姜黄素通过抑制α-突出核蛋白聚集保护鱼藤酮致多巴胺能细胞损伤的机制研究

Mechanism of curcumin's protection of dopaminergic cells from rotenone-induced injury by inhibiting α-synuclein aggregation

目的探讨姜黄素(Curcumin)对α-突触核蛋白(α-synuclein)聚集的影响,及其拮抗鱼藤酮(Ro)诱导的PC12细胞损伤的作用机制。方法选用大鼠嗜铬细胞瘤株PC12细胞,利用鱼藤酮诱导其损伤建立帕金森病细胞模型,利用姜黄素进行干预;采用MTT法检测细胞活力、荧光酶标仪检测蛋白酶体水解酶活性、Western blotting法检测α-突出核蛋白表达、免疫荧光法检测细胞内α-突出核蛋白聚集、流式细胞仪检测PC12细胞凋亡。结果鱼藤酮组PC12细胞活力及蛋白酶体水解酶活性明显降低,α-突出核蛋白表达和聚集以及细胞凋亡率明显增加;经0.5、1.0、5.0、10μmol/L各浓度姜黄素预处理4h后与0.1μmol/L鱼藤酮共同孵育PC12细胞24h,0.5和1.0μmol/L的姜黄素使细胞活力以及蛋白酶体水解酶活性明显升高、α-突出核蛋白的表达和聚集明显减少、细胞凋亡率明显降低;5.0和10μmol/L姜黄素对鱼藤酮的拮抗作用明显减弱,细胞活力和蛋白酶体水解酶活性、以及细胞凋亡率与鱼藤酮组比较均无明显差异。结论低浓度姜黄素能够通过诱导PC12细胞蛋白酶体水解酶活性、抑制α-突出核蛋白的表达和聚集,从而拮抗鱼藤酮诱导的PC12细胞的损伤。

Objective Purpose To investigate the effect of curcumin on aggregtion of alph-synuelein and its mechanism to resist the rotenone-induced injury to PC12 cells. Methods The cellular modee of parkinson＇s disease was established by adoption of rat pheochromocytoma strain PC12 cells and utilization of rotenone was induced injury to them. Cell viability was assessed with M＇fT, enzymatic activity of three hydrolases in proteasome was measured by detection of the fluoro- phore of various cleavaged synthetic lluorogenic peptides. The expression and aggregation of α-synuclein in PC12 cells was observed by Western blot and immunofluorescence respectively and cell apoptosis by flow cytometry. Results Cell viability and activity of proteasome of rotenone group were decreased dramatically. The expression and aggregation of α-synuclein and cell apotosis of Rotenone group increased apparently. 0.5 μmol/L and 1.0μmol/L of curcumin allowed cell viability and activity of proteasome of PC12 cells to increase significantly after co-incubation of curcumin-pretreated PC12 cells with Ro for 24h and the resistant role of 5. 0μmol/L and 10μmol/L of curcumin on the injury of rotenone-induced PC12 cells decreased apparantly. Conclusion Low concentrion of curcumin was able to induce the activity of proteasome and inhibit the expression and aggregation of α-synuclein in PC12 cells and thereby to alleviate the injury of rotenone-induced PC12 cells.