The purpose of this study was to investigate whether rabbit bone marrow-derived mesenchymal stem cells (MSCs) effectively decrease alkali-induced oxidative stress in the rabbit cornea. The alkali (0.15 N NaOH) was applied on the corneas of the right eyes and then rinsed with tap water. In the first group of rabbits the injured corneas remained untreated. In the second group MSCs were applied on the injured corneal surface immediately after the injury and eyelids sutured for two days. Then the sutures were removed. In the third group nanofiber scaffolds seeded with MSCs (and in the fourth group nanofibers alone) were transferred onto the corneas immediately after the injury and the eyelids sutured. Two days later the eyelid sutures were removed together with the nanofiber scaffolds. The rabbits were sacrificed on days four, ten or fifteen after the injury, and the corneas were examined immunohistochemically, morphologically, for the central corneal thickness (taken as an index of corneal hydration) using an ultrasonic pachymeter and by real-time PCR. Results show that in untreated injured corneas the expression of malondialdehyde (MDA) and nitrotyrosine (NT) (important markers of lipid peroxidation and oxidative stress) appeared in the epithelium. The antioxidant aldehyde dehydrogenase 3A1 (ALDH3A1) decreased in the corneal epithelium, particularly in superficial parts, where apoptotic cell death (detected by active caspase-3) was high. (In control corneal epithelium MDA and NT are absent and ALDH3A1 highly present in all layers of the epithelium. Cell apoptosis are sporadic). In injured untreated cornea further corneal disturbances developed: The expressions of matrix metalloproteinase 9 (MMP9) and proinflammatory cytokines, were high. At the end of experiment (on day 15) the injured untreated corneas were vascularized and numerous inflammatory cells were present in the corneal stroma. Vascular endothelial growth factor (VEGF) expression and number of macrophages were high. The results obtained in injured corneas covered with nanofiber scaffolds alone (without MSCs) or in injured corneas treated with MSCs only (transferred without scaffolds) did not significantly differ from the results found in untreated injured corneas. In contrast, in the injured corneas treated with MSCs on nanofiber scaffolds, ALDH3A1 expression remained high in the epithelium (as in the control cornea) and positive expression of the other immunohistochemical markers employed was very low (MMP9) or absent (NT, MDA, proinflammatory cytokines), also similarly as in the control cornea. Corneal neovascularization and the infiltration of the corneal stroma with inflammatory cells were significantly suppressed in the injured corneas treated with MSCs compared to the untreated injured ones. The increased central corneal thickness together with corneal opalescency appearing after alkali injury returned to normal levels over the course of ten days only in the injured corneas treated with MSCs on nanofiber scaffolds. The expression of genes for the proinflammatory cytokines corresponded with their immunohistochemical expression. In conclusion, MSCs on nanofiber scaffolds protected the formation of toxic peroxynitrite (detected by NT residues), lowered apoptotic cell death and decreased matrix metalloproteinase and pro-inflammatory cytokine production. This resulted in reduced corneal inflammation as well as neovascularization and significantly accelerated corneal healing.

• MeSH
• alkálie toxicita   MeSH
• chemické popálení patologie   chirurgie   MeSH
• hojení ran MeSH
• králíci MeSH
• mezenchymální kmenové buňky cytologie   MeSH
• modely nemocí na zvířatech MeSH
• nanovlákna terapeutické užití   MeSH
• oxidační stres * MeSH
• poranění rohovky MeSH
• rohovka patologie   chirurgie   MeSH
• tkáňové podpůrné struktury * MeSH
• transplantace mezenchymálních kmenových buněk metody   MeSH
• zvířata MeSH
• Check Tag
• králíci MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH

