A Study Of Cydonia Oblanga Mill.Extracts On Human Umbilical Vein Endothelial Cell Injury Induced By Hydrogen Peroxide

Objectives: To study the oxidative injury model of human umbilical vein endothelial cell（HUVEC） was established by treatment with H₂O₂, to investigate the protective effect of Cydonia oblanga Mill.extracts on the injury of HUVEC induced by hydrogen peroxide（H₂O₂） in vitro and to explore its mechanism. Methods: The HUVEC was subcultured in vitro and used for experiment. By the enzyme digestion method to digestion to extend HUVEC, the oxidative injury model of HUVEC was established by treatment with H₂O₂. Purified Cydonia oblonga Mill polysaccharide by sevage method and determination the content by Anthronesulfuric acid method the oxidative injured HUVEC were cultured with different concentrations of Cydonia oblanga Mill. extracts.They were divided into eight groups as follows: Control group, H₂O₂-injured group（10μmo L/L）, low-dosage of TFCOM group（10μg/m L）, mid-dosage of TFCOM group（20μg/m L）, high-dosage of TFCOM group（40μg/m L）, low-dosage of PCOM group（15μg/m L）, mid-dosage of PCOM group（30μg/m L） and high-dosage of PCOM group（60μg/m L）, in which the cells were injured by H₂O₂ for 4 hours.The supemates and the cells were collected respectively after different treatments for measuring the level of ROS,LDH and MDA, as well as SOD with the corresponding detection kit according to the manufacturers instructions.The gene expression of Bax, Bcl-2 and caspase-3 were analysized by Real-time PCR. The protein expression of Bax, Bcl-2 and caspase-3 were analysized by western blotting to further investigation of the mechanism of Cydonia oblanga Mill.extracts in inhibiting HUVEC apoptosis induced by H₂O₂. Results: The apoptosis model of HUVEC with cell volume reduction, cytoplasm shrinkage,condensation and fragmentation of chromatin can be induced by 10μmo L/L H₂O₂. It offered a good instrument for further study the protective effect and mechanism ofCydonia oblanga Mill.extracts on H₂O₂ induced HUVEC injury.Compared with the normal group, H₂O₂ can obviously increase the release of ROS, decrease cell viability,induce cell apoptosis, increase contents of LDH and MDA, decrease the activities of SOD of the cultured medium, reduce the percentage of cells of S phase and G2/M phase.Compared with the H₂O₂ model group, different concentrations of Cydonia oblanga Mill.extracts were shown to alike distinctly attenuate these responses elicited by H₂O₂（P<0.05）. Bcl-2 m RNA level was increased, Bax and caspase-3 m RNA level was decreased by Cydonia oblanga Mill. extracts. Its more consistent with the results of western blot analysis of protein content, and data was statistically significant（P<0.01）.Conclusion: Cydonia oblanga Mill. extracts protective effects against H₂O₂ induced injury in HUVEC, the Cydonia oblanga Mill. extracts can decrease the apoptosis and ease the injury of HUVEC injured by H₂O₂.Key wordsECOMHUVECHOOxidative stress injury...