Association Between MiRNAs Expression And IgA Nephropathy Progression

BackgroundImmunoglobulin A nephropathy (IgAN) is the most common form of primaryglomerulonephritis throughout the world. Meanwhile, it is also an important cause ofend stage renal disease (ESRD), and about40%of patients with biopsy-proven IgANprogress to ESRD in10to20years. It has been well known that renal biopsy isconsidered as the gold standard for diagnosis of IgAN. However, renal biopsy haspotential complications that cannot be easily accepted by most patients, and repeatedmonitoring is technically difficult. Consequently, highly sensitive and specificnon-invasive biomarkers reflecting disease severity and progression are urgentlyneeded in the clinical management of patients with IgAN. MiRNAs are a group ofshort noncoding RNA molecules that regulate gene express at post-transcriptinal levelby degrading or repressing translation of target mRNA. And abnormal expression ofmiRNAs may lead to many kinds of diseases including IgAN by changing kidneytissue structure, immune molecular structure, cell signaling etc. Moreover, miRNAs,present in body fluids, such as plasma and urine, are in a remarkably stable form thateven withstands repetitive freezing/thawing cycles and are protected against RNAses.They can display unique expression profiles in pathological conditions and it has be proposed that distinctive miRNA signatures may be exploited as innovativediagnostic/prognostic tools. However, there has been no certain conclusion about theassociation between expression of miRNAs and progression of IgAN until now. Inthe present study, we chose specific miRNAs targets, based on their reportedinvolvement in pathogensis and renal fibrosis in IgAN, exploring the possibility ofusing these miRNAs as non-invasive biomarkers for it.ObjectiveTo study intra-renal, plasmic and urinary levels of miR-148b, miR-194,miR-200a and miR-382in patients with IgAN, and to explore the association betweenmiRNAs expression and IgA nephropathy progression, investigating the possibility ofusing these miRNAs as non-invasive biomarkers for this disease.Methods1. We used real-time PCR to quantify the expression of miR-148b, miR-194,miR-200a and miR-382in kidney biopsy of7patients with biopsy-proven IgAN,plasma and urine of34patients with biopsy-proven IgAN. We studied normal renaltissue from the nephrectomy specimen of7patients with renal cell carcinoma, andplasma and urine from13healthy volunteers as controls for intra-renal, plasmic andurinary expression study, respectively.2. Total RNA was extracted from kidney tissue, plasma and urine using TRIzolLS Reagent(Invitrogen life technologies).The concentration of all total RNA sampleswere measured by NanoDrop ND-1000(Nanodrop, Wilmington, Delaware,USA).RNU6B was used as house-keeping genes to normalize the miRNA expression.The2-△△Ctmethod for relative quantitation was used.3. Data ananysis of miR-148b, miR-194, miR-200a and miR-382expression inkidney tissue, plasma and urine of patients with IgAN and in healthy controls, and thecorrelation between miRNAs expression and clinical parameters, including serumcreatinine,24hours urine protein, glomerular filtration rate(GFR) and histologicalparameters, etc., were performed using SPSS v.19.0software (IBM Corp., Armonk, NY, USA). Date were compared by test or One-way ANOVA. A Bonferronicorrection was applied to adjust the level of significance for multiple comparisons; a“p” value of less than0.017(0.05/3) was considered statistically significant. Pearsoncorrelation or Spearman rank correlation analysis were used to quantify linearcorrelation. A P value of below0.05was considered statistically significant.Results(1) The levels of intra-renal miR-148b、 miR-194、 miR-200a weredown-regulated in patients with IgAN while miR-382was up-regulated (P＜0.05).The levels of plasmic miR-148b、miR-194、miR-382were down-regulated inpatients with IgAN while miR-200a was up-regulated (P＜0.05). The levels ofurinary miR-148b、miR-382were up-regulated while miR-194、miR-200a weredown-regulated (P＜0.05).(2) The levels of plasmic miR-148b、miR-194inversely correlated with Hassgrade (P＜0.017), and the levels of plasmic miR-382were significantly lower in Hassgrade III and grade IV-V than those in grade I-II(P＜0.017). Plasmic levels ofmiR-200a did not correlate with Hass grade(P＞0.05).(3) The levels of urinary miR-148b were significantly higher in Hass grade IV-Vthan those in grade III(P＜0.017). The levels of urinary miR-194were significantlyhigher in Hass grade I-II and grade III than those in grade IV-V(P＜0.017). Urinarylevels of miR-200a and miR-382did not correlate with Hass grade(P＞0.05).(4) Urinary expression of miR-148b negatively significantly correlated withSerum uric acid (UA) and complement4(C4)(P＜0.05), and urinary expression ofmiR-382positively significantly correlated with urine red blood cell counts (P＜0.05).Urinary expression of miR-194、miR-200a and plasmic expression of all fourmiRNAs did not correlate with serum creatinine(Scr), glomerular filtration rate(GFR),blood urea nitrogen(BUN), etc.(P＞0.05).(5) The levels of urinary miR-200a in IgAN patients with T0of Oxfordclassification of IgAN were higher than those in T1-2(P＜0.05). There was nosignificant difference in levels of urinary miR-148b、miR-194、miR-382and plasm- -ic four miRNAs among sub-types of Oxford classification of IgAN(P＞0.05).ConclusionsWe determined in the present study that the intra-renal, plasmic and urinaryexpressions of miR-148b, miR-194, miR-200a and miR-382in patients with IgANwere significantly different with healthy controls, and some of these miRNAsexpressions correlated with histological parameters and clinical paremeters. Theresults suggested that these miRNA species might correlate with progression of IgAN,and urinary expression of miRNAs has the potential of further development asnon-invasive biomarkers of IgAN.