Úvod: Cílem naší studie je zanalyzovat příčiny vedoucí k explantaci nitroočních čoček. Metodika: Soubor retrospektivní studie zahrnuje celkem 22 očí 21 pacientů, kteří v letech 2008–2011 podstoupili na Evropské oční klinice Lexum explantaci nitrooční čočky. Soubor tvoří dvě skupiny očí. Do první skupiny A jsou zahrnuty oči s explantovanou monofokální čočkou (14 očí, 13 pacientů), druhou skupinu B tvoří oči s explantovanou multifokální čočkou (8 očí, 7 pacientů). Analýza příčin vedoucích k explantaci nitroočních čoček byla provedena jednak na základě subjektivních výpovědí pacientů a jednak na základě objektivního nálezu ve zdravotnické dokumentaci. Výsledky: Mezi nejčastější příčiny explantace monofokálních čoček se řadí refrakční chyba, luxace čočky, decentrace čočky a zkalení nitrooční čočky. Ve skupině multifokálních čoček byly nejčastější příčinou vedoucí k explantaci čočky fotické fenomény (halo, glare), dále nerealistické očekávání pacienta a refrakční chyba. Závěr: Zatímco ve skupině monofokálních čoček dominovala jako příčina jejich explantace refrakční chyba, ve skupině multifokálních čoček zaujímaly přední místo rušivé fotické fenomény. Správný výběr pacienta, kvalitně provedená operace, optimálně zvolená dioptrická síla nitrooční čočky na základě precizní biometrie stejně jako typ a materiál čočky patří mezi nejdůležitější faktory, které mohou minimalizovat riziko komplikací vedoucích k explantaci nitrooční čočky.

Aim: To analyze the reasons of intraocular lenses (IOL) explantation. Methods: Retrospective study of 22 eyes of 21 patients. Those patients underwent explantation of intraocular lenses between the years 2008 to 2011. The study group included two eye groups. Group A included eyes with exptanted monofocal IOL (14 eyes of 13 patients) and group B included eyes with explanted multifocal IOL (8 eyes of 7 patients). Reasons requiring explantation were analyzed based on subjective patient complaints and on objective ocular findings in health documentation as well. Results: The most common indications of explantation in Group A of monofocal lenses were incorrect IOL power, followed by IOL luxation, decentration and IOL opacity. In Group B of multifocal IOL, the most common reasons for IOL removing were halo and glare, inadequate postoperative expectations and incorrect IOL power. Conclusion: The main reason for IOL exchange in Group A of monofocal lenses was the incorrect power, while in Group B of multifocal IOL the main reason were the disturbing photic phenomenon like glare and halo. Careful patient selection, good surgical technique, optimally selected IOL power measurements based on precise biometry readings as well high quality of IOL materials are the most important factors in minimizing the risk for IOL explantation.

• MeSH
• implantace nitrooční čočky * MeSH
• lidé MeSH
• nitrooční čočky * škodlivé účinky   MeSH
• oftalmologické chirurgické výkony MeSH
• statistika jako téma MeSH
• výsledek terapie MeSH
• výsledky a postupy - zhodnocení (zdravotní péče) * MeSH
• Check Tag
• lidé MeSH

Limbal transplantation or limbal stem cell (LSC) transfer represents the only way to treat severe ocular surface damage or LSC deficiency. However, limbal allografts are promptly rejected in spite of extensive immunosuppressive therapy. To characterize immune response after limbal transplantation, we established an experimental model of limbal transplantation in the mouse. Syngeneic, allogeneic and xenogeneic (rat) limbal grafts were grafted orthotopically in BALB/c mice and graft survival was evaluated. The presence of graft donor cells and the expression of IL-2, IL-4, IL-10, IFN-γ and inducible nitric oxide synthase (iNOS) mRNA in the grafts were detected by real-time PCR. While syngeneic grafts survived permanently, allografts were rejected in 9.0±1.8 days and xenografts in 6.5±1.1 days. The manifestation of clinical symptoms of rejection correlated with the disappearance of donor cells in the graft and in the recipient cornea. Intragraft expression of iNOS mRNA and distinct expression patterns of Th1 (IL-2, IFN-γ) and Th2 (IL-4, IL-10) cytokines were detected during rejection of limbal allografts and xenografts. The limbal graft rejection was prevented with anti-CD4, but not anti-CD8 monoclonal antibody therapy. The results indicate that limbal grafts do not enjoy immune privilege of the eye and are promptly rejected by Th1 (allografts) or by a combined Th1 and Th2 (xenografts) type of immune response involving CD4+ cells and iNOS expression. Targeting this pathway may be an effective way to prevent and treat limbal graft rejection.

