Number, size and type of tumors in DMBA groups at 20th week.
\r\n\tThe LED can be lingering further into three major categories are (i) Traditional inorganic LEDs, (ii) Organic LEDs (Small Molecule OLED, Polymer LED, Passive Matrix OLED Active Matrix OLED), (iii) High brightness LEDs, (iv) Deep-UV LEDs, (v) Active Matrix Organic Light-Emitting Diodes (AMOLED).
",isbn:"978-1-83968-886-7",printIsbn:"978-1-83968-885-0",pdfIsbn:"978-1-83968-887-4",doi:null,price:0,priceEur:0,priceUsd:0,slug:null,numberOfPages:0,isOpenForSubmission:!1,hash:"97e861d1556a639f0e5cc6ee8bdb0a0f",bookSignature:"Prof. Jagannathan Thirumalai",publishedDate:null,coverURL:"https://cdn.intechopen.com/books/images_new/10559.jpg",keywords:"Aluminum Gallium Arsenide, Gallium Arsenide Phosphide, Indium Phosphide, Thin-Film-Display, Organic Rare-Earth Complexes, Colour Rendering Index, High Brightness Leds, Luminous Control, Air Purification, Skin Therapy, Organic Compounds Form the Electroluminescent Material, Specific Type of Thin-Film-Display",numberOfDownloads:30,numberOfWosCitations:0,numberOfCrossrefCitations:0,numberOfDimensionsCitations:0,numberOfTotalCitations:0,isAvailableForWebshopOrdering:!0,dateEndFirstStepPublish:"September 21st 2020",dateEndSecondStepPublish:"October 19th 2020",dateEndThirdStepPublish:"December 18th 2020",dateEndFourthStepPublish:"March 8th 2021",dateEndFifthStepPublish:"May 7th 2021",remainingDaysToSecondStep:"5 months",secondStepPassed:!0,currentStepOfPublishingProcess:4,editedByType:null,kuFlag:!1,biosketch:"As an expert in the optoelectronics and nanotechnology area, Dr.Thirumalai has been invited to examine several MSc and Ph.D. theses, invited to give a talk in various forums, and to review papers for international and national journals.",coeditorOneBiosketch:null,coeditorTwoBiosketch:null,coeditorThreeBiosketch:null,coeditorFourBiosketch:null,coeditorFiveBiosketch:null,editors:[{id:"99242",title:"Prof.",name:"Jagannathan",middleName:null,surname:"Thirumalai",slug:"jagannathan-thirumalai",fullName:"Jagannathan Thirumalai",profilePictureURL:"https://mts.intechopen.com/storage/users/99242/images/system/99242.png",biography:"Dr. J. Thirumalai received his Ph.D. from Alagappa University, Karaikudi in 2010. \n\nHe was awarded the Post-doctoral Fellowship from Pohang University of Science and Technology (POSTECH), Republic of Korea, in 2013.\nHe worked as an Assistant Professor of Physics, B.S. Abdur Rahman University, Chennai, India (2011 to 2016). \nCurrently, he is working as an Assistant Professor & Head of the Department of Physics, SASTRA Deemed to be University, Kumbakonam (T.N.), India. \n\nHis research interests focus on luminescence, self-assembled nanomaterials, thin-film optoelectronic devices & Supercapacitors. \n\nHe has published more than 60 SCOPUS/ISI indexed papers, 11 book chapters, and he edited 5 books. He is serving as a member in various national and international societies. Currently, he is acting as a principal investigator for a funded project towards the application of luminescence-based thin-film optoelectronic devices, funded by the Science and Engineering Research Board (SERB), India. \nAs an expert in optoelectronics and nanotechnology area, he has been invited to examine several MSc and Ph.D. theses, invited to give a talk in various forums and to review papers for international and national journals.",institutionString:"SASTRA University",position:null,outsideEditionCount:0,totalCites:0,totalAuthoredChapters:"10",totalChapterViews:"0",totalEditedBooks:"6",institution:{name:"SASTRA University",institutionURL:null,country:{name:"India"}}}],coeditorOne:null,coeditorTwo:null,coeditorThree:null,coeditorFour:null,coeditorFive:null,topics:[{id:"11",title:"Engineering",slug:"engineering"}],chapters:[{id:"74673",title:"Economic Applications for LED Lights in Industrial Sectors",slug:"economic-applications-for-led-lights-in-industrial-sectors",totalDownloads:30,totalCrossrefCites:0,authors:[{id:"150046",title:"Prof.",name:"Muhammad M.A.S.",surname:"Mahmoud",slug:"muhammad-m.a.s.