Empirical Study On The Effects Of Vitrectomy And The Intervene Of Tea Polyphenols Ophthalmic Gel On The Lens Epithelial Cells

Objective：To observe ROS expression in mitochondria，membrane potential，Caspases-3and endoplasmic reticulum of rabbit lens epithelial cells cultured aftervitrectomy and the changes of them after the intervention of tea polyphenols ophthalmicgel.To study the causes and effects of these changes，and to further explore the complicatedcataract and tea polyphenols ophthalmic gel，s curative effect.Methods：1. Rabbit lens epithelial cells were cultured in vitro in different specialselective medium made from BSS，silicone oil，O₂，C₃F₈concentration to simulate thevitrectomy in vivo.The morphous changes of the cells were then observed when they wereintervened in different concentration of tea polyphenols ophthalmic gel.2. Prepare mediumwith final concentrations of1000μg/ml、100μg/ml、10μg/ml、1μg/ml、0μg/ml teapolyphenols ophthalmic gel to clture rabbit lens epithelial cells in vitro，and use CCK-8tochoose the safe tea polyphenols ophthalmic gel concenrations.3.The rabbit lens epithelialcells were divided into4groups：A group were normal cells-control group；B group wereRLECs burned by BSS，silicone oil，O₂，C₃F₈injury group；C group were RLECs culturedwith medium mixed with final concentrations of10μg/ml and induced damage by BSS，silicone oil，O₂，C₃F₈；D group were RLECs cultured with medium mixed with finalconcentrations of100μg/ml and induced damage by BSS，silicone oil，O₂，C₃F₈.4. Usemitochondrial membrane potential detection kit and mitochondrial reactive oxygen speciesassay kit to detecte by flow cytometry of mitochondrial reactive oxygen species，mitochondrial membrane potential，The expression of endocytoplasmic reticulum and Caspases-3in rabbit lens epithelial cells was detected with immunocytochemistry andimmunofluorescence to analyze the effect of tea polyphenols ophthalmic gel on rabbit lensepithelial cells exposed to BSS，silicone oil，O₂，C₃F₈damage and the possible mechanism.Results：1. Thougth inverted phase contrast microscope：rabbit lens epithelial cellscultured in vitro were spindle or polygonal，adherent monolayer growth，and cell borderswere clearly.2. Thougth test of CCK-8：（1） There were statistically significant differencesabout the OD values between cells with medium mixed with final concentration of1000μg/ml tea polyphenols ophthalmic gel and nomal retinal pigment epithelial cellgroup（P<0.05）；（2） The OD values of cells with medium mixed with final concentrations of100μg/ml，10μg/ml and1μg/ml tea polyphenols ophthalmic gel were no significantdifferences with nomal retinal pigment epithelial cell group（P>0.05）.3. Observe cells ofgroup A^D by inverted phase contrast microscope after establishing the BSS，silicone oil，O₂，C₃F₈damage model：Cells of group A were spindle or polygonal，adherent monolayergrowth，and cell borders were clearly. Some of cells of group B were shrinkage，rounding，refraction changes，and around the cells coule be seen bright ring；nucleus shrink，split intomultiple or a nuclear margination；there were cell debris suspension cells in the culturemedium. Cells of group C^D were spindle or polygonal，adherent monolayer growth，andcell borders were clearly.few of cells were shrinkage，rounding，refraction changes，therewere no cell debris suspension cells in the culture medium.4. In BSS，silicone oil，O₂，C₃F₈，ROS generated from the lens epithelial cells increased.Inversely，it decreased ingroups interfered with tea polyphenols ophthalmic gel.5. In BSS，silicone oil，O₂，C₃F₈，MMP decreased.It showed reverse outcomes in groups interfered with tea polyphenolsophthalmic gel.6. The positive expression of endocytoplasmic reticulum in rabbit lensepithelial cells decreased in BSS，silicone oil，O₂，C₃F₈. On the contrary，the expressionincreased in groups interfered with tea polyphenols ophthalmic gel.7. The positiveexpression of Caspases-3in rabbit lens epithelial cells increased in BSS，silicone oil，O₂，C₃F₈.On the contrary，the expression decreased in groups interfered with tea polyphenolsophthalmic gel. Conclusions：Vitrectomy can be triggered by various means lens epithelial cellsdeath，the death of epithelial cells may be diabetic cataract in one of the reasons. It isclosely related to the expression changes of endocytoplasmic reticulum and Caspases-3protein which is the most important enzyme in the final implementation in the apoptoticprocess. And tea polyphenols ophthalmic gel may inhibit the formation of mitochondrialreactive oxygen species can be protected from damage to lens epithelial cells andmaintaining its normal structure and function，which may delay the occurrence of diabeticcataract，further study of the disease and put forward the preventive therapeutic drug.