谷氨酸激活神经干细胞SHH通路机制的研究

Study on SHH pathway of activating neural stem cells by glutamate

目的探讨谷氨酸通过N-甲基-D-天冬氨酸受体1(NMDAR1)激活SHH通路促进神经干细胞增殖的机制。方法将5只3日龄SD大鼠断头取脑分离神经干细胞(NSC);将NSC分为5组:正常组、谷氨酸刺激组、Conantokin-R组、SHH-shRNA组和shRNA对照组;通过Real-time RCR、Western Blotting、CCK-8检测NSC在NMDAR1受到抑制及SHH被shRNA下调情况下受到谷氨酸刺激后SHH、Nestin的表达变化及细胞增殖能力的变化;采用单因素方差分析中的LSD法进行统计分析。结果与正常组相比,谷氨酸刺激组和shRNA对照组SHH和Nestin的mRNA和蛋白表达明显上升(均P<0.05),Conantokin-R组SHH和Nestin的mRNA和蛋白表达无明显变化(均P>0.05)。SHHshRNA组SHH的mRNA和蛋白表达明显下调(均P<0.05),Nestin的mRNA和蛋白表达无明显变化(均P>0.05)。结论谷氨酸通过NMDAR1介导激活SHH通路。

Objective To investigate the mechanism of glutamate activating SHH pathway through N-methyl-D-aspartic acid receptor（ NMDAR1 ） to promote neural stem cell （NSC） proliferation. Methods Five 3-day-old SD rats were decapitated and neural stem ceils （NSC） were isolated. NSC was divided into five groups ： normal group, glutamic acid stimulation group, Conantokin-R group, SHH-shRNA group, and shRNA control group;the change in the expression of SHH and Nestin, as well as the change of cell proliferation, under the conditions of inhibited NMDAR1 and down-regulated SHH by shRNA, were tested by Real-time RCR, Western Blotting. LSD method in ANOVA was used for statistical analysis. Results Compared with the control group, the mRNA and protein of SHH and Nestin in the glutamine stimulation group and the shRNA control group increased （ all P 〈 0. 05 ） ; the mRNA and protein of SHH and Nestin in Conantokin-R group had no significant difference （ all P 〉 0. 05 ）. The mRNA and protein of SHH in SHH-shRNA group was significantly decreased （ all P 〈 0. 05 ） ; the expression of mRNA and protein of Nestin had no significant difference （ all P 〉 0.05 ）. Conclusion Glutamate activates SHH pathway by inducing NMDAR1.