组织金属蛋白酶抑制物对大鼠脑外伤后神经细胞凋亡及基质金属蛋白酶9、胱天蛋白酶3表达的影响

The Effect of Tissue Inhibitor of Metalloproteinase on the Apoptosis of Neuron and Expressions of Matrix Metalloproteinase 9,Caspase-3 after Brain Trauma in Rats

目的探讨组织金属蛋白酶抑制物（TIMP）对大鼠脑外伤后海马区脑组织中神经细胞凋亡及基质金属蛋白酶9（MMP-9）、胱天蛋白酶3（caspase-3）表达的影响,以了解TIMP在大鼠脑外伤后的脑保护作用及其机制。方法将90只体质量为250~350 g的SD大鼠按抽签法随机分为3组,各30只。正常对照组：不作处理;生理盐水组：制作大鼠弥漫性轴索损伤模型前0.5 h向侧脑室注入0.9%氯化钠溶液10μL;TIMP组：制作大鼠弥漫性轴索损伤模型前0.5 h向侧脑室注入TIMP10μL。将生理盐水组及TIMP组大鼠利用颅脑瞬间旋转模型致鼠脑弥漫性轴索损伤。每组分别于制作脑损伤模型后1、3、6、24、72、168 h共6个时间点处死大鼠,每个时间点随机选取大鼠5只,取3组大鼠海马区脑组织标本,用脱氧核糖核苷酸末端转移酶介导的缺口末端标记法检测凋亡神经细胞,用免疫组织化学法测定MMP-9、caspase-3表达,并比较分析同一时间点3组间的差异。结果 1正常对照组中少见凋亡神经细胞,受损伤两组中凋亡神经细胞数明显增多（P〈0.05）,TIMP组各时间点凋亡神经细胞数均显著低于生理盐水组（P〈0.05）。2与正常对照组比较,生理盐水组和TIMP组中MMP-9、caspase-3有明显表达（P〈0.05）,与生理盐水组比较,TIMP组中在各时间点MMP-9、caspase-3表达显著下降（P〈0.05）。结论脑外伤后大鼠海马区脑组织中存在神经细胞凋亡及MMP-9、caspase-3表达的变化,而TIMP能抑制MMP-9表达及caspase-3的激活,使抗凋亡作用增强,减少脑外伤后大鼠海马区脑组织中神经细胞凋亡,表明其对脑外伤后大鼠具有一定的脑保护作用。

Objective To explore the impact of the tissue inhibitor of metalloproteinase（ TIMP） on the nerve cells＇ apoptosis and the expressions of matrix metalloproteinase 9（ MMP-9） s,caspase-3 in the rats＇ hippocampus tissue after brain trauma,to study the protections and its＇ mechanisms of the TIMP for the rats＇ brain after brain trauma. Methods A total of 90 Sprauge-Dawley rats weighted from 250 to 350 grams were randomly divided into three groups,30 in each. Control group： no treatment; physiological saline group（ PS group） ： 0. 9% Na Cl 10 μL was injected to the cerebral ventricle 0. 5 h before making brain—diffuse axonal injury model; TIMP group： TIMP 10 μL was injected to the cerebral ventricle 0. 5 h before making brain—diffuse axonal injury model. Then PS group and TIMP group were made into diffuse axonal injury models by instant rotation model. All rats were killed at 1,3,6,24,72 and 168 h after injury（ five rats randomly each time） respectively. The nerve cells＇ apoptosis in the rats＇ hippocampus tissue was detected by TUNEL and the expressions of MMP-9,caspase-3 in the rats＇ hippocampus tissue were detected by immunohistochemistry staining. The differences among three groups at the same time point were compared and analyzed. Results1 Neural cell apoptosis was rarely observed in normal control group,while in the other two groups the numbers increased significantly（ P〈0. 05）,the number of apoptosis neuronal cells of TIMP group at each time point was significantly lower than the PS group（ P〈0. 05）. 2 Compared with the normal control group,MMP-9 and caspase-3 of the injury group had obvious expression（ P〈0. 05）,compared with the PS group,the caspase-3 and MMP-9 expression in TIMP group at each time point was significantly lower（ P〈0. 05）.Conclusion There are nerve cells＇ apoptosis and changes of the expressions of MMP-9 and caspase-3 in the rats＇ hippocampus tissue after diffuse axonal injury. TIMP can inhibit the expression of MMP-9 and the activation of caspase-3,enhance the anti-apoptosis function,reduce the nerve cells＇ apoptosis in the rats＇ hippocampus tissue after injury,indicating that TIMP has a certain protection for the rats＇ brain after brain injury.