-mahmoud",fullName:"Muhammad M.A.S. Mahmoud"}]}],productType:{id:"1",title:"Edited Volume",chapterContentType:"chapter",authoredCaption:"Edited by"},personalPublishingAssistant:{id:"297737",firstName:"Mateo",lastName:"Pulko",middleName:null,title:"Mr.",imageUrl:"https://mts.intechopen.com/storage/users/297737/images/8492_n.png",email:"mateo.p@intechopen.com",biography:"As an Author Service Manager my responsibilities include monitoring and facilitating all publishing activities for authors and editors. From chapter submission and review, to approval and revision, copyediting and design, until final publication, I work closely with authors and editors to ensure a simple and easy publishing process. I maintain constant and effective communication with authors, editors and reviewers, which allows for a level of personal support that enables contributors to fully commit and concentrate on the chapters they are writing, editing, or reviewing. I assist authors in the preparation of their full chapter submissions and track important deadlines and ensure they are met. I help to coordinate internal processes such as linguistic review, and monitor the technical aspects of the process. As an ASM I am also involved in the acquisition of editors. Whether that be identifying an exceptional author and proposing an editorship collaboration, or contacting researchers who would like the opportunity to work with IntechOpen, I establish and help manage author and editor acquisition and contact."}},relatedBooks:[{type:"book",id:"5348",title:"Luminescence",subtitle:"An Outlook on the Phenomena and their Applications",isOpenForSubmission:!1,hash:"d982c49fed4423a0ea7367af4f917b82",slug:"luminescence-an-outlook-on-the-phenomena-and-their-applications",bookSignature:"Jagannathan Thirumalai",coverURL:"https://cdn.intechopen.com/books/images_new/5348.jpg",editedByType:"Edited by",editors:[{id:"99242",title:"Prof.",name:"Jagannathan",surname:"Thirumalai",slug:"jagannathan-thirumalai",fullName:"Jagannathan Thirumalai"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"6489",title:"Light-Emitting Diode",subtitle:"An Outlook On the Empirical Features and Its Recent Technological 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by"}},{type:"book",id:"72",title:"Ionic Liquids",subtitle:"Theory, Properties, New Approaches",isOpenForSubmission:!1,hash:"d94ffa3cfa10505e3b1d676d46fcd3f5",slug:"ionic-liquids-theory-properties-new-approaches",bookSignature:"Alexander Kokorin",coverURL:"https://cdn.intechopen.com/books/images_new/72.jpg",editedByType:"Edited by",editors:[{id:"19816",title:"Prof.",name:"Alexander",surname:"Kokorin",slug:"alexander-kokorin",fullName:"Alexander Kokorin"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}}]},chapter:{item:{type:"chapter",id:"64239",title:"The Wound Healing Responses and Corneal Biomechanics after Keratorefractive Surgery",doi:"10.5772/intechopen.81886",slug:"the-wound-healing-responses-and-corneal-biomechanics-after-keratorefractive-surgery",body:'\nOver the past 30 years, corneal refractive surgery has successfully corrected the refractive error for millions of patients. The spread of laser corneal refractive surgery is increasing the interest in the study of the safety and predictability. Many studies showed that the wound healing process influences the predictability and safety. Corneal wound healing is a major contributor to the success of refractive surgeries. Biological differences in wound healing responses are thought to be a major factor limiting the predictability of refractive surgery [1]. In some cases, mechanical instability or an abnormal wound healing process can lead to serious complications such as keratectasia or severe haze.
\nHence, it is important to investigate and understand the corneal biomechanics and wound healing process for a better vision after corneal refractive surgery.
\nCorneal refractive surgery changes the corneal curvature to correct the refractive error. The most common laser refractive procedures performed today are small incision lenticule extraction surgery (SMILE) [2, 3], femtosecond laser in situ keratomileusis (FS-LASIK), and surface ablation procedures, i.e., photorefractive keratectomy (PRK), laser epithelial keratomileusis (LASEK), and epi-LASIK [4]. With the development of the femtosecond laser, the SMILE surgery and FS-LASIK have become the most commonly used procedures in China for myopic subjects.
