CAJ | 학술논문

目的探讨参麦注射液对大鼠皮层神经元内质网应激诱导凋亡的保护作用.方法体外培养SD乳鼠皮层神经元细胞,免疫组织化学、免疫荧光染色鉴定神经元纯度,流式细胞术AnnexinV、PI双标检测凋亡率及活性caspase-3、-9表达,Western印迹免疫分析GRP78、细胞色素C蛋白、Bcl-2蛋白表达,Fura-2/AM法荧光分光光度计检测细胞内钙浓度([Ca~(2+)]i).结果 SD乳鼠皮层神经元可纯化体外培养,2 μmol/L毒胡萝卜素作用神经元24、48 h细胞凋亡率分别是17.88%、21.38%,参麦治疗组分别是7.42%、8.16%,两组差异显著(P<0.05).胡萝卜素诱导神经元糖调节蛋白GRP78表达上调,活化caspase-3、-9,使细胞色素C蛋白表达增加,Bcl-2表达减少.参麦注射液促进细胞Bcl-2表达,抑制细胞色素C释放,减少活性caspase-3、-9含量,降低[Ca~(2+)]i浓度.结论参麦注射液能抑制体外培养神经元内质网应激所致的凋亡可能与其降低[Ca~(2+)]i浓度有关.
목적 탐토삼맥주사액대대서피층신경원내질망응격유도조망적보호작용.방법 체외배양SD유서피층신경원세포,면역조직화학、면역형광염색감정신경원순도,류식세포술AnnexinV、PI쌍표검측조망솔급활성caspase-3、-9표체,Western인적면역분석GRP78、세포색소C단백、Bcl-2단백표체,Fura-2/AM법형광분광광도계검측세포내개농도([Ca~(2+)]i).결과 SD유서피층신경원가순화체외배양,2 μmol/L독호라복소작용신경원24、48 h세포조망솔분별시17.88%、21.38%,삼맥치료조분별시7.42%、8.16%,량조차이현저(P<0.05).호라복소유도신경원당조절단백GRP78표체상조,활화caspase-3、-9,사세포색소C단백표체증가,Bcl-2표체감소.삼맥주사액촉진세포Bcl-2표체,억제세포색소C석방,감소활성caspase-3、-9함량,강저[Ca~(2+)]i농도.결론 삼맥주사액능억제체외배양신경원내질망응격소치적조망가능여기강저[Ca~(2+)]i농도유관.
Objective To observe the protect effects of Shenmai (SM) injection on endoplasmic reticulum stress-inducing rat cortical neuronal apoptosis. Methods Primary cortial neurons were cultured in vitro and NSE-positive cells were detected by immunofluorescence staining and immunohistochemistry. The percentage of apoptotic cells and protein expression of caspase-3,-9 were determined by flow cytometric analysis. Protein levels of GRP78, cytochrome C, Bcl-2 were assessed by immunoblotting. Intracellular free calcium concertration [Ca~(2+)]i was measured with fluorophotometry. Results Cortical neurons of neonate rats could be cultured in vitro, the percentage of apoptosis induced by thapsigargin after intervened for 24 and 48 h was 17.88% and 21.38% respectively. GRP78 and cytochrome C protein levels were elevated in cortical neurons in response to thapsigargin. The expression of caspase-3 and -9 were activated and the protein level of Bcl-2 was decreased. SM significantly decreased the percentage of neuronal apoptosis and increased the expression of Bcl-2, reduced neuronal [Ca~(2+)]i and the expression of caspase-3 and -9, inhibited the leakage of cytochrome C.Conclusions SM has effect of anti-neuronal apoptosis induced by endoplasmic reticulum stress in vitro, which might be related to decreasing intracellular free calcium concentration.