缺血-再灌注损伤诱导大鼠视网膜AQP4内化及其溶酶体降解  

Ischemia-reperfusion injury induces internalization and lysosomal degradation of retinal AQP4 in mice

作　　者：甘胜伟[1] 孙善全[1] 冉建华[1] 陈海[2] 樊萍[3] 朱淑娟[1] 黄娟[1] 陈臻[4] 

机构地区：[1]重庆医科大学神经科学研究中心,重庆400016 [2]重庆市第三人民医院神经内科,重庆400014 [3]重庆市第五人民医院妇产科,重庆400062 [4]云南省第一人民医院眼科,昆明650031

出　　处：《第三军医大学学报》2013年第14期1459-1465,共7页Journal of Third Military Medical University

基　　金：国家自然科学基金(30470608;30500171);重庆市自然科学基金(CSTC2012jjA10022;CSTC2011jjA10003;CSTC2010BB5098)~~

摘　　要：目的研究在急性高眼压作用后不同再灌注时间水通道蛋白4(aquaporin-4,AQP4)内化及其向溶酶体分选(降解)的规律,探讨AQP4的内化及其降解与视网膜胶质细胞水肿的关系。方法取72只体质量190～210 g的雌性SD大鼠(8周龄),在其左眼建立急性高眼压模型,并根据再灌注时间分为0、1、2、4、8、12 h组(n=12),应用免疫荧光双标的方法鉴定视网膜内胶质细胞的分布部位,并检测各组大鼠视网膜AQP4与早期内涵体抗原1(early endosome antigen 1,EEA1)、晚期内涵体标记物甘露糖-6-磷酸受体(mannose-6-phosphate receptor,MPR)及溶酶体相关膜蛋白-1(lysosomal-asso-ciated membrane protein-1,LAMP1)的共表达,分别计数AQP4阳性细胞及AQP4/EEA1、AQP4/LAMP1共表达细胞,计算其比例并进行统计学比较。结果视网膜内星形胶质细胞位于神经纤维层及节细胞层,Müller细胞位于内核层;在高眼压介导的再灌注损伤0～8 h,可见AQP4分别与EEA1、MPR有共表达,再灌注12 h则仅见AQP4与EEA1的共表达;再灌注1～12 h都可见AQP4与LAMP1的共表达,再灌注4 h,胶质细胞中AQP4与LAMP1共表达细胞的比例达到最大值(55.60±13.03)%,与其他时间点相比,差异有统计学意义(P<0.05),此后共表达细胞比例逐渐减少。结论高眼压所致的视网膜缺血-再灌注损伤可诱导AQP4发生内化,内化的AQP4可分选至溶酶体降解。AQP4的内化/降解可能在视网膜水肿的发生、发展中起作用。Objective To investigate the internalization and lysosomal degradation of retinal aquapor- in-4 （AQP4） induced by different reperfusion phases after acute ocular hypertension （AOH） in rats, so as to explore the relationship of AQP4 internalization and degradation with the glial edema of retinae. Methods A total of 72 rats, 8-week-old, and weighting 190 to 210 g, were induced to AOH models in the left eye, and then divided into 6 reperfusion groups （n = 12 for each group） according to the different duration of reperfu- sion, that is, 0, 1, 2, 4, 8 and 12 h following 60 minutes＇ ischemia. Double-immunofluorescence labeling was applied to identify the localization of glial cells and detect the co-expression of AQP4/early endosome anti- gen 1 （ EEA1 ）, AQP4/mannose-6-phosphate receptor （ MPR）, and AQP4/lysosomal-associated membrane protein-1 （LAMP1）. The numbers of single-labeled AQP4＋ cells, and double-labeled AQP4＋/EEA1＋, and AQP4 ＋/LAMP1 ＋ cells were estimated and statistical compared between the consecutive groups. Results The astrocytes which were positive to GFAP were located in the nerve fiber layer and ganglion cell layer, and the Muller ceils positive to GS were located in the inner nuclear layer. AQP4/EEA1 and AQP4/MPR were co-local- ized during an 8-hour reperfusion window following 60 min ischemia, but only AQP4 and EEA1 were stillobserved till 12 h of reperfusion. The co-expression of AQP4 and LAMP1 was seen from 1 to 12 h of reperfu- sion, with increased first and then decreased gradually, At the 4-hour reperfusion, the cells co-expressing AQP4 and LAMP1 reached the largest count, accounting for （55.60 ± 13.03 ） % （ P 〈 0.05, compared with other time points）. Conclusion Ischemia-reperfusion injury in the retina resulted from ocular hypertension in-duces the internalization of retinal AQP4, which are then degraded by lysosome. Internalization and lysosomal degradation of AQP4 may play a role in the pathogenesis and progress of retinal edema.

关 键 词：水通道蛋白-4 胶质细胞 再灌注损伤 内化 内涵体 溶酶体 

分 类 号：R362[医药卫生—病理学] R774.1[医药卫生—基础医学]