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.
出处
《中国中药杂志》
CAS
CSCD
北大核心
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)
关键词
帕金森病
葛根素
线粒体
细胞凋亡
Parkinson’s disease； puerarin； mitochondria； apoptosis；
