敲减水通道蛋白1对小鼠雪旺细胞形态及水转运的影响  

Effects of knocking down AQP1 by RNAi on the morphology and water transport in mouse Schwann cells

作　　者：章杰[1,2] 江华[2] 刘安堂[2] 朱鴷[2] 张文俊[2] 芦立轩[2] 

机构地区：[1]南昌大学第一附属医院整形美容科,江西南昌330006 [2]第二军医大学上海长征医院整形外科,上海200003

出　　处：《基础医学与临床》2014年第1期29-35,共7页Basic and Clinical Medicine

基　　金：国家自然科学基金(31271264)

摘　　要：目的探讨水通道蛋白1(AQP1)基因表达和雪旺细胞肿胀之间的关系。方法细胞免疫荧光双标法验证小鼠面神经来源的原代雪旺细胞AQP1表达;应用慢病毒RNA干扰技术下调雪旺细胞AQP1表达;AQP1-shRNA感染雪旺细胞后,与CTRL组(scr-shRNA)比较,每天在倒置相差显微镜下观察细胞形态变化,连续6d;细胞容积测定量化AQP1-shRNA细胞与CTRL(scr-shRNA)细胞水含量变化;MTT法检测细胞增殖;流式细胞术检测AQP1-shRNA对雪旺细胞凋亡的影响。结果小鼠面神经来源的原代雪旺细胞表达AQP1;构建的小鼠AQP1-shRNA慢病毒载体感染雪旺细胞后,能在mRNA和蛋白水平明显抑制AQP1表达(P<0.05);AQP1-shRNA能使雪旺细胞形态皱缩,细胞水含量明显降低(P<0.01);AQP1-shRNA细胞较CTRL组细胞增殖活力低(P<0.05),但不会引起雪旺细胞明显的凋亡。结论 AQP1是控制雪旺细胞快速水转运的主要因素,抑制AQP1表达可能提供一种临床治疗面神经水肿的新方法。Objective To explore the relationship between the expression of AQP1 gene and Schwann cells swell-ing. Methods The AQP1 expression in Schwann cells of mouse facial nerve tissues was detected by immunofluo-rescent staining. The RNA interference by lentiviral transduction was used to specifically down-regulate AQP1 ex-pression in Schwann cells. AQP1-shRNA and scr-shRNA transduced cells were observed during AQP1-KD by phase contrast microscopy. Cell volume of scr-shRNA and AQP1 shRNA treated cells was measured daily for six days. Cell viability was measured by MTT assay. Apoptosis effects of AQP1 shRNA on Schwann cells were detected by flow cytometry. Results Schwann cell primary cultures maintained a high level of AQP1 water channels,repre-senting an ideal cell model to study the role of AQP1 in the facial nerve. AQP1 expression in AQP1-shRNA cells was significantly lower compared to cells transduced with only the scr-shRNA by Real-time PCR and Western Blot analysis（P 〈0. 05）. AQP1 gene silencing resulted in a cell shrinkage phenotype,as validated by cell volume deter-minations（P 〈0. 01）. MTT assay showed that the cell viability in AQP1-shRNA cells was lower compared to cells transduced with scr-shRNA（P 〈0. 05）. Flow cytometry assay showed that AQP1 gene silencing fail to affect the apoptosis and necrosis rates of Schwann cells. Conclusions AQP1 is an important factor responsible for the fast water transport of cultured Schwann cells; AQP1 inhibition may provide a new therapeutic alternative for the treat-ment of some forms of facial nerve edema.

关 键 词：水通道蛋白 RNA干扰 慢病毒 面瘫 基因治疗 

分 类 号：R363[医药卫生—病理学]