摘要
目的探讨CAPN1 RNAi对Aβ25-35诱导的原代神经细胞的神经毒性及凋亡的影响。方法利用Aβ25-35诱导原代胎鼠皮质神经元毒性损伤,观察CAPN1 RNAi对神经元细胞活力、凋亡的影响。实验分为4组:对照组(NC)、NC+Aβ、CAPN1 shRNA+Aβ、CAPN1 shRNA+Aβ+P25。CCK-8法检测神经元细胞活力的变化情况;Annexin V-PI方法进行细胞凋亡检测;Real-Time PCR检测转染CAPN1 RNAi后CAPN1 mRNA的表达水平;Western blotting检测CAPN1、CDK5、GSK3β、p-tau/tau蛋白表达的变化。结果 CCK8检测结果显示,与对照组相比,NC+Aβ组细胞活力明显下降,转染CAPN1 shRNA后细胞活力有明显改善(P<0.05);与CAPN1 shRNA+Aβ组相比,CAPN1 shRNA+Aβ+P25组细胞活力无显著性差异(P>0.05)。Annexin V-PI凋亡实验结果表明:与NC+Aβ组相比,转染CAPN1 shRNA后细胞早期凋亡比例显著降低(P<0.01)。RT-PCR检测结果表明:与NC+Aβ组相比,CAPN1 shRNA+Aβ组CAPN1 mRNA的表达显著得到抑制。Western blot发现NC+Aβ组中CAPN1、CDK5、GSK3β、p-tau蛋白表达水平均显著升高,而转染CAPN1 shRNA后均显著下调(P<0.01)。结论 CAPN1RNAi可以显著改善Aβ25-35诱导的原代神经细胞的神经毒性及凋亡。
Objective To investigate the effect of CAPN1 RNAi on the neurotoxicity and apoptosis of primary neurons induced by Aβ 25-35. Methods The primary cultured rat cortical neurons were induced by Aβ 25-35 and the effect of CAPN1 RNAi on neuronal cell viability and apoptosis was observed.The experiment was divided into four groups：control group （NC）,NC＋Aβ,CAPN1 shRNA＋Aβ,CAPN1 shRNA＋Aβ＋P25.Cell viability was detected by CCK-8 assay.Annexin V-PI was used to detect apoptosis.CAPN1 mRNA expression was detected by Real-Time PCR and CAPN1,CDK5,GSK3β,p-tau/tau protein expression were detected by western blot. Results Compared with control group,the cell viability of NC＋Aβ group was significantly lower than that of control group （ P 〈0.05）.Compared with CAPN1 shRNA＋Aβ group,there was no significant difference in cell viability between CAPN1 shRNA＋Aβ＋P25 group （ P 〉0.05）.The results of Annexin V-PI apoptosis assay showed that compared with NC＋Aβ group,the rate of early apoptosis of CAPN1 shRNA transfected cells was significantly decreased （ P 〈0.01）.The results of RT-PCR showed that CAPN1 mRNA expression was significantly inhibited in CAPN1 shRNA＋Aβ group compared with NC＋Aβ group.Western blot showed that the expression of CAPN1,CDK5,GSK3β and p-tau in NC＋Aβ group were significantly increased,while the CAPN1 shRNA transfected significantly reduced （ P 〈0.01）. Conclusion CAPN1 RNAi can significantly improve the neurotoxicity and apoptosis of primary neurons induced by Aβ 25-35.
作者
杨保平
罗拯
邹可
唐震宇
YANG Baoping;LUO Zheng;ZOU Ke(The Departments of Neurology,The Second Affiliated Hospital of Nanchang University,Nanchang 330000,Chin)
出处
《中风与神经疾病杂志》
CAS
2018年第6期488-491,共4页
Journal of Apoplexy and Nervous Diseases
基金
江西省自然科学基金(20151BAB205024)
