vijay",middleName:null,surname:"Gayam",slug:"vijay-gayam",fullName:"Vijay Gayam",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/273100/images/system/273100.jpeg",biography:"Dr. Vijay Bhaskar Reddy Gayam is currently practicing as an internist at Interfaith Medical Center in Brooklyn, New York, USA. He is also a Clinical Assistant Professor at the SUNY Downstate University Hospital and Adjunct Professor of Medicine at the American University of Antigua. He is a holder of an M.B.B.S. degree bestowed to him by Osmania Medical College and received his M.D. at Interfaith Medical Center. His career goals thus far have heavily focused on direct patient care, medical education, and clinical research. He currently serves in two leadership capacities; Assistant Program Director of Medicine at Interfaith Medical Center and as a Councilor for the American\r\nFederation for Medical Research. As a true academician and researcher, he has more than 50 papers indexed in international peer-reviewed journals. He has also presented numerous papers in multiple national and international scientific conferences. His areas of research interest include general internal medicine, gastroenterology and hepatology. He serves as an editor, editorial board member and reviewer for multiple international journals. His research on Hepatitis C has been very successful and has led to multiple research awards, including the 'Equity in Prevention and Treatment Award” from the New York Department of Health Viral Hepatitis Symposium (2018) and the 'Presidential Poster Award” awarded to him by the American College of Gastroenterology (2018). He was also awarded 'Outstanding Clinician in General Medicine” by Venus International Foundation for his extensive research expertise and services, perform over and above the standard expected in the advancement of healthcare, patient safety and quality of care.",institutionString:"Interfaith Medical Center",institution:{name:"Interfaith Medical Center",country:{name:"United States of America"}}},{id:"93517",title:"Dr.",name:"Clement",middleName:"Adebajo",surname:"Meseko",slug:"clement-meseko",fullName:"Clement Meseko",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/93517/images/system/93517.jpg",biography:"Dr. Clement Meseko obtained DVM and PhD degree in Veterinary Medicine and Virology respectively. He has worked for over 20 years in both private and public sectors including the academia, contributing to knowledge and control of infectious disease. Through the application of epidemiological skill, classical and molecular virological skills, he investigates viruses of economic and public health importance for the mitigation of the negative impact on people, animal and the environment in the context of Onehealth. \r\nDr. Meseko’s field experience on animal and zoonotic diseases and pathogen dynamics at the human-animal interface over the years shaped his carrier in research and scientific inquiries. He has been part of the investigation of Highly Pathogenic Avian Influenza incursions in sub Saharan Africa and monitors swine Influenza (Pandemic influenza Virus) agro-ecology and potential for interspecies transmission. He has authored and reviewed a number of journal articles and book chapters.",institutionString:"National Veterinary Research Institute",institution:{name:"National Veterinary Research Institute",country:{name:"Nigeria"}}},{id:"158026",title:"Prof.",name:"Shailendra K.",middleName:null,surname:"Saxena",slug:"shailendra-k.-saxena",fullName:"Shailendra K. Saxena",position:null,profilePictureURL:"https://s3.us-east-1.amazonaws.com/intech-files/0030O00002bRET3QAO/Profile_Picture_2022-05-10T10:10:26.jpeg",biography:"Professor Dr. Shailendra K. Saxena is a vice dean and professor at King George's Medical University, Lucknow, India. His research interests involve understanding the molecular mechanisms of host defense during human viral infections and developing new predictive, preventive, and therapeutic strategies for them using Japanese encephalitis virus (JEV), HIV, and emerging viruses as a model via stem cell and cell culture technologies. His research work has been published in various high-impact factor journals (Science, PNAS, Nature Medicine) with a high number of citations. He has received many awards and honors in India and abroad including various Young Scientist Awards, BBSRC India Partnering Award, and Dr. JC Bose National Award of Department of Biotechnology, Min. of Science and Technology, Govt. of India. Dr. Saxena is a fellow of various international societies/academies including the Royal College of Pathologists, United Kingdom; Royal Society of Medicine, London; Royal Society of Biology, United Kingdom; Royal Society of Chemistry, London; and Academy of Translational Medicine Professionals, Austria. He was named a Global Leader in Science by The Scientist. He is also an international opinion leader/expert in vaccination for Japanese encephalitis by IPIC (UK).",institutionString:"King George's Medical University",institution:{name:"King George's Medical