\nThe cornea is a highly specialized transparent avascular tissue and is composed of five layers. They are epithelium, stroma, Descemet’s membrane, and endothelium. Stroma is the main part of the cornea, and any factor that changes the corneal structure may obviously influence the biomechanical properties of the cornea [5, 6].
\nThe epithelium’s contribution to corneal biomechanics was significantly lower than that of the stroma with respect to the stiffness. Bowman’s membrane tissue is a transparent sheet of approximately 12 μm. It is acellular and is composed of densely packed collagen fibrils that are in random direction. The fibrils are continuous with those in the stroma, which is believed to stabilize the corneal curvature [5].
\nThe stroma constitutes nearly 90% of the corneal thickness. And its biomechanical properties are influenced by the collagen fibers and extracellular matrix (ECM), which further determine the corneal strength, shape, and transparency [7, 8].
\nThe stroma is a fibrous layer of lamellae made up of connective tissue. Interlamellar branching is more extensive in the anterior stroma than in the posterior stroma. The density of the collagen lamellae is higher, and their arrangement and directionality are more complicated anteriorly than posteriorly. The collagen lamellae in the corneal stroma are organized into a complex, highly intertwined three-dimensional meshwork of transversely oriented fibers, which contributes to the corneal shape and stromal stiffness. Another critical component for corneal stromal biomechanics is the ECM. The ECM is mostly composed of proteoglycans (PGs), which comprise a core protein and are located in the spaces among the collagen fibers in the corneal stroma. PGs play a critical role in collagen fibril assembly and spacing, and their mechanical importance may be greater than currently recognized [9].
\nDescemet’s membrane is approximately 10-nm thick and considered as a secretion of endothelial cells. The membrane is comprised of type IV collagen fibers. It is highly elastic and represents a barrier against punctures. Descemet’s membrane serves as an endothelial basement membrane. Bowman’s layer and Descemet’s membrane accounted for 20% of the bending rigidity of the cornea through the Finite element evaluation [5].
\nThe endothelium is composed of one layer of cells, which adhere to the Descemet’s membrane. The endothelium cells cannot regenerate after damage or aging, but can spread and enlarge to maintain the cornea clear and transparent, and further prevent the cornea from becoming hydrated. The corneal endothelium may indirectly affect the corneal stiffness by regulating corneal hydration. The loss of corneal endothelial cells will result in increased water absorption by the corneal stroma [8, 9].
\nIt is noteworthy that the corneal biomechanics and wound healing responses are linked in time and space. Specifically, the corneal biomechanics involves the stromal healing responses; the better stromal healing process will contribute to better corneal biomechanics after surgery and more stable visual results.
\nEpithelial wound healing involves three main steps: sliding, proliferation, and stratification of epithelial cells. Specifically, the epithelium cells migrate to the wound surface; then the cells increase and divide; lastly the cells cover the wound area and multiple layers of the epithelium are regenerated [9].
\nSome studies suggest that many cytokines are involved in the healing process including the epithelial growth factor (EGF), hepatocyte growth factor (HGF), keratinocyte growth factor (KGF), and transforming growth factor β (TGF-β). [9] These changes permit cells to migrate, establishing dynamic adhesion with other epithelial cells and extracellular matrix components. In the epithelial cells surrounding the wound edge, there is an increased expression of CD44. After the migration of epithelial cells, the phase of proliferation begins.
\nWhen the basal membrane is damaged, cytokines, neuropeptides, growth factors, chemokines, and matrix metalloproteinases can diffuse into the stroma and interact with keratocytes. These factors could stimulate the transformation of the keratocytes into myofibroblast cells [9]. One recent study [10] used the exosomes extracted from the epithelial cells, and cultured these exosomes with fibroblast cells; they found that the stroma cells transformed into myofibroblast cells with higher expression of TGF-β, CD63, and PDGF-B. This study indicates that the epithelial cells are very important for stromal wound healing, and they may use exosomes to transmit the wound healing signals in order to regulate the process [10].
