摘要
目的探讨PDE4抑制剂咯利普兰(rolipram)对H2O2致PC12细胞氧化应激损伤的保护作用及其作用机制。方法以H2O2损伤PC12细胞为氧化应激损伤模型,MTT法检测细胞存活率;碘化丙啶(Propidium iodide,PI)染色流式细胞术检测细胞周期变化;化学比色法测定细胞清除羟自由基(·OH)和超氧阴离子自由基(O2·)的能力;以及培养上清液中的丙二醛(MDA)、谷胱甘肽(GSH)和一氧化氮(NO)含量、超氧化物歧化酶(SOD)活性。Western blot和Real-time RT-PCR检测细胞内硫氧还蛋白(Trx)和诱导型一氧化氮合酶(iNOS)的表达。结果咯利普兰能明显提高H2O2损伤的PC12细胞存活率,恢复细胞增殖;提高细胞清除·OH和O2·的能力;降低MDA和NO含量,提高GSH含量和SOD活性;使Trx蛋白和mRNA表达增加,同时下调iNOS蛋白和mRNA表达。结论咯利普兰对H2O2致PC12细胞损伤具有保护作用,其作用机制可能与提高PC12细胞的抗氧化能力相关。
Aim To investigate the potential protective effects of PDE4 inhibitor rolipram on PC12 cells injury induced by H2O2 and the mechanisms involved. Methods The survival rate of PC12 cells subject to the oxidative damage induced by H2O2 was determined by MTF assay. The PC12 cells were stained with PI and the percentage of different phases in the cell cycle was analyzed by flow cytometry. The ability of clearing ·OH and O2 the content of MDA, GSH and NO and the activity of SOD were determined by colorimetric Assay Kit. Effects of rolipram on Trx and iNOS expression levels of damaged PC12 cells induced by H2O2 were detected both by Western blot and real-time RT-PCR. Results Rolipram significantly increased the survival rate of PC12 cells injured by H2O2 in a concentration dependent manner. And rolipram rescued the mitosis of damaged PC12 cell and stimulated the synthesis of DNA in S-phase of cell cycle, which were shown in rising level of cell ratio in different phases. The biochemical assay data showed rolipram improved the ability of clearing hydroxy radical and superoxide anion in impaired cells induced by oxidative stress with H2O2. Meanwhile, inhibition of PDE4 increased the level of GSH and SOD in cell supernatant, while it decreased the content of MDA and NO. Furthermore, the impaired cells incubated with rolipram expressed higher level of Trx and lower level of iNOS compared with those treated with the vehicle. Conclusion PDFA inhibitor rolipram protects the PC12 cell from the oxidative and antioxidative system.
出处
《中国药理学通报》
CAS
CSCD
北大核心
2010年第11期1429-1435,共7页
Chinese Pharmacological Bulletin
基金
国家自然科学基金资助项目(No30672453
30973518)
广东省自然科学基金重点项目(No7117782)