University",country:{name:"India"}}},{id:"94928",title:"Dr.",name:"Takuo",middleName:null,surname:"Mizukami",slug:"takuo-mizukami",fullName:"Takuo Mizukami",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/94928/images/6402_n.jpg",biography:null,institutionString:null,institution:{name:"National Institute of Infectious Diseases",country:{name:"Japan"}}},{id:"233433",title:"Dr.",name:"Yulia",middleName:null,surname:"Desheva",slug:"yulia-desheva",fullName:"Yulia Desheva",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/233433/images/system/233433.png",biography:"Dr. Yulia Desheva is a leading researcher at the Institute of Experimental Medicine, St. Petersburg, Russia. She is a professor in the Stomatology Faculty, St. Petersburg State University. She has expertise in the development and evaluation of a wide range of live mucosal vaccines against influenza and bacterial complications. Her research interests include immunity against influenza and COVID-19 and the development of immunization schemes for high-risk individuals.",institutionString:'Federal State Budgetary Scientific Institution "Institute of Experimental Medicine"',institution:null},{id:"238958",title:"Mr.",name:"Atamjit",middleName:null,surname:"Singh",slug:"atamjit-singh",fullName:"Atamjit Singh",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/238958/images/6575_n.jpg",biography:null,institutionString:null,institution:null},{id:"333753",title:"Dr.",name:"Rais",middleName:null,surname:"Ahmed",slug:"rais-ahmed",fullName:"Rais Ahmed",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/333753/images/20168_n.jpg",biography:null,institutionString:null,institution:null},{id:"252058",title:"M.Sc.",name:"Juan",middleName:null,surname:"Sulca",slug:"juan-sulca",fullName:"Juan Sulca",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/252058/images/12834_n.jpg",biography:null,institutionString:null,institution:null},{id:"191392",title:"Dr.",name:"Marimuthu",middleName:null,surname:"Govindarajan",slug:"marimuthu-govindarajan",fullName:"Marimuthu Govindarajan",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/191392/images/5828_n.jpg",biography:"Dr. M. Govindarajan completed his BSc degree in Zoology at Government Arts College (Autonomous), Kumbakonam, and MSc, MPhil, and PhD degrees at Annamalai University, Annamalai Nagar, Tamil Nadu, India. He is serving as an assistant professor at the Department of Zoology, Annamalai University. His research interests include isolation, identification, and characterization of biologically active molecules from plants and microbes. He has identified more than 20 pure compounds with high mosquitocidal activity and also conducted high-quality research on photochemistry and nanosynthesis. He has published more than 150 studies in journals with impact factor and 2 books in Lambert Academic Publishing, Germany. He serves as an editorial board member in various national and international scientific journals.",institutionString:null,institution:null},{id:"274660",title:"Dr.",name:"Damodar",middleName:null,surname:"Paudel",slug:"damodar-paudel",fullName:"Damodar Paudel",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/274660/images/8176_n.jpg",biography:"I am DrDamodar Paudel,currently working as consultant Physician in Nepal police Hospital.",institutionString:null,institution:null},{id:"241562",title:"Dr.",name:"Melvin",middleName:null,surname:"Sanicas",slug:"melvin-sanicas",fullName:"Melvin Sanicas",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/241562/images/6699_n.jpg",biography:null,institutionString:null,institution:null},{id:"337446",title:"Dr.",name:"Maria",middleName:null,surname:"Zavala-Colon",slug:"maria-zavala-colon",fullName:"Maria Zavala-Colon",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"University of Puerto Rico, Medical Sciences Campus",country:{name:"United States of America"}}},{id:"338856",title:"Mrs.",name:"Nur Alvira",middleName:null,surname:"Pascawati",slug:"nur-alvira-pascawati",fullName:"Nur Alvira Pascawati",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Universitas Respati Yogyakarta",country:{name:"Indonesia"}}},{id:"441116",title:"Dr.",name:"Jovanka M.",middleName:null,surname:"Voyich",slug:"jovanka-m.-voyich",fullName:"Jovanka M. Voyich",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Montana State University",country:{name:"United States of America"}}},{id:"330412",title:"Dr.",name:"Muhammad",middleName:null,surname:"Farhab",slug:"muhammad-farhab",fullName:"Muhammad Farhab",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"University of Agriculture Faisalabad",country:{name:"Pakistan"}}},{id:"349495",title:"Dr.",name:"Muhammad",middleName:null,surname:"Ijaz",slug:"muhammad-ijaz",fullName:"Muhammad Ijaz",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"University of Veterinary and Animal Sciences",country:{name:"Pakistan"}}}]}},subseries:{item:{id:"26",type:"subseries",title:"Machine Learning and Data Mining",keywords:"Intelligent Systems, Machine Learning, Data Science, Data Mining, Artificial Intelligence",scope:"The scope of machine learning and data mining is immense and is growing every day. 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