\nAfter the epithelial and stromal damage, soluble mediators could be secreted through the epithelium and move to the stroma area. These molecules, like TGF-β and TSP-1, could stimulate the wound healing process and make keratocytes transform into myofibroblasts. The myofibroblasts lay down the ECM and generate alpha-smooth muscle actin (α-SMA) to close the wound. However, abnormal wound healing synthesizes excessive α-SMA, exerts traction forces across the ECM, and causes unorganized tissue architecture, haze, and regression after corneal refractive surgery [11, 12]. Only when the EBM is appropriately re-established, proper stromal levels of TGF-β and PDGF cause myofibroblast apoptosis, keratocyte repopulation, clearing of the abnormal ECM, and restoring of corneal transparency [13]. A delay in the regeneration of the EBM, due to damage, dystrophy, or elevated levels of MMP-2 and MMP-9, causes TGF-β and PDGF to continue entering the corneal stroma.
\nThe wound healing of the stroma is end when the collagen fibrils fully connected the wound edge. Activated cells migrate to the wound area. The keratocytes are changed through the reorganization of the cytoskeleton and the development of stress fibers and focal adhesion structures. Genes that encode fibronectin, metalloproteinases, and integrins are activated. The early matrix consists of fibronectin [14], which was conducive to cell migration and proliferation. Then the matrix is converted to a collagen and proteoglycan matrix that increases the tissue tensile strength and resilience. Growth factors increased stiffness and enhanced mechanical load through enhanced collagen fiber formation and cross-linking. The geometry of the collagen network will determine the mechanical properties of the wound. The collagen fiber diameter increases with time during the wound healing process and is related to tensile strength. Interweaving of collagen bundles between neighboring lamellae provides an important structural foundation for shear resistance and transfer of tensile loads between lamellae.
\nThe transformation of keratocytes into myofibroblasts is curial in the wound healing process. These cells are characterized by the expression of α-smooth muscle actin, stress fibers, and focal adhesion complexes. The microfilament bundles of myofibroblasts form stress fibers, and they contract and remodel the adjacent ECM. Myofibroblasts extend from the anterior stroma to the posterior stroma in a progressive manner. These cells develop fibrotic tissue for repair. Besides that, deposition of the ECM is beneficial for the matrix stiffening and global cellular stress. However, excess myofibroblasts cause the deposition of disorganized collagen and glycosaminoglycan [15]. The underlying mechanism for the interaction between myofibroblast cells and matrix is the focal adhesions. They play the role of a mechanotransduction system, transmitting the force generated by stress fibers to the surrounding ECM and also transducing the extracellular mechanical signals into the intracellular signaling. Further investigations are needed to find whether we could regulate the mechanotransduction system to influence the corneal wound healing process.
\nCorneal wound healing is important for the predictability and safety of corneal refractive surgery. The refractive outcome and its stability over time are strongly influenced by the corneal biomechanics and wound healing process. And the abnormal healing process or biomechanical instability could cause some complications after corneal refractive surgery.
\nRefractive regression is defined as a gradual loss of the attempted correction that limits prediction. Many studies showed loss of surgical outcome and the main cause seems to be the regression. The regression is mainly due to epithelial hyperplasia and stromal remodeling, two processes related to corneal wound healing. Refractive regression is a major challenge for myopia, especially for high levels of correction. Apoptosis, keratocyte proliferation, and myofibroblast cellular density have proved to be more intense following treatment for high myopia compared to treatments for mild myopia. Myofibroblasts are important effectors of regression. The changes of corneal biomechanics also induce the changes of the corneal shape and cause regression.
\nCorneal ectasia is a rare complication induced by the corneal refractive surgery. It may occur due to an insufficient residual stromal thickness or unidentified subclinical keratoconus. Many advanced examinations have been used clinically to exclude the potential subclinical keratoconus. Moreover, surgeons have also used many ways to preserve as thicker corneal thickness as possible. However, it is still difficult to avoid the onset of keratectasia. It may be because the corneal stiffness or biomechanics is different among individuals [16]. And the postoperative stromal tensile strength is different for each procedure. This indicates that the risk evaluations for ectasia should take the residual stromal bed thickness and corneal biomechanical properties into account. Moreover, biomechanical changes can manifest clinically as changes of the corneal shape and increased sensitivity to shape changes. The role of biomechanics is therefore important to consider in routine refractive procedures and in special cases where the biomechanical status of the cornea is abnormal.
