Human antigen R (HuR) and Cold inducible RNA‐binding protein (CIRP) influence intestinal mucosal barrier function in ulcerative colitis by competitive regulation on Claudin1
Human antigen R (HuR) and Cold inducible RNA‐binding protein (CIRP) influence intestinal mucosal barrier function in ulcerative colitis by competitive regulation on Claudin1
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DOI：
10.1002/biof.1719
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发表时间：
2021-02
期刊：
影响因子：
6
通讯作者：
Yan Xu;Yuxi Tian;Ying Wang;Junwen Yang;Fujun Li;Xiaoping Wan;Ouyang Miao
中科院分区：
文献类型：
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作者：
Yan Xu;Yuxi Tian;Ying Wang;Junwen Yang;Fujun Li;Xiaoping Wan;Ouyang Miao
目的：探讨冷诱导RNA结合蛋白(CIRP)和人源抗原R(HUR)在溃疡性结肠炎(UC)中对Claudin1表达和粘膜屏障功能的影响。收集UC患者和健康志愿者的临床标本。在临床实验中，分别用qRT-PCR、Western印迹和Pearson相关系数分析了CIRP、Claudin1和Hur在UC患者组织中的表达及其相关性。采用卡方检验评估CIRP/HUR/Claudin1水平与UC患者临床病理特征的相关性。分别采用脂多糖治疗和葡聚糖硫酸钠注射法建立UC的体内外模型。细胞实验采用流式细胞仪和CCK-8比色法检测失活和失活后CIRP或HUR对肠上皮细胞凋亡和增殖的影响。测定跨上皮电阻值、辣根过氧化物酶通透性和紧密连接蛋白(occludin、ZO-1、JAM-1)的表达，检测肠上皮屏障功能。用RNA免疫沉淀和双荧光素酶报告基因检测HUR、CIRP和Claudin1之间的关系。体内实验通过对疾病活动性指数评分、体重减轻和结肠长度的评估来观察CIRP或HUR对UC小鼠模型的影响。采用HE染色对结肠组织进行组织学分析。FITC-葡聚糖示踪法检测UC模型小鼠肠黏膜屏障功能。在本研究中，在UC患者、细胞和小鼠模型中，CIRP高表达，Hur和Claudin1低表达。CIRP、HUR和Claudin1的表达与UC的严重程度相关。CIRP与Claudin1呈负相关，Hur与Claudin1呈正相关。Claudin1可被CIRP抑制，而被HUR增强。HUR和CIRP可以竞争性地与Claudin1结合。HUR上调或CIRP下调可促进肠上皮细胞增殖，抑制细胞凋亡，改善肠细胞屏障功能受损。体内实验证明，Hur过表达或CIRP基因敲除可改善UC小鼠肠黏膜屏障功能的损害。CIRP和HUR通过竞争性结合Claudin1mRNA，在UC的肠粘膜屏障功能中发挥关键作用。
To investigate the effects of RNA‐binding proteins cold‐inducible RNA binding protein (CIRP) and human antigen R (HuR) on expression of Claudin1 and mucosal barrier function in ulcerative colitis (UC). The clinical specimens of UC patients and healthy volunteers were collected. In the clinical experiments, the expressions of CIRP, Claudin1, and HuR, along with their correlations in tissues of UC patients were analyzed by qRT‐PCR, Western blot and Pearson correlation coefficient, respectively. The chi‐square test was utilized to assess the relevance between CIRP/HuR/Claudin1 level and clinicopathological characteristics of UC patients. The in vitro and in vivo models of UC were established by lipopolysaccharide treatment or dextran sulfate sodium injection. For cell experiments, after loss‐ and gain‐of‐function, the roles of CIRP or HuR in the apoptosis and proliferation of enterocytes were examined by flow cytometry and CCK‐8 assay. The intestinal epithelial barrier function was inspected after determination on transepithelial electrical resistance value, horseradish peroxidase permeability and expressions of tight junction proteins (Occludin, ZO‐1, and JAM‐1). The relationship between HuR, CIRP, and Claudin1 was performed by RNA immunoprecipitation and dual‐luciferase reporter gene assay. For in vivo experiments, the disease activity index score, weight loss and colon length of mice were assessed to observe the effect of CIRP or HuR on the UC mouse models. Histological analysis of colon tissues was conducted by H&E staining. FITC‐dextran tracking was applied to inspect the intestinal mucosal barrier function of UC mouse models. In this study, high expression of CIRP and low expressions of HuR and Claudin1 were observed in patients, cells and mouse models of UC. The expressions of CIRP, HuR, and Claudin1 were correlated with the severity of patients with UC. There was a negative correlation between CIRP and Claudin1, and as a positive correlation between HuR and Claudin1. Claudin1 can be suppressed by CIRP, while enhanced by HuR. HuR and CIRP can competitively bind to Claudin1. HuR upregulation or CIRP downregulation promoted proliferation, suppressed apoptosis and ameliorated the damage of the barrier function in enterocytes. The in vivo experiments verified that the ameliorated damage of the intestinal mucosal barrier function in UC mice occurred with HuR overexpression or CIRP knockdown. CIRP and HuR confer pivotal effect on the intestinal mucosal barrier function of UC through competitively binding to Claudin1 mRNA.