补肾方含药血清拮抗谷氨酸兴奋性神经毒作用及其机制的初步研究  

Preliminary studies on effect and mechanism of serum with Bu Shen Decoction against excitatory neurotoxicity induced by glutamate

作　　者：贺文彬[1] 张俊龙[1] 陈乃宏[2] 张岭[2] 朱海波[2] 

机构地区：[1]山西中医学院,太原030024 [2]中国医学科学院药物研究所,北京100050

出　　处：《中国实验方剂学杂志》2005年第4期28-32,共5页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家863计划资助项目(2004AA2Z3815);山西省自然科学基金资助项目(No:19991091)

摘　　要：目的:研究补肾复方含药血清(BS)对谷氨酸诱导凋亡PC12细胞的保护作用;并观察BS对谷氨酸诱导凋亡的PC12细胞胞浆内钙信号转导机制的影响。方法:采用MTT法测定谷氨酸致损伤后PC12细胞的存活力;采用荧光显微镜技术观察BS对谷氨酸所诱导PC12细胞凋亡的影响;以流式细胞术观察BS对谷氨酸诱导凋亡的PC12细胞胞浆内游离钙离子浓度的影响;运用蛋白免疫印迹(Westernblotting)技术观察BS对凋亡的PC12细胞CaMKⅡ的影响。结果:20%浓度BS含药血清可以拮抗谷氨酸的兴奋性神经毒作用,抑制PC12细胞凋亡;20%浓度BS含药血清能够拮抗谷氨酸诱导凋亡的PC12细胞胞浆内钙离子浓度的升高,抑制CaMKⅡ的磷酸化水平增高。结论:BS含药血清具有拮抗谷氨酸兴奋性神经毒的作用,其神经保护作用的机制可能与减轻钙超载,调控钙信号通路关键信号分子的磷酸化有关。Objectives:The antagonistic effects of serum with Bu Shen Decoction (BS) on apoptosis of PC12 cells induced by glutamate (Glu) were investigated in vitro. So were the effects of serum with BS on the calcium signal transduction system.Methods:The antagonistic action of the BS on neurotoxicity induced by Glu was observed with MTT method.The fluorescence microscope technique was employed to observe the effect of BS on apoptosis of PC12 cells induced by Glu. Its intracellular free calcium concentration was determined by flow cytometry.The Western-blotting technique was applied for observing the effect of BS on phosphorylation of CaMK Ⅱ in apoptotic PC12 cells.Results:The serum with BS in 20% concentration was discovered to be able to exert antagonistic effect on Glu-induced excitatory neurotoxicity and to inhibit the apoptosis of PC12 cells induced by Glu. Moreover,it was discovered that the increase in intracellular free calcium concentration and the excessive phosphorylation of CaMK Ⅱ during apoptosis of PC12 cells induced by Glu could be inhibited by serum with BS in 20% concentration.Conclusion:BS exert an antagonistic action on excitatory neurotoxicity induced by Glu.The mechanism of neuroprotective effect afforded by serum with BS might be related with relieving calcium overload and adjusting phosphorylation of the key signal molecules in the downstream of calcium ion signal transduction system.

关 键 词：补肾复方 细胞凋亡 细胞内钙 钙/钙调素依赖性蛋白激酶Ⅱ 中药血清药理学 

分 类 号：R285.5[医药卫生—中药学]