\nCorneal haze refers to the cornea opacity. It is commonly seen in surface ablation surgeries. Haze can potentially form in the interface between the LASIK flap and the stromal bed or directly underneath the newly formed epithelium overlying the stromal tissue after PRK surgery [17]. In modern refractive surgery, haze tends to be mild and resolves very quickly. In extremely rare instances, haze can cause decreased visual acuity and increased glare.
\nAbnormal regulation of the wound healing process can result in the formation of stromal haze with decreased corneal crystalline expression, increased light scattering, and production of a disorganized extracellular matrix. Myofibroblasts are major contributors to corneal opacity with reduced expression of crystallin, greater secretion of type III collagen, and spread morphology. Over a period of time ranging from several weeks to several months, the myofibroblasts tend to gradually disappear through a series of remodeling processes. This process may be closely related to the expression of matrix metalloproteinases. These proteins are a family of proteolysis enzymes, which could degrade abnormal collagen fibrils. The cytokines, growth factors, and inflammatory mediators could also regulate the synthesis of metalloproteinase [18, 19, 20].
\nLaser refractive surgeries are effective for the correction of refractive errors. A better understanding of corneal wound healing from the biomechanical point of view is mandatory if refractive surgery is ever to achieve more predictable and safer refractive results.
\nOne of the first models of experimental carcinogenesis in animals was carried out by Salley in 1954 [1]. After applying various carcinogens, including 9,10-dimethyl-1,2 benzanthracene on the oral epithelium of Syrian hamsters for 3 months, Salley was able to verify the existence of squamous cell carcinomas (SCC) and lymphatic metastases. Subsequently, several authors have standardized this model and repeated it in order to achieve new knowledge about DMBA and the process of experimentally induced carcinogenesis [2, 3].
\n7,12-Dimethylbenz[a]anthracene (DMBA) is a polycyclic aromatic hydrocarbon which may, on its own, induce premalignant lesions and carcinomas within a few weeks after it is administered in mucosae [4, 5]. Commonly, it has been used in combination with ethanol as a promoter. DMBA is released after the combustion of tobacco -especially with cigarettes- or from animal fat when meat is grilled, and is also found in smoked meat and fish. This substance is, therefore, strongly involved in the carcinogenesis of oral, bronchiopulmonary and digestive tract malignancies [6, 7, 8, 9].
\nIn order to discover new drugs with cancer preventive effects, some authors have obtained promising outcomes at basic research level, specifically with substances such as salvinolic acid B [10] -derived from Salvia miltiorrhiza, used in fluorescence-, isothiocyanates [11] -synthetic derivatives of cabbages, squash, turnips and turnip greens-, Buddleja incana leaves, a tree that grows in Peru and Bolivia, Toona sinensis leaves [12], and olive oil extracts [9, 13, 14, 15]. In relation to the latter, and especially regarding its phenolic compounds, its antioxidant and cardiovascular protective properties are well known. In this sense, we have data stating that olive oil may act as preventive or inhibitor of carcinogenesis, and could even modify the nature of premalignant lesions that have already arisen, providing them a more benign and indolent behavior [14, 16].
\nTo experimentally test the inhibitory effect on the carcinogenesis process of Picual variety extra virgin olive oil.
\nForty-four male hamsters (Syrian Golden), 4-6 weeks old and weighing 60-80 g, were divided into four groups (two control and two experimental):
Experimental DMBA-PO group (DMBA from Sigma Chemical Co.), 12 animals. The oral pouches were brushed daily with paraffin oil (PO) in the first two weeks. Then, a solution of 0.5% DMBA and PO was administered on Mondays and Fridays for five weeks; alternatively, PO was applied on Tuesdays, Wednesdays and Thursdays at the same time. Thereafter, animals received daily PO until the twentieth week. All hamsters were fed with standard feed, and ad libitum water. \nFigure 1\n.
Experimental DMBA-OO group (DMBA from Sigma Chemical Co.), 12 animals. The oral pouches were brushed daily with OO in the first two weeks. Then, a solution of 0.5% DMBA and OO was administered on Mondays and Fridays every two days, for five weeks. Thereafter, animals received daily OO until the twentieth week. These hamsters were fed with standard feed, Picual variety ripe olives, and ad libitum water.
Control DMBA-PO group, 10 hamsters. The oral pouches were brushed daily with PO for twenty weeks.
