Objective: To investigate the cerebral protective effect of shRNA down-regulated matrix metalloproteinase-9(MMP-9) on cerebral infarction rats. Methods: 32 SD rats were randomly divided into four groups (8 for each group): group A (sham operation group): the branches of external carotid artery were only separated and ligated and were not be inserted thread; group B (MCAO group): rat MCAO models were made referring to the conventional Longa method; group C (physiological saline group): rats were injected with 5 μl saline two weeks before MCAO models; group D (lentivirus intervention group): rats were injected with 5 μl lentivirus-mediated shRNA two weeks before MCAO models.The diffusion tensor imaging(DTI) was used to evaluate the tissue of cerebral ischemic of middle cerebral artery occlusion(MCAO) in rats.At 4, 8, 12, 18 and 24 h after MCAO, the ADC value and FA value of the lesion side were calculated.The expression of MMP-9 protein in brain tissue of ischemic side was measured by Western blot, and the expression of MMP-9 protein was observed by paraffin section and immunohistochemistry.SPSS 17.0 software was used for statistical processing, and the difference was statistically significant in P<0.05. Results: At 4, 8, 12, 18, 24 h after ischemia, the values of ADC and FA in the ischemic area of the lentivirus intervention group were higher than those in the MCAO group, and the difference was statistically significant (the t values of ADC values at different time points were 8.78, 3.30, 3.58, 4.55 and 13.12, respectively P<0.05.The t values of FA values at different time points were 2.51, 5.87, 6.91, 4.97 and 4.58, respectively P<0.05). The expression of MMP-9 protein increased gradually after focal cerebral ischemia.The expression of MMP-9 protein in lentivirus intervention group was lower than MCAO group.The differences were statistically significant at each point (t values at different time points were 3.55, 6.11, 4.67, 4.63 and 2.77, respectively P<0.05). Conclusion: Lentivirus mediated shRNA inhibits the expression of MMP-9, reduces the vasogenic edema and plays a protective role in cerebral infarction.Magnetic resonance DTI can monitor dynamically the changes of ischemic area in rats with ischemic cerebral infarction.
目的: 通过对大鼠大脑中动脉栓塞(MCAO)模型脑缺血区组织的扩散张量成像(DTI)评价,探讨shRNA下调基质金属蛋白酶9(MMP-9)对脑梗死大鼠的脑保护作用。 方法: 将32只SD大鼠随机分为4组(每组8只):A组(假手术组):仅分离结扎颈外动脉分支,不插入栓线;B组(MCAO组):参照传统的Longa法,建立大鼠MCAO模型;C组(生理盐水组):MCAO两周前给予脑组织内注射生理盐水5 μl;D组(慢病毒干预组):MCAO两周前给予脑组织内注射慢病毒介导的shRNA 5 μl。MCAO后4、8、12、18、24 h时间点,计算脑梗死侧的表观扩散系数(ADC)值和部分各向异性(FA)值。Western印迹测定缺血侧脑组织MMP-9蛋白表达;免疫组化染色观测MMP-9蛋白表达。使用SPSS 17.0软件进行统计学处理,以P<0.05为差异有统计学意义。 结果: 缺血后4、8、12、18、24 h,慢病毒干预组缺血区的ADC值和FA值高于MCAO组,且差异有统计学意义(不同时间点ADC值差异的t值分别为8.78、3.30、3.58、4.55和13.12,P<0.05。不同时间点FA值差异的t值分别为2.51、5.87、6.91、4.97和4.58,P<0.05)。局灶性脑缺血后MMP-9蛋白表达逐渐增多;慢病毒干预组相应时间点的MMP-9蛋白的表达低于MCAO组,且在相应的每个时间点差异均具有统计学意义(不同时间点t值分别为3.55、6.11、4.67、4.63和2.77,均P<0.05)。慢病毒干预组的mNSS评分低于MCAO组,差异有统计学意义(不同时间点t值分别为2.25、2.90、5.19、4.36和7.31,P<0.05)。 结论: 慢病毒介导的shRNA通过抑制MMP-9基因表达,减轻血管源性水肿,发挥脑梗死后的保护作用;磁共振DTI可动态监测缺血性脑梗死大鼠缺血区的改变。.
Keywords: Diffusion tensor imaging; Infarction, middle cerebral artery; Lentivirus; Matrix metalloproteinase 9.