补阳还五汤介导Cav1调控Wnt通路对脑缺血小鼠神经细胞凋亡的影响

Effects of Buyang Huanwu Decoction Mediating Cav1 in Regulating Wnt Pathway on Neuronal Cell Apoptosis in Cerebral Ischemia Mice

目的基于小窝蛋白(Cav1)调控Wnt通路探讨补阳还五汤对脑缺血小鼠神经细胞凋亡的影响。方法将雄性野生型(WT)及Cav1-/-(KO)C57BL/6小鼠分别随机分为假手术组、模型组和补阳还五汤组(18.5 g/kg),采用大脑中动脉栓塞法制备脑缺血小鼠模型,并予药物干预14 d。对小鼠进行神经行为学评分,HE染色观察脑组织缺血侧皮质区形态,TUNEL染色检测缺血侧皮质区神经细胞凋亡情况,免疫组化检测缺血侧皮质区凋亡相关蛋白表达及Wnt1、糖原合成酶激酶3β(GSK3β)、β-连环蛋白(β-catenin)蛋白表达。结果与同基因型假手术组比较,模型组小鼠神经行为学评分显著升高,缺血侧皮质区神经细胞呈空泡样改变,细胞核固缩,细胞间隙增宽,神经细胞凋亡率显著升高,Bax、GSK3β表达升高,Bcl-2、Wnt1、β-catenin表达降低(P<0.01);与同基因型模型组比较,补阳还五汤组小鼠神经行为学评分显著降低,缺血侧皮质区病理损伤改善,神经细胞凋亡率降低,Bax、GSK3β表达降低,Bcl-2、Wnt1、β-catenin表达升高(P<0.01);与WT模型组比较,KO模型组小鼠神经行为学评分升高,缺血侧皮质区损伤加重,神经细胞凋亡率显著升高,GSK3β表达升高(P<0.05);与WT补阳还五汤组比较,KO补阳还五汤组小鼠神经行为学评分升高,缺血侧皮质区损伤加重,神经细胞凋亡率显著升高,Bax、GSK3β表达升高,Bcl-2、Wnt1、β-catenin表达降低(P<0.01)。结论补阳还五汤能抑制脑缺血后神经细胞凋亡,其作用机制可能与介导Cav1调控Wnt信号通路,进而影响凋亡相关蛋白表达有关。

Objective To explore the effects of Buyang Huanwu Decoction on neuronal cell apoptosis after cerebral ischemia based on mediating Cav1 in regulating Wnt pathway.Methods Male wild-type(WT)and Cav1-/-(KO)C57BL/6 mice were randomly divided into sham-operation group,model group and Buyang Huanwu Decoction group(18.5 g/kg).Cerebral ischemia model was prepared using middle cerebral artery occlusion method,and drug intervention was given for 14 days.Neurobehavioral score was performed,HE staining was used to observe the morphology of ischemic cortical area of brain tissue,TUNEL staining was used to detect neuronal apoptosis in ischemic cortical area,immunohistochemistry was used to detect the expressions of apoptosis related proteins and Wnt1,glycogen synthase kinase 3β(GSK3β)andβ-catenin protein in ischemic cortical area.Results Compared with the same genotype sham-operation group,the neurobehavioral score of the model group mice significantly increased,neuronal cells in the ischemic cortical area showed vacuolar changes,with nuclear condensation and widened intercellular spaces,the apoptosis rate of nerve cells significantly increased,with increased expressions of Bax,GSK3βand decreased expressions of Bcl-2,Wnt1 andβ-catenin(P<0.01).Compared with the same genotype model group,the neurobehavioral score of mice in Buyang Huanwu Decoction group were significantly decreased,the pathological damage of the ischemic cortical area improved,the apoptosis rate of nerve cells decreased,the expressions of Bax and GSK3βdecreased,and the expressions of Bcl-2,Wnt1 andβ-catenin increased(P<0.01).Compared with the WT model group,the KO model group showed an increase in neurobehavioral score,aggravated damage in ischemic cortical area,significantly increased neuronal apoptosis rate,and increased expression of GSK3β(P<0.05).Compared with the WT Buyang Huanwu Decoction group,the KO Buyang Huanwu Decoction group showed an increase in neurobehavioral score,aggravated damage in ischemic cortical area,significantly increased neuronal apoptosis rate,increased expressions of Bax and GSK3β,and decreased expressions of Bcl-2,Wnt1 andβ-catenin(P<0.01).Conclusion Buyang Huanwu Decoction can inhibit neuronal cell apoptosis after cerebral ischemia,and its mechanism may be related to regulating the expressions of apoptosis-related proteins by mediating Cav1 to regulate the Wnt signaling pathway.