Control DMBA-OO group, 10 hamsters. The oral pouches were brushed daily with extra virgin OO. Also, animals received diet with standard feed, Picual variety ripe olives, and ad libitum water for twenty weeks.
The hamsters.
The animals in each group were sacrificed after twenty weeks. Then, a macroscopic description and histological analysis of the induced tumors in the oropharynx, esophagus and stomach were performed.
\nA carcinogenic effect of 100% was assigned to the total number of induced tumors in hamsters who received DMBA-PO. The inhibitory effect in the DMBA-OO group was established by the percentage difference over 100. An inhibitory effect >50% was considered significant in the DMBA-OO group.
\nThis research work was examined and approved by the Ethical Committee for Animal Experimentation of the University of Seville (November 7, 2005). It met the requirements for experimentation with animals and was in accordance with the regulations in force in Spain and elsewhere the European Union.
\nThe groups exposed to DMBA showed tumors of different characteristics. Nonspecific lesions and others more suggestive of malignancy were found in the oral pouches of the DMBA-OO group, with a predominance of the former. These findings included leukoplakia, denudation of the mucosa, ulcerations or tumors with a benign appearance. However, tumors in DMBA-OO group were less common and smaller than in DMBA-PO (\nFigures 2A\n and \nB\n).
\nMacroscopic comparison of the digestive tract of two animals belonging to the DMBA-OO group (A) and DMBA-PO group (B). (A) The oral pouches, esophagus and stomach showed few small lesions and benign appearance. (B) The oral pouches, esophagus, and stomach showed tissue retractions, larger tumors and apparently more malignant lesions.
On the other hand, the DMBA-PO group mostly showed malignant-looking neoplastic formations in the oral mucosa, such as ulcerated nodules, necrosis areas, exophytic and verrucous tumors, and areas with abundant vascularization.
\nIn addition to the oral pouches, both DMBA groups presented tumors in the esophagus and stomach. Maximum and minimum measures of all lesions are shown in \nTable 1\n.
\nHistological lesion | \nLocation | \nNumber of lesions | \nMean diameter (Ø mm) | \n||
---|---|---|---|---|---|
\n | \n | \nDMBA-PA\n | \n\nDMBA-OO\n | \n\nDMBA-PA\n | \n\nDMBA-OO\n | \n
\nSQUAMOUS PAPILLOMA\n | \nOral epithelium | \n1 | \n11 | \n1 | \n0.14 (0.1-0.2) | \n
Esophagus | \n9 | \n19 | \n0.6 (0.2-1.2) | \n0.34 (0.2-1) | \n|
Stomach | \n14 | \n21 | \n0.7 (0.3-1.2) | \n0.5 (0.2-1) | \n|
\nINTRAEPITHELIAL CARCINOMA\n | \nOral epithelium | \n20 | \n2 | \n0.2 (0.1-0.4) | \n0.3 (0.3) | \n
Esophagus | \n0 | \n1 | \n0 | \n0.3 | \n|
Stomach | \n1 | \n0 | \n0.6 | \n0 | \n|
\nVERRUCOUS CARCINOMA\n | \nOral epithelium | \n1 | \n2 | \n1.7 | \n4 (1-7) | \n
Esophagus | \n3 | \n0 | \n0.9 (0.8-1) | \n0 | \n|
Stomach | \n5 | \n2 | \n1.5 (0.8-2) | \n1.5 (1.2-1.8) | \n|
\nINVASIVE CARCINOMA\n | \nOral epithelium | \n1 | \n0 | \n0.5 | \n0 | \n
Esophagus | \n2 | \n0 | \n3 (2-4) | \n0 | \n|
Stomach | \n2 | \n0 | \n5.3 (1.5-9) | \n0 | \n
Number, size and type of tumors in DMBA groups at 20th week.
No visible lesions were found in the control groups which only received paraffin oil or olive oil.
\nThe histological study at 20 weeks evidenced different types of lesions, demonstrating a complete carcinogenesis process in both DMBA groups: Squamous papillomas, intraepithelial carcinomas, verrucous carcinomas and invasive SCC.
\nSQUAMOUS PAPILLOMAS: Papillary projections lined with squamous epithelium were noted, showing hyperkeratosis and epithelial thickening. No atypia or mitotic activity was observed (\nFigure 3\n). Twelve papillomas were found among the groups exposed to DMBA (one in the DMBA-PO group and eleven in the DMBA-OO group). The differences regarding incidence of this kind of lesion were statistically significant (p .004).
