| Oxidative damage is the first risk factor of age-related cataract. Lens epithelial cells (LECs) are responsible for material and energic metabolism during lens growth, differentiation and repair. It is well studied that calcium concentration elevates in LECs of many kinds of cataractous patients. The family of calpains is one kind of neutral restrictive proteolytic enzyme which distributes diffusely. Under pathological condition, abnormal elevated calcium can activate calpains and induce diseases. Calpain II is the predominant form in lenses. PD150606 is a specific inhibitor of calpains with high permeability and high combinative ability. It is known that PD150606 can protect neural system from proteolyticenzyme.Our data showed the changes of opacity of lens, proportion of protein, hydration oflens, apoptosis of LECs, concentration of intracellular calcium, expression and proteolytic activity of calpains induced by H2O2 and inhibition of oxidative cataract by PD 150606, tostudy the possibility of using animal H2OHnduced cataract as the model of humanage-related cataract, the role of calpains in the mechanism of oxidative cataract and inhibition of oxidative cataract by PD 150606.1. Establishment of the model of oxidative cataractPurpose: To study the possibility of using animal H2(>2-induced cataract as the model of human age-related cataract by determination of lens opacity, proportion and electrophoresis bands of water-soluble protein (WSP) and LECs apoptosis of rat HjOr induced cataract.Methods: Rat lenses were cultured in vitro and cataract was induced by H2O2. The lenses were observed under microscope. Simultaneously, photograph and picture analysis were done in order to detect the variation of opacity; the proportion of WSP was determined by the method of Lowry; WSP was detected by SDS-PAGE electrophoresis stained with Coomassie Blue; the hydration of lens was determined; the number of apoptotic LECs was measured by flow cytometry.Results: Opacity of lens was enhanced in 24h when cultured with H2O2. There were significant differences of relative gray scale between H2O2-induced and control group at 6h, 12h and 24h (P=0.032, 0.000,0.000), significant differences of the proportion of WSP between H2O2-induced and control group at 6h, 12h and 24h (P=0.011,0.001,0.000) with lighten bands at 25kDa, 29kDa and 30kDa of WSP, significant differences of hydration of lens at 6h, 12h and 24h (P=0.004,0.001, 0.000) and significant differences of the number of apoptotic LECs at 6h, 12h and 24h (P=0.000, 0.000, 0.000).Conclusions: H2O2 can induce cataract in vitro. The changes of lens morphology, protein and cells were similar with human age-related cataract. It is a recommendable way to use H2O2-induced animal cataract as the model of human age-related cataract.2. Variation of calcium concentration in LECs of oxidative cataractPurpose: To study the role of calcium in the mechanism of oxidative cataract by determination of calcium concentration of rat LECs in H2O2-induced cataract.Methods: Rat lenses were cultured in vitro and cataract was induced by H2O2. Intracellular Ca2+was measured by fluoresence determination with Fura-2/AM.Results: There were significant differences of the concentration of intracellular calcium between H2C>2-induced and control group at 6h, 12h and 24h (P=0.001,0.000, 0.000).Conclusions: The concentration of calcium is elevated in LECs of rat H2O2-induced cataract and may contributes to activation of calcium-dependent proteolytic enzymes such as calpains.3. Protein expression and proteolytic activity of calpains in LECs of oxidative cataractPurpose: To study the role of calpains in the machanism of oxidative cataract by detecting the expression and proteolytic activity of calpains in LECs of H2O2-induced cataract.Methods: Rat lenses were cultured in vitro and cataract was induced by H2O2. The expression of calpain II in LECs was detected with immunohistochemical method; the proteolytic activity in LECs was measured using a fluorogenic...
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