来氟米特通过调节ESRP1的水平促进肺组织纤维化的机制研究  

Mechanism of Leflunomide Promoting Pulmonary Fibrosis by Regulating ESRP1 Level

作　　者：刘冬 赖伟男[2] LIU Dong;LAI Wei-nan(Department of Pharmacy,Xinqiao Hospital,Army Military Medical University,Chongqing 400037,China;Department of Rheumatology and Immunology,Nanfang Hospital,Southern Medical University,Guangzhou 510515,China)

机构地区：[1]陆军军医大学新桥医院药剂科,重庆400037 [2]南方医科大学南方医院风湿免疫科,广州510515

出　　处：《解放军医药杂志》2019年第11期11-14,共4页Medical & Pharmaceutical Journal of Chinese People’s Liberation Army

基　　金：广东省自然科学基金资助项目(2017A030313508)

摘　　要：目的探究来氟米特(LEF)通过调节上皮细胞剪接调节蛋白(ESRP1)的水平促进肺组织纤维化的机制。方法将人肺上皮细胞BEAS-2B随机分为对照组、LEF组、ESRP1+LEF组和ESRP1组。使用细胞转染技术上调ESRP1的水平,在5 mg/L的LEF条件下培养48 h。使用免疫荧光染色检测α平滑肌肌动蛋白(α-SMA)。分别使用Western blot和RT-qPCR检测蛋白和mRNA的水平。结果 LEF组ESRP1的mRNA和蛋白的水平显著低于对照组,且ESRP1+LEF组的ESRP1的表达水平显著低于LEF组(P<0.05)。各组细胞的活力比较差异物统计学意义(P>0.05)。对照组α-SMA表达水平很低,并且可看出细胞轮廓为椭圆形。LEF组的α-SMA的荧光强度高于对照组,ESRP1组的α-SMA的荧光强度低于对照组,ESRP1+LEF组的α-SMA相对荧光强度高于ESRP1组(P<0.05)。与对照组比较,LEF组α-SMA水平升高,E-cadherin水平降低(P<0.05)。ESRP1组的α-SMA低于对照组,E-cadherin高于对照组(P<0.05)。与ESRP1组比较,ESRP1+LEF组α-SMA水平升高,E-cadherin水平降低(P<0.05)。结论 LEF可能通过抑制ESRP1的水平,促进α-SMA的表达而抑制E-cadherin的水平,促进肺上皮细胞向间质细胞转化,引起肺纤维化。Objective To investigate mechanism of Leflunomide(LEF) promoting fibrosis in lung tissue by regulating levels of epithelial splicing regulatory proteins1(ESRP1). Methods Human lung epithelial cells BEAS-2 B were randomly divided into control group, LEF group, ESRP1+LEF group and ESRP1 group. ESRP1 level was up-regulated using cell transfection technique and cultured for 48 h at 5 mg/L LEF. Level of α-smooth muscle actin(α-SMA) was detected by using immunofluorescence staining. Protein and mRNA levels were detected by using Western blot and real time-quantitative polumerase chain reaction(RT-qPCR) respectively. Results Levels of ESRP1 mRNA and protein in LEF group were significantly lower than those in ESRP1+LEF group, and levels of ESRP1 mRNA and protein in ESRP1+LEF group were significantly lower than those in ESRP1 and LEF groups(P<0.05). There were no significant differences in viability of cells among four group(P>0.05). The α-SMA expression in control group was very low, and cell outline could be found elliptical. Fluorescence intensity of α-SMA in LEF group was more than that in control group, and fluorescence intensity of α-SMA in ESRP1 group was less than that in control group, while relative fluorescence intensity of α-SMA in ESRP1+LEF group was more than that in ESRP1 group(P<0.05). In LEF group, E-cadherin expression was inhibited by promoting α-SMA level. In ESRP1 group, α-SMA level was lower, while E-cadherin was higher than those in control group(P<0.05), and partial reverse LEF had role of promoting α-SMA expression and inhibiting E-cadherin expression. Conclusion LEF may inhibit E-cadherin expression and promote transformation of lung epithelial cells to stromal cells by inhibiting ESRP1 level and promoting α-SMA expression to induce pulmonary fibrosis.

关 键 词：肺纤维化 来氟米特 上皮细胞剪接调节蛋白 上皮-间充质转化 Α平滑肌肌动蛋白 

分 类 号：R563.13[医药卫生—呼吸系统]