\nSquamous papilloma is an exophytic lesion which shows typically papillary growth and highly differentiated epithelium.
INTRAEPITHELIAL CARCINOMAS (\nFigure 4\n): Twenty four intraepithelial carcinomas were identified. Twenty one occurred in the DMBA-PO group, and three in the DMBA-OO group. The differences observed between both groups were statistically significant (p .003).
\nIntraepithelial carcinoma is classically characterized by full-thickness with hyperkeratosis and parakeratosis, hypercellurality, nuclear atypia and mitotic figures. The epithelium-stroma interface is preserved.
VERRUCOUS CARCINOMAS: Several exophytic lesions with papillomatosis and infiltrative growth (\nFigure 5\n). Thirteen verrucous carcinomas were found, nine in the in the DMBA-PO group and four in the DMBA-OO group. This was not statistically significant (p .523).
\nSCC, verrucous carcinoma. Verrucous carcinoma is warty-appearing, highly differentiated, and shows hyperkeratosis. There is minimal atypia, abundant eosinophilic cytoplasm and normal mitotic figures. No invasion of the stroma by isolated neoplastic cells was observed.
INVASIVE CARCINOMAS (\nFigure 6\n): Light microscopy revealed epithelial proliferations that, like cords, invaded the adjacent stroma. In addition, the proliferating cells showed marked atypia and mitotic activity. Five invasive carcinomas were found in the DMBA-PO group.
\nInvasive SCC (A). The SCC is composed of infiltrating islands or nets of malignant cells, which form an irregular growth pattern (B).
The carcinogenic effect in the DMBA-PA group (35 carcinomas) corresponded to 100%, while in the DMBA-OO group (7 carcinomas), it was of 20%. According to the observed results, inhibitory effect seen in the DMBA-OO group was 86% for intraepithelial carcinoma, 56% for verrucous carcinoma, and 100% for SCC (\nTable 2\n).
\nLesion | \n\n | DMBA-PA (%) | \nDMBA-OO (%) | \n
---|---|---|---|
\nINTRAEPITHELIAL CARCINOMA\n | \nCarcinogenic effect | \n21 (100) | \n3 (14.3) | \n
Inhibitory effect | \n\n | (85.7) | \n|
\nVERRUCOUS CARCINOMA\n | \nCarcinogenic effect | \n9 (100) | \n4 (44.4) | \n
Inhibitory effect | \n\n | (55.6) | \n|
\nINVASIVE CARCINOMA\n | \nCarcinogenic effect | \n5 (100) | \n0 | \n
Inhibitory effect | \n\n | (100) | \n|
\nTOTAL CARCINOMAS\n | \nCarcinogenic effect | \n35 (100) | \n7 (20) | \n
Inhibitory effect | \n\n | (80) | \n
Carcinogenic and inhibitory effects of DMBA-PO/DMBA-OO, according to lesion subtypes.
No tumors were observed in the control animals.
\nThis research work about carcinogenesis is based on an experimental model of induced SCC after the administration of DMBA at 0.5% -dissolved in mineral oil- into the oral pouches of the hamster. We think, like Nagini and Kowshik [3], that the DMBA carcinogenesis model in hamster oral pouches is characteristic and highly representative of the “cancer induction”. In addition, it is advantageous for the reproducibility of lesions, facilitates experimental research, and can be used as a test for chemotherapy and preventive agents. Also, in this work, the olive oil inhibitory effect on carcinogenesis has been studied alone -extra virgin olive oil applied before, during, and after DMBA, and ad libitum diet with ripe olives ripe of the Picual variety, from the olive harvest-, and combined -as a solvent for DMBA- [16].
\nThe carcinomas produced in the upper gastrointestinal tract were SCC, similar to SCC of the oral mucosa in humans. These results coincide with those obtained in other experimental works [17, 18].
\nIn oral carcinogenesis, using DMBA in hamsters, some authors have described the development of precancerous lesions and, subsequently, their progression towards intraepithelial carcinoma and invasive carcinoma after a few months. At 8 weeks, precancerous lesions usually appear. At 12 weeks, these evolve to intraepithelial carcinoma; eventually developing into invasive carcinomas at 18 weeks. This phenomenon, although slower, also occurs in humans [19]. The results obtained in our work resemble those of oral cancer progression described in the literature.
