缺血半暗带组织中水通道蛋白4表达的初步研究

背景在组织学和细胞学上对半暗带的认识已取得了进展,而关于水通道蛋白(aquaporin-4,AQP4)与半暗带和磁共振成像相结合的报道不多.目的探讨水通道蛋白-4(aquaporin-4,AQP4)在缺血半暗带(ischemicpenumbra,IP)中的表达规律.设计随机对照的实验研究.地点和对象健康Wistar大鼠35只,体质量300～400g,雌雄不拘,随机分为对照组和栓塞组;栓塞组又分为大脑中动脉栓塞(MCAO)后15,30min,1,3,6,24h组,每组各5只大鼠.干预对照组的大脑中动脉不栓塞,其余操作同栓塞组.在对照组术后30min、各栓塞组于栓塞结束时行脑部T1WI,T2WI和DWI扫描.采用刘梅丽等方法界定影像半暗带,计算影像半暗带和中心梗死区的相对表观扩散系数(ADC1和ADC2),将缺血脑组织进行红四氮唑(TTC)染色,并与DWI图像对比.取最大层面的影像半暗带组织进行病理观察、免疫组织化学及原位杂交实验,用图像分析检测AQP4蛋白、基因的表达量,并进行统计分析.主要观察指标①影像半暗带组织病理观察结果.②影像半暗带和中心梗死区的相对表现扩散系数.③AQP4蛋白、基因的表达量.结果对照组的DWI和T2WI未见异常,ADCR为(98.7±0.8)%,AQP4蛋白表达平均值为(7.9±0.5)%,基因表达平均值为0.5±0.06.栓塞后15min即在DWI上出现高信号;平均于栓塞后1.4h在T2WI显示高信号,ADC1在24 h内始终呈下降趋势,在1 h内迅速下降;而ADC2呈现先降后升的变化规律.在1h内AQP4表达迅速上调,1～24 h仍缓慢增加,AQP4表达与ADC1呈显著负相关(r=-0.989,P＜0.001).影像半暗带与组织半暗带都具有相同的病理改变-细胞性水肿.结论影像半暗带与组织半暗带是等值的,半暗带的病理基础之一是细胞内水肿,AQP4表达增强是缺血半暗带形成的重要原因,半暗带组织的ADC值下降可以间接反映AQP4表达上调的水平.

Expression of aquaporin-4 in ischemic penumbra tissues

BACKGROUND:The histological and cytological understanding of penumbra has received progresses. However, there are not any reports regarding the combination of aquaporin-4(AQP4) and penumbra and Magnetic Resonance Imaging.OBJECTIVE: To investigate the expressive rule of AQP4 in the graphic ischemic penumbra(IP) tissues.DESIGN: A randomised controlled trial.SETTING and MATERIALS: Thirty-five healthy Wistar rats with a body mass from 300 g to 400 g in either gender were randomly divided into control and occlusion group by computerized random method. The occlusion group was again divided into 15 minutes, 30 minutes, 1 hour, 3 hours, 6hours and 24 hours after middle cerebral artery occlusion(MCAO)sub-groups with 5 rats each.INTERVENTION: The operations performed in rats of control group were as same as that of occlusion group except MCAO. Brain T1 WI, T2WI and DWI scanning was performed at 30 minutes after operation in rats of control group,and immediately after occlusion in each occlusion sub-groups. Penumbra was determined with the method reported by Mei-Li Liu et al for the calculation of the relative apparent diffusion coefficient(ADC) of penumbra(ADC1) and central infarct area(ADC2) . The ischemic brain tissue was stained by TTC,which was compared with DWI images as well. The brain tissue with the largest penumbral area was used for pathological observation, immunohistochemistry(△S) and in situ hybridization (α) tests.The expressions of AQP4protein and gene were assayed by image analysis, which were also analyzed statistically.MAIN OUTCOME MEASURES: ① results of histopathological observation of penumbral tissues. ② ADC1 and ADC. ③ the expressions of AQP4protein and gene.RESULTS: There were no significant changes on DWI and T2WI and the mean values of ADCR, △S and o were(98.7 ±0. 8) ％, (7.9 ±0. 5)％ and (0.5 ±0. 06) respectively in rats of control group. Abnormal high-intensity was found on DWI at 15 minutes after MCAO. T2WI detected high-intensity at 1 hour(average time) after MCAO. ADC1 decreased quickly within 1 hour after MCAO followed. By a descending tendency during 24 hours inpenumbra. Whereas, the changes in theinfarct tissue, the ADC2 had a rule of decreasing followed by increasing. The expression of AQP4 increased rapidly within I hour after MCAO followed by a slow increase within 24 hours.The expression of AQP4 negatively correlated witl ADC1 in the penumbra (r= -0.989, P < 0.001) . Same major pathologic change in both penumbra and histological penumbra, which was intracellular edema.CONCLUSION: The penumbra is as same as histological penumbra with the pathologic characteristics of intracellular edema. The enhanced expression of AQP4 may be an important cause of IP. The decrease of ADC values in IP may indirectly reflect the up-regulation of AQP4 expression.