The Role Of Mitochondrial Permeability Transition Pore In The Rabbit Lens Epithelial Cells Apoptosis Induced By Hydrogen Peroxide

Lens epithelial cells (LECs) are responsible for material and energy metabolism during lens growth, differentiation and repair. It has been well studied that LECs' apoptosis is the cytology basis of all sorts of cataract formation except that of congenital. Hydrogen peroxide is an important factor that can induce cataract formation. There is a common view of point that cell nucleus is the control center of the apoptosis, furthermore, analysis on cell ultrastructure have showed that the mitochondrial structure can maintain to the last stage of cell apoptosis. But recently a plenty of researches have represented that mitochondrial membrane is responsible for the occurrence of all sorts of cells' apoptosis. Mitochondrial plays an importantrole in the process of apoptosis and even death. For these reasons, view diversion on cell apoptosis' control center from nucleus to mitochondrial has arose.The changes of mitochondrial structure and function play a key role in the process of apoptosis. And the fundamental change of mitochondrial during the process of apoptosis is the mitochondrial permeability transition (MPT). Affected by different kinds of factors that could intrigue apoptosis, the mitochondrial permeability transition pore (MPTP) opened which makes the mitochondrial permeability transition changes and subsequently cause the mitochondrial membrane potential ( m) reduce and a sequence of prompting apoptosis materials including apoptosis induced factor(AIF), cytochrome C release. All these changes induce the chromatin condense and DNA rapture and other apoptosis representation. Because the open of MPTP occurred prior to the changes of mitochondrial permeability transition, it is important to clarify the role and function of MPTP for investigating the mechanism of cells' apoptosis. This experiment is designed to investigate the possible role of mitochondrial permeability transition pore (MPTP) in the control of the apoptosis of rabbit lens epithelial cells during oxidation damage and elucidate the detail of oxidation damage on lens epithelial cells.[Objective]To investigate the role of mitochondrial in the lens epithelial cells apoptosis. By blocking the rabbit lens epithelial cells' mitochondrial permeability transition pore, and measuring the effect on the apoptosis process induced by hydrogen peroxide (H2O2), which include the changes of cells' ultrastructure, mitochondrial function and apoptosis ratio, we investigate the role of mitochondrial permeability transition pore (MPTP) during the process of rabbit lens epithelial cells' apoptosis which induced by hydrogen peroxide.[Methods]So far cyclosporine A is regarded as the most effective blocking reagent to mitochondrial permeability transition pore. By means of blocking mitochondrial permeability transition porewith different concentrations of cyclosporine A is regarded as the main method to investigate whether mitochondria! permeability transition pore enroll in the control of apoptosis. For this reason, we pre-treat cultured cells with different concentrations of cyclosporine A to block the mitochondrial permeability transition pore.At the first stage, we culture the rabbit lens epithelial cells and establish the model of oxidation damage by treating cells with 0.1mmol-L"' hydrogen peroxide for 2 hour. Different concentration of cyclosporine A have been employed to pre-treat the lens epithelial cells for 30 minutes which would block the mitochondrial permeability transition pore in different degrees (The solute, DMSO, of all the concentrations of cyclosporine A solutions selected have been measured and guaranteed to have no anti-effect on lens epithelial cells' growth and proliferation). Then the cells were treated with 0.1mmol-L"' hydrogen peroxide for 2 hour. To detect the effect on the cells caused by the block of mitochondrial permeability pore, we use MTT assay to measure the cell mitochondrial function; flow cytometer with AnnexinV and propidium iodide double staining to measure the cells apoptosis percentage, and finally, tr...