脑脉通联合骨髓干细胞动员对脑缺血大鼠神经细胞凋亡及Bax、Bcl-2表达的影响

Influences of Naomaitong Granules combined with mobilization of BMSC on neuron apoptosis and expressions of Bax and Bcl-2 in rats with cerebral ischemia

目的观察脑脉通联合骨髓干细胞(BMSCs)动员对脑缺血大鼠神经细胞凋亡的作用,并从对凋亡相关基因Bax、Bcl-2表达及Bax/Bcl-2变化的影响探讨其可能的作用机制。方法大鼠随机分组。线栓法制备局灶性脑缺血(MCAO)动物模型。动员组和联合组大鼠分别于术前3d和术后2d皮下注射人重组粒细胞集落刺激因子(rG-CSF);脑脉通灌胃用药。术后2、3、7、14d取材,测定外周血和脑组织CD34+细胞;观察脑组织病理改变;检测神经细胞凋亡;测定脑组织Bax、Bcl-2表达,计算Bax/Bcl-2比值。结果模型组大鼠外周血和脑组织CD34+细胞增加,分别于3d和7d达到峰值;各治疗组尤其联合组脑组织CD34+增强明显。缺血后7d大鼠脑组织损伤明显,动员组14d和各联合组的损伤减轻。各模型组神经细胞凋亡增多、Bax表达增加、2d和3d的Bax/Bcl-2增高。各联合组神经细胞凋亡减轻、Bax表达减弱,7d及14d的Bcl-2表达增强,各时间点的Bax/Bcl-2降低。联合组较动员组2d和3d的凋亡细胞、各时间点的Bax表达减轻和Bax/Bcl-2降低,7d及14d的Bcl-2表达增强。结论脑缺血可引起BMSCs进入外周血及向脑组织归巢,达到峰值的时间存在差异,脑脉通可使动员后的骨髓干细胞增多;脑缺血引起的神经细胞凋亡与Bax表达上调相关,脑脉通可使BMSCs动员抑凋亡的作用增强并延长,其作用可能与早期(2d、3d)下调促凋亡基因Bax和后期(7d、14d)上调Bcl-2表达有关。

Objective To observe the effect of Naomaitong Granules combined with the mobilization of bone marrow stem cells （ BMSC） on neuron apoptosis in rats with cerebral ischemia,and investigate the possible mechanism from the influences of the combination on the expressions of relevant genes-Bax and Bcl-2,and changes of ratio of Bax to Bcl-2. Methods All rats were randomly divided into groups and the model of middle cerebral artery occlusion （ MCAO） was established by nylon thread approach. The rats of the mobilization group and combination group were injected hypodermically recombinant human granulocyte colony-stimulating factor （ rG-CSF） 3 days before the operation and 2 days after the operation. Naomaitong Granules was administrated intragastrically. The samples were taken 2,3,7 and 14 days after the operation respectively. The levels of CD34 ＋ cells in peripheral blood and brain weredetected,the pathological changes of cerebral tissue were observed,the neuron apoptosis was examined, the expressions of Bax and Bcl-2 were detected,and the ratio of Bax to Bcl-2 was calculated. Results In the model group the levels of CD34 ＋ cells in peripheral blood and brain increased and reached the peak on the 3rd day and 7th day respectively. In all treatment groups,and especially in the combination group, the levels of CD34 ＋ cells increased significantly. The lesion of rat brain was significant 7 days after the ischemia,and after 14 days the lesion was alleviated in the mobilization group and combination group. In the model group the neuron apoptosis was improved,Bax expression increased,and the ratio of Bax to Bcl-2 increased on 2nd and 3rd day. In the combination group the neuron apoptosis was reduced,Bax expression decreased,Bcl-2 expression increased on 7th day and 14th day,and the ratio of Bax to Bcl-2 decreased at different time points. The comparison between the combination group and mobilization group showed that the neuron apoptosis was reduced on 2nd〗day and 3rd day,Bax expression decreased,ratio of Bax to Bcl-2 decreased,and Bcl-2 expression decreased on 7th day and 14th day at different time points in the combination group. Conclusion BMSC can be induced by cerebral ischemia entering into peripheral blood and homing in cerebral tissue,and the peak time is different. Naomaitong Granules can increase mobilized BMSC. The neuron apoptosis induced by cerebral ischemia is correlated to up-regulation of Bax expression. Naomaitong Granules can improve and prolong the anti-apoptosis effect of BMSC mobilization,which may be related to that it regulates downward Bax expression at early time （ 2nd day and 3rd day） and regulates upward Bcl-2 expression at later time （ 7th day and 14th day） .