补阳还五汤对去势脑缺血雌性大鼠海马神经干细胞增殖及ERK/CREB信号通路的影响

目的 探讨补阳还五汤对去势脑缺血雌性大鼠海马神经干细胞（neural stem cells，NSCs）增殖及ERK/CREB信号通路的影响。方法 采用双侧卵巢切除术（OVX）结合大脑中动脉阻塞（MCAO）法复制去势雌性大鼠脑缺血复合模型，将造模后的36只大鼠随机分为双假手术组、复合模型组、雌激素组、雌激素+G15组、补阳还五汤组及补阳还五汤+G15组6组，每组6只，术后24 h给予相应药物灌胃连续干预14 d，同时每天腹腔注射BrdU、G15分别标记增殖细胞和阻断GPER-1。采用免疫荧光BrdU/Nestin双标法检测各组大鼠缺血侧海马齿状回NSCs增殖情况，免疫组化SP法检测ERK1/2、CREB1磷酸化蛋白表达水平。结果 补阳还五汤组和雌激素组缺血侧海马DG区BrdU/Nestin双标阳性细胞及pERK1/2、pCREB1阳性细胞表达均增加，与复合模型组比较有显著性差异（P<0.05），但补阳还五汤组要多于雌激素组（P<0.05）。与补阳还五汤组比较，补阳还五汤+G15组缺血侧海马DG区BrdU/Nestin双标阳性细胞及及pERK1/2、pCREB1阳性细胞表达均减少（P<0.05）。结论 补阳还五汤可以促进去势脑缺血雌性大鼠缺血侧海马DG区神经干细胞增殖，激活ERK/CREB信号通路，可能是其发挥类雌激素作用、促进内源性神经再生作用机制之一。

Effects of Buyang Huanwu Decoction on Neural Stem Cells Proliferation and ERK/CREB Signaling Pathway in Hippocampus after Cerebral Ischemia in Ovariectomized Female Rats

Objective To investigate the effects of Buyang Huanwu Decoction on neural stem cells(NSCs) proliferation and ERK/CREB signaling pathway in hippocampus after cerebral ischemia in ovariectomized female rats. Methods The ovariectomized female rats with cerebral ischemia model was established by the methods of ovariectomy(OVX)and middle cerebral artery occlusion (MCAO). The 36 rats were randomly divided into 6 groups, double sham-operation group, compound model group, estrogen group, estrogen+G15 group and Buyang Huanwu Decoction group and Buyang Huanwu Decoction+G15 group, 6 rats in each group. The rats were given corresponding medicine intragastric administration for 14 days from 24 hours after operation, at the same time, BrdU and G15 were intraperitoneal injected to label the proliferation cells and block GPER-1 in each day, respectively. The NSCs proliferation of ischemic hippocampal dentate gyrus in rats was detected using immunofluorescence BrdU/nestin double staining with paraffin sections, the SP method of immunohistochemistry was used to detect the expression level of ERK1/2, CREB1 phosphorylation protein. Results Compared with the composite model group, the number of double positive cells and pERK1/2, pCREB1 positive cells in ischemic hippocampal DG area of Buyang Huanwu Decoction group and estrogen group were increased (P<0.05), but the indicators in Buyang Huanwu decoction group were higher than estrogen group (P<0.05). Compared with Buyang Huanwu Decoction group, the number of double positive cells and pERK1/2, pCREB1 positive cells in ischemic hippocampal DG area of Buyang Huanwu Decoction+G15 group were decreased (P<0.05). Conclusion Buyang Huanwu Decoction can promote ischemic hippocampal NSCs proliferation in ovariectomized female rats after cerebral ischemia, and to activate ERK/CREB signaling pathway, which may be one of the mechanism of its estrogen-like effect and endogenous neurogenesis.