葛根素对MPP^+诱导的SH-SY5Y细胞线粒体途径凋亡的保护作用  

Protective effects of puerarin against 1-methyl-4-phenylpyridinium-induced mitochondrial apoptotic death in differentiated SH-SY5Y cells

作　　者：程月发[1] 朱国旗[2] 关亚丽[1] 刘颖硕[1] 胡艳[1] 李庆林[2] 

机构地区：[1]河北联合大学冀唐学院,河北唐山063000 [2]安徽中医学院安徽省现代中药重点实验室,安徽合肥230038

出　　处：《中国中药杂志》2011年第9期1222-1226,共5页China Journal of Chinese Materia Medica

基　　金：国家自然科学基金面上项目(30973894);安徽省高等学校省级自然科学研究计划重点项目(KJ2008A051)

摘　　要：目的:探讨葛根素对MPP+诱导SH-SY5Y细胞损伤的保护机制。方法:台盼蓝染色检测细胞存活率;PI单染流式细胞术分析细胞凋亡率、细胞周期分布;ELISA和Western blotting法检测胞浆Cyt C的变化;ELISA方法检测Caspase-3,Caspase-8和Caspase-9酶活性;DNA ladder分析药物对细胞凋亡的干预作用。结果:在12.5~50μmol.L-1,葛根素对MPP+诱导的SH-SY5Y细胞损伤具有一定的保护作用,并呈一定的量效关系;PI单染流式细胞术检测结果显示MPP+诱导SH-SY5Y细胞发生凋亡,并阻滞细胞于G2/M期,预先加入不同剂量的葛根素可以逆转MPP+对SH-SY5Y细胞损伤作用。West-ern blotting和ELISA分析结果均显示葛根素能够抑制MPP+诱导SH-SY5Y细胞的Cyt C的释放。不同剂量的葛根素也能够抑制MPP+诱导SH-SY5Y细胞的Caspase-3和Caspase-9的活性,但对Caspase-8的活性影响不明显;DNAladder分析也显示葛根素可以抑制MPP+诱导SH-SY5Y细胞凋亡。结论:葛根素可以提高MPP+诱导的SH-SY5Y损伤的细胞存活率,抑制细胞凋亡的发生,其机制可能与调节线粒体Caspase通路和干预细胞周期有关。It is well known that puerarin possesses protective activity on neurodegenerative diseases.However,the exact pathway involved in the protective effect of puerarin on MPP＋-induced cell death is unclear.In this study,we focused on mitochondria impairment in the apoptotic process of MPP＋-elicited SH-SY5Y cells and detected the protection of puerarin.As evidenced by Trypan blue assay,the cell viability was significantly decreased by 1 mmol·L-1 MPP＋,but reversed by different concentrations puerarin pretreatment.Flow cytometer analysis revealed that MPP＋-induced SH-SY5Y cells apoptosis and arrested the cells in G2/M phase,whereas puerarin pretreatment concentration dependently reversed the apoptosis ratio.In addition to the apoptosis ratio,50.0 μmol·L-1 puerarin pretreatment even altered the MPP＋-induced G2/M phase arrest.JC-1 assay suggested that MPP＋ significantly opened MMP of the SH-SY5Y cells;pretreatment with puerarin attenuated the deterioration of the MMP.Both ELISA and Western blotting showed that puerarin prevented the release of cytochrome c from the mitochondrial interior to the cystol elicited by MPP＋.DNA ladder showed that typical DNA ladder was present in the MPP＋-induced SH-SY5Y cells.Additionally,MPP＋ enhanced caspase-9 and caspase-3 activity,respectively,while not caspase-8.However,the enhancement was concentration dependently blocked by puerarin pretreatment.Taken together,puerarin can modulate mitochondrial membrane potential and inhibit the cytochrome c releasing-caspase cascade to prevent MPP＋-induced cell injury.

关 键 词：帕金森病 葛根素 线粒体 细胞凋亡 

分 类 号：R285.5[医药卫生—中药学]