| Objective:Lens is in hypoxia in development, which may induce the expression of VEGF in lens epithelial cells. The expression of VEGF may be related not only to the formation of hyaloid vessels, but also to the growth and differentiation of the lens itself through autocrine or paracrine. There have been a lot of studies that indicated the effect of VEGF on the differentiation and survival of multiple sources of stem cells. There are stem/progenitor cell like cells in the germinal zone of lens. The effects of VEGF on growth and differentiation of stem cells in lens aren’t elucidated. Because the expression of VEGF in lens will decline in hyperoxia, newborn mice were fed in hyperoxia to observe the expression of VEGF and the stem cell markers, to study the effect of VEGF on the differentiation and development of lens epithelial cells, and to investigate the potential mechanism and preventive treatment for cataract and posterior capsule opacification.Methods:1, the newborn mice were fed since postnatal (P)0.5in75%high oxygen partial pressure, and the enucleation was manipulated at P3.5and P8.5. The changes of morphology, histology, and ultrastructure of lens epithelial cell were observed.2, the effect of hyperoxia on VEGF and nestin in lens epithelial cells was investigated to analyze characteristics of stem cells and its relationship with VEGF at the above time points with fluorescence quantitative PCR and immunohistochemistry.3, to observe the effect of hyperoxia on proliferation of rat lens epithelial cells. PCNA mRNA transcription was investigated at the above time with fluorescence quantitative PCR. The immunohistochemistry was done to observe the changes of PCNA, BrdU, to analyze the proliferation of lens epithelial cells. Moreover the apoptosis of epithelial cells was observed with DAPI staining.Results:1, Development of mice’s lens in vivo could be influenced in hyperoxia. At P3.5, morphology of lens had no significant difference between hyperoxia group and control group. At P8.5, there was no significant difference in the lens of the horizontal diameter, but the lens was thinner and turbider in hyperoxia group. There was no significant difference in histology of lens at P3.5between groups. However at P8.5, the epithelial cells in anterior subcapsular were, the nuclei were ellipse, regular arranged, but the nuclei of monolayer epithelial cells were round or irregular in hyperoxia group. The epithelial cells were mainly monolayer, occasionally double-deck (possible germinal zone) in control group at equatorial region, with gradual transition of nuclei from round at anterior to spindle backward. More double-deck cells were existence in hyperoxia group at equatorial regions, with the same thansition of nuclei as the contral group. Transmission electron microscopy showed that the anterior capsule deletion in hyperoxia group at P8.5.2, the expression of VEGF in lens epithelial cells increased, but the expression of Nestin decreased. At P3.5and P8.5, the transcription of VEGF mRNA in hyperoxia group significantly decreased than control group than the control, and immunohistochemistry showed the expression of VEGF was weakened iin hyperoxia group at P3.5. PCR and immunohistochemistry showed that the expression of Nestin did not differ significantly between groups at P3.5, while at P8.5, it increased in hyperoxia group than that in the control group.3, hyperoxia induced more proliferation of lens epithelial cell. The expression of PCNA increased in hyperoxia group than that in contral group at P3.5and P8.5through PCR and immunohistochemistry. At the same time, P3.5and P8.5, BrdU staining in epithelial cells were more significantly obvious in hyperoxia group than that incontrol group. DAPI nuclear staining showed that apoptosis could be observed in both groups at P3.5and P8.5.Conclusion:Hyperoxia maybe reduce the synthesis of crystallin and the volume of lens. The vanishment of lens capsule membrane may be related to the abnormal synthesis of collagen, or special collagen hydrolase. The expression of VEGF in lens epithelial cells reduced, while more stem cells are existence and cell proliferation is enhanced, which is contrary to the previous reports that showed that VEGF can promote the proliferation of human lens epithelial cell in culture, which may be concerned with different experimental conditions, or hyperoxia can induce other ways of promoting the proliferation of lens epithelial cells in vivo. The possible mechanism of the differentiation and maturation of lens epithelial cell was discussed, which provides preliminary theoretical basis for further study of the pathogenesis of cataract, and certain theoretical basis for the prevention of cataract and posterior capsule opacific ation. |
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