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IgA肾病中尿微小RNA作为生物标志物的分析及初步验证

Profiling and initial validation of urinary microRNAs as biomarkers in IgA nephropathy.

作者信息

Wang Nannan, Bu Ru, Duan Zhiyu, Zhang Xueguang, Chen Pu, Li Zuoxiang, Wu Jie, Cai Guangyan, Chen Xiangmei

机构信息

Department of Nephrology, Chinese PLA General Hospital, Chinese PLA Institute of Nephrology, State Key Laboratory of Kidney Diseases, National Clinical Research Center for Kidney Diseases , Beijing , China.

出版信息

PeerJ. 2015 Jun 2;3:e990. doi: 10.7717/peerj.990. eCollection 2015.

Abstract

Background. MicroRNAs (miRNAs) have been found in virtually all body fluids and used successfully as biomarkers for various diseases. Evidence indicates that miRNAs have important roles in IgA nephropathy (IgAN), a major cause of renal failure. In this study, we looked for differentially expressed miRNAs in IgAN and further evaluated the correlations between candidate miRNAs and the severity of IgAN. Methods. Microarray and RT-qRCR (real-time quantitative polymerase chain reaction) were sequentially used to screen and further verify miRNA expression profiles in urinary sediments of IgAN patients in two independent cohorts. The screening cohort consisted of 32 urine samples from 18 patients with IgAN, 4 patients with MN (membranous nephropathy), 4 patients with MCD (minimal changes disease) and 6 healthy subjects; the validation cohort consisted of 102 IgAN patients, 41 MN patients, 27 MCD patients and 34 healthy subjects. The renal pathological lesions of patients with IgAN were evaluated according to Lee's grading system and Oxford classification. Results. At the screening phase, significance analysis of microarrays analysis showed that no miRNA was differentially expressed in the IgAN group compared to all control groups. But IgAN grade I-II and III subgroups (according to Lee's grading system) shared dysregulation of two miRNAs (miR-3613-3p and miR-4668-5p). At the validation phase, RT-qPCR results showed that urinary level of miR-3613-3p was significantly lower in IgAN than that in MN, MCD and healthy controls (0.47, 0.44 and 0.24 folds, respectively, all P < 0.01 by Mann-Whitney U test); urinary level of miR-4668-5p was also significantly lower in IgAN than that in healthy controls (0.49 fold, P < 0.01). Significant correlations were found between urinary levels of miR-3613-3p with 24-hour urinary protein excretion (Spearman r = 0.50, P = 0.034), eGFR (estimated glomerular filtration rate) (r = - 0.48, P = 0.043) and Lee's grades (r = 0.57, P = 0.014). Similarly, miR-4668-5p was significantly correlated with eGFR (r = - 0.50, P = 0.034) and Lee's grades (r = 0.57, P = 0.013). For segmental glomerulosclerosis according to Oxford classification, patients scored as S0 had significantly lower levels of urinary miR-3613-3p and miR-4668-5p than those scored as S1 (0.41 and 0.43 folds, respectively, all P < 0.05). Conclusions. The expression profile of miRNAs was significantly altered in urinary sediments from patients with IgAN. Urinary expression of miR-3613-3p was down-regulated in patients with IgAN. Moreover, urinary levels of both miR-3613-3p and miR-4668-5p were correlated with disease severity. Further studies are needed to explore the roles of miR-3613-3p and miR-4668-5p in the pathogenesis and progression of IgA nephropathy.

摘要

背景。微小RNA(miRNA)几乎在所有体液中都有发现,并已成功用作各种疾病的生物标志物。有证据表明,miRNA在IgA肾病(IgAN)中起重要作用,IgA肾病是肾衰竭的主要原因。在本研究中,我们寻找IgAN中差异表达的miRNA,并进一步评估候选miRNA与IgAN严重程度之间的相关性。方法。依次使用微阵列和RT-qRCR(实时定量聚合酶链反应)来筛选并进一步验证两个独立队列中IgAN患者尿沉渣中的miRNA表达谱。筛选队列包括来自18例IgAN患者、4例膜性肾病(MN)患者、4例微小病变肾病(MCD)患者和6名健康受试者的32份尿液样本;验证队列包括102例IgAN患者、41例MN患者、27例MCD患者和34名健康受试者。根据Lee分级系统和牛津分类法评估IgAN患者的肾脏病理损伤。结果。在筛选阶段,微阵列分析的显著性分析表明,与所有对照组相比,IgAN组中没有miRNA差异表达。但IgAN I-II级和III级亚组(根据Lee分级系统)共有两种miRNA(miR-3613-3p和miR-4668-5p)失调。在验证阶段,RT-qPCR结果显示,IgAN患者尿液中miR-3613-3p水平显著低于MN、MCD和健康对照组(分别为0.47、0.44和0.24倍,通过Mann-Whitney U检验,均P<0.01);IgAN患者尿液中miR-4668-5p水平也显著低于健康对照组(0.49倍,P<0.01)。发现miR-3613-3p尿液水平与24小时尿蛋白排泄量(Spearman r = 0.50,P = 0.034)、估算肾小球滤过率(eGFR)(r = -0.48,P = 0.043)和Lee分级(r = 0.57,P = 0. = 0.014)之间存在显著相关性。同样,miR-4668-5p与eGFR(r = -0.50,P = 0.034)和Lee分级(r = 0.57,P = 0.013)显著相关。根据牛津分类法的节段性肾小球硬化,评分为S0的患者尿液中miR-3613-3p和miR-4668-5p水平显著低于评分为S1的患者(分别为0.41和0.43倍,均P<0.05)。结论。IgAN患者尿沉渣中miRNA的表达谱发生了显著改变。IgAN患者尿液中miR-3613-3p的表达下调。此外,miR-3613-3p和miR-4668-5p的尿液水平均与疾病严重程度相关。需要进一步研究来探索miR-3613-3p和miR-4668-5p在IgA肾病发病机制和进展中的作用。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b3e3/4458130/430386c25af7/peerj-03-990-g001.jpg

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