\nAs in the field of experimental carcinogenesis, research on cancer prevention has continued to grow in recent decades, focusing on agents proposed for this purpose, although with few results yet. This is the case of the mediterranean diet, which is largely based on extra virgin olive oil, and that has been explored in the prevention of breast cancer [11, 13], and colorrectal cancer [9]. In the present work, the combination of olive oil as dissolvent, extra virgin olive oil applied before, during, and after DMBA application, and ad libitum diet with Picual variety olives, have been used as a preventive agent of DMBA carcinogenesis.
\nMenéndez et al. have shown that extra virgin olive oil polyphenols can inhibit erbB-2 malignant transformation of human breast cancer epithelial cells [14]. Owen et al. pointed out the importance of phenolic compounds isolated from olive oil as antioxidants and their anticancer potential [20].
\nIn this sense, olive oil is composed of 99% different fatty acids, the most important being oleic acid, a monounsaturated fatty acid, with a richness of 60-80%, and other fatty acids -palmitic, stearic, palmitoleic, linoleic, and linolenic-. The remaining 1% is made up of vitamin E and natural antioxidants. The most important antioxidants are phenolic compounds, present in the mesocarp of the olive and in extra virgin olive oil, which are mainly responsible for the antioxidant properties and which are not present in any other vegetable oil. For this reason, the diet added to the standard feed that the hamsters received was ripe olives from the tree, recently harvested and not spoiled. The variety of olive richest in phenolic compounds is the Picual variety.
\nKeys et al. demonstrated an inversely proportional relationship between the incidence of cardiovascular diseases and the adoption of eating habits established in seven countries in the Mediterranean area [21]. It seems that this “cardiovascular protection” resides in the creation of an anti-atherosclerotic plasma profile, which is defined by a decrease in total cholesterol and low-density lipoprotein (LDL) cholesterol levels, as well as by an increase in high-density lipoprotein (HDL) cholesterol. Some studies have attributed these properties to the high content of oleic acid -monounsaturated grade acid of the omega-9 series- of olive oil [22].
\nAnalyzing our results, we can affirm that the combination of olive oil as a solvent for DMBA, extra virgin olive oil applied before, during, and after DMBA administration, and ad libitum diet with Picual olives has shown the capability to reduce the malignant progression of lesions already started, and modify the malignant phenotype of some neoplasms, making it less aggressive.
\nIt is possible that in the DMBA-OO group, -COOH groups and unsaturated bonds of the vegetable oil could absorb or react with carcinogen, decreasing the effective concentration of the carcinogen. The antioxidant effect and anticancer properties of extra virgin olive oil expressed by some authors are reinforced [18, 19].
\nThe study of the lesions at 20 weeks showed a total of 59 neoplasms in the DMBA-PO group and 58 in the DMBA-OO group, so there were no differences in the absolute incidence of tumors. However, clear differences were observed regarding the type of neoplasms and malignancy. Eighty-eight percent of the tumors in the DMBA-OO group corresponded to benign squamous papilloma-type tumors, compared to 41% that developed in the DMBA-PO group; the rest were carcinomas.
\nIn addition, hamsters that did not eat ripe olives and did not receive extra virgin olive oil, developed 21 intraepithelial carcinomas, 9 verrucous carcinomas, and 5 invasive carcinomas; while animals that received the olive oil as a solvent for DMBA, extra virgin olive oil -before, during, and after DMBA-, and ad libitum diet with Picual olive developed 3 intraepithelial carcinomas, 4 verrucous carcinomas, and no invasive squamous carcinoma.
\nThe inhibitory effect of extra virgin olive oil (Picual variety) on the experimental chemical carcinogenesis is higher than 50% for carcinomas, especially for intraepithelial carcinoma and invasive squamous carcinoma.
\nFurthermore, the tumors originated in animals who received DMBA mixed with olive oil were predominantly benign, specifically of the squamous papilloma subtype.
\nTherefore, these data suggest that the extra virgin olive oil and the diet with ripe olives extracted from the harvesting of the tree may modulate the experimental carcinogenesis with DMBA, originating very well differentiated and not very aggressive tumors.
\nThe authors declare no conflict of interest.
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