C/EBPβ介导NPHP1敲低的肾小管上皮细胞TNF-α通路下游炎症因子的表达  

C/EBPβmediates expressions of downstream inflammatory factors of the tumor necrosis factor-αsignaling pathway in renal tubular epithelial cells with NPHP1 knockdown

作　　者：黄丹梅 刘雅清 李丹彤 张静兰 杨翌晨 孙良忠 HUANG Danmei;LIU Yaqing;LI Dantong;ZHANG Jingan;YANG Yichen;SUN Liangzhong(Department of Pediatrics,Nanfang Hospital,Southern Medical University,Guangzhou 510515,China)

机构地区：[1]南方医科大学南方医院儿科,广东广州510515

出　　处：《南方医科大学学报》2024年第1期156-165,共10页Journal of Southern Medical University

基　　金：广东省自然科学基金(2022A1515012307;2020A1515010286);北京白求恩公益基金会(SCE093DS);南方医科大学南方医院院长基金青年项目(2020C021)。

摘　　要：目的探讨NPHP1基因缺陷肾小管上皮细胞TNF-α信号通路激活及其调控的炎症因子表达。方法使用重组慢病毒LV-NPHP1-RNAi构建敲低NPHP1表达的人近端肾小管细胞株(HK2)细胞(NPHP1^(KD)HK2)。通过实时荧光定量PCR、Western blot、酶联免疫吸附实验法检测各细胞株TNF-α表达、p38和C/EBPβ激活状态及其调控炎症因子CXCL5、CCL20、IL-1β、IL-6、MCP-1等表达情况。通过构建siRNA敲低野生型和NPHP1^(KD)HK2细胞C/EBPβ表达,观察上述指标变化。结果敲低NPHP1表达后,NPHP1^(KD)HK2细胞TNF-α、C/EBPβ、CXCL5、IL-1β和IL-6的mRNA表达量增加(P<0.05);Western blotting结果显示,phospho-p38、C/EBPβ表达上调(P<0.05);培养液上清IL-6水平增加(P<0.05)。使用siRNA敲低C/EBPβ表达后,NPHP1KDHK2细胞CSF2、CCL20、IL-1β和IL-6的mRNA表达水平下调(P<0.05);Western blot显示phospho-p38表达下调(P<0.05);培养液上清IL-6水平下降(P<0.001)。结论NPHP1基因缺陷的NPHP1KDHK2细胞中TNF-α信号通路被激活,其调控的下游印证因子表达上调。C/EBPβ可能是介导NPHP1KDHK2细胞TNF-α信号通路相关炎症因子表达的关键转录因子。Objective To explore the activation of tumor necrosis factor-α(TNF-α)signaling pathway and the expressions of the associated inflammatory factors in NPHP1-defective renal tubular epithelial cells.Methods A human proximal renal tubular cell(HK2)model of lentivirus-mediated NPHP1 knockdown(NPHP1^(KD))was constructed,and the expressions of TNF-α,p38,and C/EBPβand the inflammatory factors CXCL5,CCL20,IL-1β,IL-6 and MCP-1 were detected using RT-qPCR,Western blotting or enzyme-linked immunosorbent assay.A small interfering RNA(siRNA)was transfected in wild-type and NPHP1^(KD)HK2 cells,and the changes in the expressions of TNF-α,p38,and C/EBPβand the inflammatory factors were examined.Results NPHP1^(KD)HK2 cells showed significantly increased mRNA expressions of TNF-α,C/EBPβ,CXCL5,IL-1β,and IL-6(P<0.05),protein expressions of phospho-p38 and C/EBPβ(P<0.05),and IL-6 level in the culture supernatant(P<0.05),and these changes were significantly blocked by transfection of cells with siRNA-C/EBPβ(P<0.05).Conclusion TNF-αsignaling pathway is activated and its associated inflammatory factors are upregulated in NPHP1^(KD)HK2 cells,and C/EBPβmay serve as a key transcription factor to mediate these changes.

关 键 词：肾单位肾痨 TNF-Α C/EBPΒ 炎症因子 NPHP1 

分 类 号：R692[医药卫生—泌尿科学]