环状RNA circHIPK3在儿童哮喘中的表达和作用研究

doi:10.3969/j.issn.1673-5501.2021.01.006

摘要/Abstract

摘要： 背景环状RNA在多种疾病中发挥重要作用,但目前尚无环状RNA在儿童哮喘中的研究报道。有报道环状RNA circHIPK3在过敏性鼻炎中调控Th2细胞的分化。目的探讨环状RNA circHIPK3在儿童哮喘发病中可能的作用机制。设计收集临床样本,检测circHIPK3的表达并检测其与T细胞转录因子的相关性。构建小鼠哮喘模型进一步验证。进行相关的生物信息学分析,并分析circHIPK3与儿童哮喘相关临床指标的相关性。方法纳入2017年8月26日至2019年8月5日复旦大学附属儿科医院呼吸科门诊确诊的哮喘患儿(哮喘组),以同期体检的健康儿童作为对照组。分离两组外周血单个核细胞(PBMCs),通过qRT-PCR检测circHIPK3和T盒子转录因子(T-bet)在两组的表达情况,并进行相关性分析。建立蟑螂提取物(CRE)诱导的小鼠哮喘模型,用qRT-PCR检测小鼠肺组织中的circHIPK3、T-bet的表达情况。通过生物信息学分析circHIPK3与T-bet之间的调控关系。分析circHIPK3与外周血嗜酸性粒细胞的相关性。主要结局指标 circHIPK3和T-bet的相对表达量以及二者的相关系数。结果哮喘组43例,均为轻度哮喘,男35例,中位年龄6.5岁。对照组28例,男22例,中位年龄6.4岁。与对照组相比,circHIPK3(t=4.627,P<0.001)和T-bet (t=2.727, P<0.01)在哮喘组PBMCs中均呈低表达;在哮喘患儿中circHIPK3与T-bet呈正相关(R=0.483 0,P=0.000 7)。哮喘小鼠肺组织中的检测结果与之一致。检索GEO数据库发现,哮喘患儿miR-485-3p增高。生物信息学分析发现:circHIPK3与miR-485-3p存在竞争结合关系,T-bet是miR-485-3p的可能靶基因。哮喘患儿中,circHIPK3与外周血中嗜酸性粒细胞数量呈负相关(R=-0.369 2,P=0.019 1)。结论 circHIPK3在儿童哮喘和哮喘模型小鼠中表达下调,可能通过吸附miR-485-3p的方式影响T-bet的表达,引起Th1/Th2失衡,影响哮喘的炎症表现。

关键词: 环状RNA, 儿童哮喘, T细胞分化

Abstract: Background CircRNAs play an important part in various dieases. But circRNAs in pediatric asthma have not been reported so far. It had been reported that circHIPK3 promoted Th2 differentiation in allergic rhinitis. Objective To explore the expression and potential function of circHIPK3 in children with asthma. Design The study developed by firstly collecting peripheral blood mononuclear cells (PBMCs) from asthmatic patients and healthy controls. The expression of circHIPK3 was detected and the correlations between circHIPK3 and T cell's transcription factors were analyzed. Relative results were validated in asthma mice. Sequently, bioinformatics analysis was used to analyze possible mechanisms and correlation analysis was performed between circHIPK3 and clinical indicators. Methods Peripheral blood mononuclear cells (PBMCs) were collected from asthmatic patients and healthy controls from August 26,2017 to August 5,2019 in the outpatient respirology clinic at Children's Hospital of Fudan University.qRT-PCR was used for the detection of the expression of circHIPK3 and T-bet in the PBMCs.Pearson correlation analysis was used to analyze the correlation between circHIPK3 and T-bet.A cockroach extract (CRE)-induced asthma model was adopted in the experiment. The expression of circHIPK3 and T-bet in the mice lung were detected by qRT-PCR. Bioinformatics analysis was used to explore the potential mechanism between circHIPK3 and T-bet in children with asthma. Pearson correlation analysis was used to analyze the correlation between circHIPK3 and eosinophils in peripheral blood. Main outcome measures The main outcome measures were relative expression of circHIPK3 and T-bet, as well as the correlation coefficients between them. Results A total of 43 patients of mild asthma (35 males, median age 6.5 years) and 28 normal controls (22 males, median age 6.4 years) were evaluated. Comparing with healthy controls,the expression of circHIPK3 (t=4.627,P<0.001) and T-bet (t=2.727, P<0.01) were significantly decreased in asthmatic patients.circHIPK3 was positively correlated with T-bet in pediatric asthma patients (R=0.483 0, P=0.000 7). The results in lung remodeling in asthma mice were in accordance with those in human PBMCs. miR-485-3p was up-regulated in children with asthma by analysis of a reported GEO database. circHIPK3 could bind with miR-485-3p in a sponge way. T-bet was a predicted target of miR-485-3p. A negative correlation was detected between circHIPK3 and eosinophils in peripheral blood (R=-0.369 2, P=0.019 1). Conclusion circHIPK3 was down-regulated in the asthmatic patients and mice, potentially regulating T-bet through sponge miR-485-3p.

Key words: Circular RNAs, Asthma, T cell differentiation

梁秋妍, 郑红梅, 付劲蓉, 刘丽娟, 张晓波, 钱莉玲, 周玉峰. 环状RNA circHIPK3在儿童哮喘中的表达和作用研究[J]. 中国循证儿科杂志, 2021, 16(1): 66-70.

LIANG Qiuyan, ZHENG Hongmei, FU Jinrong, LIU Lijuan, ZHANG Xiaobo, QIAN Liling, ZHOU Yufeng. The expression and function analysis of circHIPK3 in children with asthma[J]. Chinese Journal of Evidence-Based Pediatrics, 2021, 16(1): 66-70.

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