大鼠创伤性颅脑损伤后脑室下区Notchl蛋白表达的时程变化研究

Dynamic changes of Notch1 protein expression in the subventricular zone after traumaticcerebral injury of the rats

目的观察创伤性颅脑损伤（TBI）后脑室下区（SVZ）Notch1蛋白表达的动态变化，探讨其与Notch信号通路开放及TBI后神经干细胞（NSCs）增殖的相关性。方法96只SD大鼠按随机数字表法分为假手术组（仅切开头皮和颅骨开窗，n=16）、对照组（不做任何处理，n=16）和实验组（采用Feeney’s自由落体模型建立中型TBI模型，n=64），其中实验组大鼠依据取材时间不同分为1、3、7、14 d亚组，每亚组16只。采用免疫荧光化学染色检测各组大鼠SVZ区脑组织Notch1/巢蛋白（nestin）双标染色阳性细胞表达，采用Western blotting检测Notch1蛋白表达。结果免疫荧光化学染色结果显示，实验组SVZ区脑组织Notch1/nestin双标染色阳性细胞数量在TBI后1 d时即升高出现峰值，达对照组的2~3倍，其后逐渐下降，14 d时基本恢复正常，4个亚组间差异有统计学意义（P〈0.05）；实验组1、3、7 d亚组Notch1/nestin双标染色阳性细胞数量分别与对照组、假手术组相比差异均有统计学意义（P〈0.05）。Western blotting结果显示，对照组及假手术组Notch1蛋白表达量无明显差别，实验组TBI后1 d时SVZ区Notch1蛋白表达量相对较多，后逐渐下降，14 d时降至正常水平。结论大鼠TBI后可激活Notch1蛋白表达，使Notch信号通路开放，从而影响NSCs增殖的调控。

Objective To observe the dynamic changes of Notchl protein expression in the subventricular zone （SVZ） after traumatic cerebral injury （TBI） in rats, and to explore the correlation between Notch signaling pathway and neural stem cells （NSCs） proliferation after TBI. Methods Ninety-six SD rats were randomly divided into sham-operated group （only opening the scalp and skull window, n=16）, control group （without any treatment, n=16） and experimental group （establishing medium-sized TBI models with Feeney＇s free fall method, n=64）; rats in the experimental group were divided into 4 sub groups according to the execution times （one, three, 7 and 14 d after TBI, n=16）. Immunofluorescence staining was performed to detect the Notchl/nestin labeled cell expressions in SVZ of rots, and Notchl protein expression was detected by Western blotting. Results Immunofluorescence results showed that the Notchl/nestin double staining positive cells in the right SVZbrain tissues of the experimental group on the first day of TBI had peak level, which was 2-3 times of the control group; and then, it gradually declined and basically returned to normal on 146 day of TBI; the differences between the 4 subgroups were statistically different （P〈0.05）. The positive cell counts in the subgroups of first, third, and 76 d after TBI were statistically different as compared with those in the control group and sham operated group （P〈0.05）. Western blotting results indicated that there was no significant difference in the Notchl protein expression between control group and sham operated group; Notchl protein expression in SVZ of the experimental group had peak level on the first day of TBI, gradual decline was noted after that, and normal level was noted on the 146 d of TBI. Conclusion TBI can activate the Notchl protein expression, which induces the opening of Notch signaling pathway so as to affect the regulation of NSCs proliferation.