抑制整合素和黏着斑激酶促进神经干细胞对脑缺血后的神经修复

Inhibition of integrin-focal adhesion kinase promotes neural repair of neural stem cells after cerebral ischemia in rats

目的探究抑制整合素-黏着斑激酶(FAK)信号通路对脑缺血动物模型神经干细胞(NSC)移植后NSC增殖的影响。方法选择12只健康SD大鼠作为对照组,另将线栓法成功构建脑缺血模型大鼠36只。随机分为脑缺血组、NSC移植组和西仑吉肽组,每组12只。后2组大鼠注射10μl NSC溶液(细胞浓度为1×10^(6)个/L),西仑吉肽组还腹腔注射整合素抑制剂西仑吉肽100μg/kg。水迷宫实验分析大鼠神经功能,Western blot检测整合素和FAK蛋白表达,TUNEL染色检测神经细胞凋亡,免疫荧光染色检测NSC增殖。结果与对照组比较,脑缺血组逃避潜伏期显著升高,而穿越平台次数显著降低(P<0.05);NSC移植组逃避潜伏期显著低于脑缺血组[(34.02±4.02)s vs(48.79±4.94)s,P<0.05],而穿越平台次数显著高于脑缺血组[(4.86±0.54)次vs(2.74±0.42)次,P<0.05];西仑吉肽组逃避潜伏期显著高于NSC移植组,而穿越平台次数显著低于NSC移植组(P<0.05)。与对照组比较,脑缺血组大鼠整合素、FAK蛋白表达、凋亡指数和5溴脱氧尿嘧啶核苷(BrdU)/神经元核抗原(NeuN)阳性细胞数显著升高(P<0.05);NSC移植组整合素、FAK蛋白表达、BrdU/NeuN阳性细胞数显著高于脑缺血组,凋亡指数显著低于脑缺血组(P<0.05);西仑吉肽组整合素、FAK蛋白表达、BrdU/NeuN阳性细胞数显著低于NSC移植组,凋亡指数显著高于NSC移植组(P<0.05)。结论外源性NSC增殖、分化为神经元的过程中,整合素-FAK信号通路被激活,抑制整合素-FAK信号通路会影响NSC对脑缺血大鼠神经功能恢复的促进作用。

Objective To investigate the effect of inhibiting integrin-focal adhesion kinase(FAK)signaling pathway on the proliferation of neural stem cells(NSC)after transplantation in cerebral ischemic animal model.Methods The rat model of cerebral ischemia was established by wire plug inserting,and then the 36 SD rats successfully modeled were randomly divided into cerebral ischemia group,cerebral ischemia+NSC group and cerebral ischemia+NSC+cilengitide group(n=12).The rats in the latter 2 groups were given NSC transplantation(10μl,1×10^(6)cells/L),and those out of the last group received intraperitoneal injection of 100μg/kg cilengitide(integrin inhibitor).Another 12 healthy SD rats served as the control group.The nerve function of rats was analyzed by water maze test,the expression of integrin and FAK was detected by Western blotting,the apoptosis of nerve cells was detected by TUNEL,and the proliferation of NSC was detected by immunofluorescence staining.Results The escape latency was prolonged and the number of crossing platforms was decreased in the cerebral ischemia group than the control group(P<0.05).The cerebral ischemia+NSC group obtained significantly shorter escape latency(34.02±4.02 s vs 48.79±4.94 s)and higher times of crossing platforms(4.86±0.54 times vs 2.74±0.42 times)when compared with the cerebral ischemia group(P<0.05).Cilengitide treatment reversed the effects of NSC transplantation on the escape latency and the number of crossing plat-forms(P<0.05).Compared with the control group,the expression of integrin and FAK,apopto-sis index and the number of 5-bromodeoxyuridine(BrdU)/neuronal nuclear antigen(NeuN)posi-tive cells were significantly increased in the cerebral ischemia group(P<0.05).The protein lev-els and the number of BrdU/NeuN positive cells were significantly higher,and the apoptosis index was significantly lower than in cerebral ischemia+NSC group than the cerebral ischemia group(P<0.05);The protein levels and the number of double positive cells were significantly lower,and the apoptosis index was statistically higher in the cerebral ischemia+NSC+cilengitide group than the cerebral ischemia+NSC group(P<0.05).Conclusion During the proliferation of exog-enous NSC and differentiation into neurons,inhibiting the integrin-FAK signaling pathway will affect the promotion of NSC on the recovery of neural function in cerebral ischemic rats.