芪仙通络方对脑缺血大鼠内源性神经干细胞再生的影响及机制

目的:探讨芪仙通络方对脑缺血大鼠内源性神经干细胞再生的影响及可能机制。方法:采用线栓法复制大脑中动脉阻塞(middle cerebral artery occlusion,MCAO)局灶性脑缺血模型;将160只大鼠随机分为正常组,假手术组,模型组,吡拉西坦组(吡拉西坦片灌胃),芪仙通络方组(芪仙通络方灌胃);5-溴脱氧尿嘧啶核苷(5-bromo-2-deoxy uridine,Brd U)腹腔注射标记脑内增殖细胞,造模后3,7,14,28 d,分别采用免疫荧光Brd U/神经元核抗原(neuronal nuclei antigen,Neu N),Brd U/胶质纤维酸性蛋白(glial fibrillary acidic protein,GFAP)双标法检测缺血海马区新生的神经元及星形胶质细胞,并计算其双标阳性细胞率,免疫组化法检测缺血侧脑内脑源性神经营养因子(brain-derived neurotrophic factor,BDNF)的表达。结果:术后7,14,28 d,芪仙通络方组、吡拉西坦组Brd U/Neu N,Brd U/GFAP双标阳性细胞率均高于模型组(P0.01),且各时间点芪仙通络方组Brd U/Neu N阳性细胞均高于吡拉西坦组(P0.05),而仅在术后7 d,芪仙通络方组Brd U/GFAP双标阳性细胞率高于吡拉西坦组(P0.05),随后均呈下降趋势;与模型组比较,术后各时间点芪仙通络方组、吡拉西坦组缺血脑内BDNF表达均升高(P0.01),芪仙通络方组BDNF表达于术后3 d达到高峰,之后水平虽有所下降,但仍高于吡拉西坦组(P0.05)。结论:芪仙通络方可促进脑缺血大鼠内源性神经干细胞再生,以恢复期和后遗症期明显,而早期活化星形胶质细胞及上调BDNF的表达可能是其作用机制之一。

Effect and Mechanism of Qixian Tongluo Decoction on Regeneration of Endogenous Neural Stem Cells After Cerebral Ischemia in Rats

Objective: To investigate the effect and possible mechanism of Qixian Tongluo decoction on endogenous neural stem cells regeneration after cerebral ischemia in rats. Method: The focal cerebral ischemia model was established by the method of middle cerebral artery occlusion (MCAO). A total of 160 rats were randomly divided into normal group, sham operation group, model group, piracetam group (piracetam tablets intragastric administration), and Qixian Tongluo decoction group (Qixian Tongluo decoction intragastric administration). 5-Bromodeoxyuridine (BrdU) was injected intraperitoneally to label proliferating cells in brain; at day 3, 7, 14 and 28 after MCAO modeling, the neonatal neurons and astrocytes in the ischemic hippocampus were detected by immunofluorescence BrdU/neuron nuclear antigen (NeuN) and BrdU/glial fibrillary acidic protein (GFAP) double staining, and then the positive rate of double labeling cells was calculated. In addition, the expression level of brain derived neurotrophic factor (BDNF) in the ischemic brain was detected by immunohistochemistry. Result: The percentages of BrdU/NeuN and BrdU/GFAP double-labeled positive cells in Qixian Tongluo decoction group and piracetam group were all higher than those in the model group at day 7, 14 and 28 after cerebral ischemia (P<0.01). The percentages of BrdU/NeuN positive cells in Qixian Tongluo decoction group were all higher than those in piracetam group at different time points (P<0.05); however, the percentage of BrdU/GFAP double-labeled positive cells in Qixian Tongluo decoction group was higher than that of piracetam group only at day 7 after MCAO (P<0.05), and then the percentages were decreased in both groups. As compared with the model group, the expression levels of BDNF in the ischemic brain of Qixian Tongluo decoction group and piracetam group were all higher at different time points after cerebral ischemia (P<0.01). The expression level of BDNF in Qixian Tongluo decoction group reached the peak at day 3 after MCAO, and then declined at the following time points, but still higher than that in the piracetam group (P<0.05). Conclusion: Qixian Tongluo decoction can promote endogenous neural stem cells regeneration in rats with cerebral ischemia especially during convalescence and sequelae stages, and the early activation of astrocytes and upregulation of BDNF in ischemia brain may be one of its mechanisms.