慢病毒介导的shRNA对大鼠脑纹状体GCDH蛋白的沉默效应研究

Silencing of GCDH expression by Lentivirus-Mediated RNA Interference in Brain Striatal of Rats

目的：利用已成功构建的慢病毒载体，检测慢病毒介导的GCDH基因沉默对大鼠脑纹状体GCDH蛋白表达下调的能力。方法：30只健康雄性刚断乳34周SD大鼠随机分为绿色荧光蛋白（GFP）对照病毒注射组（对照组）和GCDH慢病毒注射组（GCDH组）各15只。各组分别在脑立体定位仪下定位纹状体组织后单侧给予GFP对照病毒2耻L和GCDH慢病毒原液2灿。注射后14d及1、3月，分别处死大鼠，并取注射区域的脑纹状体组织蛋白质通过Westemblot及石蜡切片免疫组化法检测大鼠纹状体GCDH蛋白表达水平，同时测定肝。肾功能、血清有机酸及Rotarod行为学。结果：GCDH慢病毒注射14d及l、3月后，GCDH组可检测到注射区域GCDH蛋白在脑纹状体区域表达有明显降低，同对照组比较差异有统计学意义（P〈0．05），2组肝肾功能、血清有机酸及行为学未发现明显差异。结论：成功注射GCDH慢病毒到刚断乳34周SD大鼠脑纹状体，并在注射后不同时间均检测到GCDH蛋白的表达稳定下调。

Objective ：To investigate the silencing effect of injecting the lentivirus-mediated shRNA to GCDH gene on the expression of GCDH in rat striatum. Methods： Three or four weeks old male SD rats were randomly divided into two groups, the GFP gene lentiviral vector injection group （Control group） and the GCDH lentiviral vector injection group （GCDH group）. Each group contained 15 rats. Lentiviral vectors expressing GCDH shRNA were administered by striatal injection. The expression of GCDH protein was detected by Western blot and im- mtmohistochemistry day 14, 1 and 3 months after the injection. Liver and renal function, serum organic acids and Rotarod Test were also detected. Results： GCDH gene lentiviral vector can down-regulate the expression of GCDH protein in rat striatum. Immunohistochemistry and Western blot detection show that the level of GCDH protein was decreased in the GCDH group compared with that in the controls（P〈0,05）. However, there was no difference in liver and renal fimctions, serum organic acids and the motor function between the two groups. Conclusion： Injecting lentivirus-mediated shRNA GCDH gene in rat striatum effectively decreases the expression of GCDH.