摘要
目的:探讨加味四君子汤对大鼠脑缺血再灌注(I/R)损伤后缺血侧海马区脑组织纤维蛋白-5(Fibulin-5),磷酸化蛋白激酶B(p-Akt)的表达及对神经细胞失巢凋亡的影响。方法:将60只SPF级雄性SD大鼠随机分为假手术组、模型组、依达拉奉组(3.2 mg·kg^(-1)),加味四君子汤高、中、低剂量组(19.08,9.54,4.77 g·kg^(-1))。采用线栓法制备大鼠大脑中动脉栓塞(MCAO)模型,7 d后处死大鼠,处死前进行神经功能缺损评分,采用苏木素-伊红(HE)染色进行组织病理学观察,原位末端标记法(TUNEL)染色检测神经细胞凋亡指数,免疫组化法和蛋白免疫印迹法(Western blot)检测缺血侧海马区脑组织Fibulin-5及p-Akt蛋白表达情况。结果:神经功能缺损评分结果显示,与假手术组比较,模型组大鼠神经功能缺损评分显著升高(P<0.01);与模型组比较,依达拉奉组、加味四君子汤高、中、低剂量组大鼠神经功能缺损评分均明显降低(P<0.05,P<0.01)。免疫组化结果显示,与假手术组比较,模型组大鼠Fibulin-5,p-Akt蛋白表达,神经细胞凋亡指数均明显增高(P<0.05,P<0.01);与模型组比较,依达拉奉组、加味四君子汤高、中、低剂量组大鼠Fibulin-5,p-Akt蛋白表达均明显增加(P<0.05,P<0.01),神经细胞凋亡指数均明显降低(P<0.05,P<0.01)。Western blot结果显示,与假手术组比较,模型组Fibulin-5,p-Akt蛋白相对表达显著下调(P<0.01);与模型组比较,依达拉奉组、加味四君子汤高、中、低剂量组Fibulin-5,p-Akt蛋白表达均明显上升(P<0.05,P<0.01)。结论:加味四君子汤可能通过稳定神经元细胞外基质(ECM)Fibulin-5,从而增加了ECM对细胞的黏附作用,促进p-Akt蛋白的表达,从而抑制神经细胞凋亡,保护脑缺血损伤。
Objective: To investigate the effect of modified Si Junzitang on the expression of fibrous protein-5(Fibulin-5),phosphorylated protein kinase B(p-Akt)in hippocampus of rats after cerebral ischemiareperfusion(I/R)injury and the anoikis of nerve cells. Method: The 60 male SD rats of SPF grade were randomly divided into sham operation group,model group,Edaravone group(3.2 mg·kg^(-1))and modified Si Junzitang high,medium and low-dose groups(19.08,9.54,4.77 g·kg^(-1)). The middle cerebral artery occlusion(MCAO)model was established by suture method,the rats were killed 7 days later,neurological deficit score was evaluated before the death,histopathological observation was performed by hematoxylin eosin staining,apoptosis index of nerve cells was detected by TdT-mediated dUTP nick end labeling(TUNEL)staining,the expression of Fibulin-5,p-Akt and protein in ischemic hippocampus were detected by immunohistochemistry and Western blot. Result:The neurological deficit score showed that,compared with the sham operation group,the neurological deficit score of the model group was significantly increased(P<0.01),compared with model group,the neurological deficit score of Edaravone group,the high,medium,low dose groups of modified Si Junzitang were decreased(P<0.05,P<0.01). Immunohistochemical results showed that,compared with the sham operation group,the expression of Fibulin-5,p-Akt protein and the apoptosis index of nerve cells in the model group were significantly increased(P<0.05,P<0.01),compared with model group,the protein expressions of Fibulin-5 and p-Akt in Edaravone group,high,medium and low-dose groups of modified Si Junzitang were significantly increased(P<0.05,P<0.01),and the apoptosis index of nerve cells was obvious,there was a significant decrease(P<0.05,P<0.01). Western blot results showed that,compared with the sham operation group,the relative expression of Fibulin-5 and p-Akt protein in the model group was significantly downregulated(P<0.01), compared with model group, the protein expressions of Fibulin-5 and p-Akt in the Edaravone group,the high,medium and low-dose groups of modified Si Junzitang were significantly upregulated(P<0.05,P<0.01). Conclusion: The modified Si Junzitang may stabilize the extracellular matrix(ECM)Fibulin-5,increase the adhesion of ECM to cells and promote the expression of p-Akt protein,thus inhibiting neuronal apoptosis and protecting cerebral ischemia injury.
作者
陈岩岩
李花
刘旺华
唐冰镕
李宇翔
王浩瞩
CHEN Yan-yan;LI Hua;LIU Wang-hua;TANG Bing-rong;LI Yu-xiang;WANG Hao-zhu(Key Laboratory of Diagnostics of Chinese Medicine in Hunan Province,Hunan University of Chinese Medicine,Changsha 410208,China)
出处
《中国实验方剂学杂志》
CAS
CSCD
北大核心
2021年第1期112-120,共9页
Chinese Journal of Experimental Traditional Medical Formulae
基金
国家自然科学基金项目(81473567)
湖南省自然科学基金项目(2019JJ40219)
湖南省中医药科研计划重点项目(202008)
湖南省大学生研究性学习和创新性实验计划项目(422)。