• MeSH
• antigeny CD4 imunologie   MeSH
• antigeny CD8 imunologie   MeSH
• homologní transplantace MeSH
• interferon gama biosyntéza   imunologie   metabolismus   MeSH
• interleukiny biosyntéza   imunologie   metabolismus   MeSH
• krysa rodu rattus MeSH
• modely u zvířat MeSH
• monoklonální protilátky imunologie   farmakologie   terapeutické užití   MeSH
• myši inbrední BALB C MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• oči cytologie   imunologie   MeSH
• potkani inbrední LEW MeSH
• přežívání štěpu účinky léků   imunologie   MeSH
• rejekce štěpu imunologie   metabolismus   prevence a kontrola   MeSH
• synthasa oxidu dusnatého, typ II imunologie   MeSH
• Th1 buňky imunologie   sekrece   MeSH
• Th2 buňky imunologie   sekrece   MeSH
• transplantace heterologní MeSH
• transplantace kmenových buněk MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Stem cell (SC) therapy represents a promising approach to treat a wide variety of injuries, inherited diseases, or acquired SC deficiencies. One of the major problems associated with SC therapy remains the absence of a suitable matrix for SC growth and transfer. We describe here the growth and metabolic characteristics of mouse limbal stem cells (LSCs) and mesenchymal stem cells (MSCs) growing on 3D nanofiber scaffolds fabricated from polyamide 6/12 (PA6/12). The nanofibers were prepared by the original needleless electrospun Nanospider technology, which enables to create nanofibers of defined diameter, porosity, and a basis weight. Copolymer PA6/12 was selected on the basis of the stability of its nanofibers in aqueous solutions, its biocompatibility, and its superior properties as a matrix for the growth of LSCs, MSCs, and corneal epithelial and endothelial cell lines. The morphology, growth properties, and viability of cells grown on PA6/12 nanofibers were comparable with those grown on plastic. LSCs labeled with the fluorescent dye PKH26 and grown on PA6/12 nanofibers were transferred onto the damaged ocular surface, where their seeding and survival were monitored. Cotransfer of LSCs with MSCs, which have immunosuppressive properties, significantly inhibited local inflammatory reactions and supported the healing process. The results thus show that nanofibers prepared from copolymer PA6/12 represent a convenient scaffold for growth of LSCs and MSCs and transfer to treat SC deficiencies and various ocular surface injuries.

• MeSH
• buněčná diferenciace MeSH
• kaprolaktam analogy a deriváty   chemie   MeSH
• kmenové buňky cytologie   MeSH
• kultivované buňky MeSH
• limbus corneae cytologie   MeSH
• mezenchymální kmenové buňky cytologie   MeSH
• myši MeSH
• nanovlákna MeSH
• polymery chemie   MeSH
• poranění oka terapie   MeSH
• proliferace buněk MeSH
• rohovkový epitel cytologie   MeSH
• tkáňové inženýrství MeSH
• tkáňové podpůrné struktury MeSH
• transplantace kmenových buněk MeSH
• transplantace mezenchymálních kmenových buněk MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

PURPOSE: To detect and isolate cells with stem cell (SC) characteristics in the limbus of the mouse. METHODS: Limbal tissues from BALB/c mice were trypsin-dissociated and separated on the gradient Percoll (Fluka, Buchs, Switzerland). Several fractions were isolated and characterized by real-time PCR for the presence of limbal SC markers and differentiation markers of corneal epithelial cells by flow cytometry for the determination of the side-population (SP) phenotype and growth properties in vitro. RESULTS: Cells retained in the lightest fraction (40% Percoll) and in the densest fraction (80% Percoll) of the gradient were both enriched for populations with a high expression of the SC markers ABCG2 and Lgr5 and also expressing the SP phenotype. However, the lightest fraction (representing approximately 12% of total limbal cells) contained cells with the strongest spontaneous proliferative capacity and expressed the corneal epithelial differentiation marker K12. In contrast the densest fraction (<7% of original cells) was K12 negative and contained small nonspontaneously proliferating cells, which instead were positive for p63. Unexpectedly, cells from this fraction had the highest proliferative activity when cultured on a 3T3 feeder cell monolayer. CONCLUSIONS: These findings demonstrate the presence of two distinct populations of corneal epithelial cells with limbal SC characteristics, based on differential expression of the keratin-specific marker K12 and transcription factor p63, and suggest a difference in developmental stage of the two populations, with the K12(-)p63(+) population being closer to the primitive limbal SC

• MeSH
• ABC transportéry analýza   genetika   MeSH
• buněčné dělení MeSH
• buňky 3T3 MeSH
• centrifugace - gradient hustoty metody   MeSH
• fibroblasty cytologie   MeSH
• financování organizované MeSH
• kmenové buňky cytologie   fyziologie   MeSH
• myši inbrední BALB C MeSH
• myši MeSH
• oxid křemičitý MeSH
• polymerázová řetězová reakce MeSH
• povidon MeSH
• průtoková cytometrie MeSH
• rohovkový epitel cytologie   MeSH
• separace buněk metody   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH