苦参碱对缺氧/复氧小鼠海马神经元HT22细胞活力、凋亡的影响及机制研究

Effect of matrine on the activity and apoptosis of HT22 cells in hippocampus neurons of hypoxia/reoxygenation mice and its mechanism

目的探讨苦参碱对缺氧/复氧(H/R)小鼠海马神经元HT22细胞活力、凋亡及脑源性神经营养因子(BDNF)/酪氨酸激酶受体B(TrkB)信号通路的影响。方法将处于对数生长期的HT22细胞分为对照组、H/R组及苦参碱低、中、高浓度组。对照组HT22细胞用无血清DMEM培养基在正常环境下培养28 h;H/R组HT22细胞在缺氧培养箱中维持4 h后在常氧条件下培养24 h;苦参碱低、中、高浓度组的H/R处理同H/R组,同时给予浓度分别为10、20、30μmol/L的苦参碱进行干预,培养24 h。检测HT22细胞活力、凋亡情况及凋亡相关蛋白[B淋巴细胞瘤-2基因(Bcl-2)、Bcl-2相关X蛋白(Bax)、半胱氨酸天冬氨酸蛋白水解酶-3(Caspase-3)]和BDNF/TrkB信号通路蛋白水平。结果与对照组比较,H/R组HT22细胞吸光度值、存活率、Bcl-2、BDNF、TrkB蛋白水平降低,细胞凋亡率、Bax、Caspase-3蛋白水平升高,差异有统计学意义(P<0.05);与H/R组比较,苦参碱低、中、高浓度组HT22细胞吸光度值、存活率、Bcl-2、BDNF、TrkB蛋白水平依次升高,细胞凋亡率、Bax、Caspase-3蛋白水平依次降低,差异有统计学意义(P<0.05),呈浓度依赖性。结论苦参碱能减轻H/R导致的神经元HT22细胞活力的降低,抑制HT22细胞凋亡,其机制可能与激活BDNF/TrkB信号通路有关。

Objective To investigate the effects of matrine on viability,apoptosis and brain derived neurotrophic factor(BDNF)/tyrosine kinase receptor B(TrkB)signaling pathway of hippocampal neurons HT22 cells in hypoxia/reoxygenation(H/R)mice.Methods HT22 cells in logarithmic growth stage were divided into control group,H/R group and matrine low,medium high concentration groups.In the control group,HT22 cells were cultured in serum-free DMEM medium under normal environment for 28 hours;in H/R group,HT22 cells were maintained in hypoxia incubator for 4 hours and cultured under normoxia for 24 hours;the H/R treatment of matrine low,medium and high concentration groups was the same as that of H/R group,at the same time,matrine with concentrations of 10,20 and 30μmol/L was given for intervention,and cultured for 24 hours.The viability and apoptosis of HT22 cells and the levels of apoptosis related proteins[B lymphocytoma-2 gene(Bcl-2),Bcl-2 related X protein(Bax),Caspase-3]and BDNF/TrkB signal pathway protein were detected.Results Compared with the control group,the absorbance value,survival rate,Bcl-2,BDNF and TrkB protein levels of HT22 cells in H/R group were decreased,while the apoptosis rate,Bax and Caspase-3 protein levels were increased,the difference was statistically significant(P<0.05);compared with H/R group,the absorbance value,survival rate,Bcl-2,BDNF and TrkB protein levels of HT22 cells in matrine low,medium and high concentration groups were increased in turn,while the apoptosis rate,Bax and Caspase-3 protein levels were decreased in turn,the difference were statistically significant(P<0.05),in a concentration dependent manner.Conclusion Matrine can reduce the decrease of neuronal HT22 cell viability caused by H/R and inhibit HT22 cell apoptosis,its mechanism may be related to the activation of BDNF/TrkB signal pathway.