MicroRNA expression in the affected skin of adult patients with IgA vasculitis

Abstract

IgA vasculitis (IgAV) represents a common systemic vasculitis in pediatric and adult population. Our current knowledge of disease pathogenesis is still very limited, without information on miRNAs in IgAV. The aim of our study was to determine the expression of five pre-selected miRNAs (miRNA-146a-5p, miRNA-148-3p, miRNA-155-5p, miRNA-223-3p, and let-7b) in the affected skin of adult IgAV patients. The study included 65 skin samples from consecutive, untreated IgAV patients (61.5% male, median age 67.6 years, range 29-91), diagnosed between October 2014 and September 2016, and 20 samples of normal skin from healthy volunteers. Total RNA was isolated from tissue sections of formalin-fixed, paraffin-embedded samples. Expression of miRNAs was measured using qRT-PCR. To present relative miRNA expression, the ΔΔCT method was used. Skin miRNA expression was correlated to clinical characteristics of adult IgAV patients. We found significantly higher levels of miRNA-155-5p, miRNA-223-3p, and let-7b in the affected skin compared to controls (18.6-fold, 6.4-fold, and 7.9-fold higher respectively). Contrary, the miRNA 148-3p expression was significantly lower (2.2-fold). The expression of the miRNA-146-5p showed near normal levels. Patients with necrotic skin lesions had significantly higher miRNA-223 tissue expression than those with non-necrotic purpura (p = 0.029). Gastrointestinal tract involvement inversely correlated with the expression of miRNA-155-5p and/or miRNA-146a-5p in affected skin. Altered expression of miRNA-148b-3p, miRNA-155-5p, miRNA-223-3p, and let-7b was found in vasculitic skin lesions in IgAV. Additionally, we found a positive association between the severity of purpura and skin miRNA-223-3p expression. Aberrantly expressed miRNAs could represent a biomarker in adult IgAV.

Association between histopathological changes and expression of selected microRNAs in skin of adult patients with IgA vasculitis

“…have recently found that levels of these cytokines are significantly higher in the blood of adult IgAV patients than in the blood of healthy blood donors, which raises the possibility that some of the miRNAs influencing the NF‐κB pathway could be used as biomarkers for IgAV. Decreased expression of miR‐146a‐5p correlated with GIT involvement in IgAV patients . The low expression levels of miR‐146a‐5p found in our present study thus seem to be related to more severe inflammation, which is not necessarily limited to the skin.…”

“…The quality and concentration of isolated RNA were assessed with aNanoDrop 1000 (Thermo Fisher Scientific, Waltham, MA, USA). Subsequent reverse transcription of isolated total RNA was performed with the miScript II RT Kit (218161; Qiagen), as described previously …”

“…All qRT‐PCR experiments were prepared with the miScript SYBR Green PCR Kit (218073; Qiagen) along with miScript Primer Assays (Qiagen), andwere performed on the Rotor‐Gene Q real‐time PCR cycler (Qiagen). Detailed information on qRT‐PCR analysis is given elsewhere . The miScript miRNA primers included in the study are shown in Table .…”

“…Subsequent reverse transcription of isolated total RNA was performed with the miScript II RT Kit (218161; Qiagen), as described previously. 16 Quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR) analysis All qRT-PCR experiments were prepared with the miScript SYBR Green PCR Kit (218073; Qiagen) along with miScript Primer Assays (Qiagen), andwere performed on the Rotor-Gene Q real-time PCR cycler (Qiagen). Detailed information on qRT-PCR analysis is given elsewhere.…”

“…10 The discovery of microRNAs (miRNAs) as modulators of gene expression has resulted in extensive investigation of the ability of miRNAs to act as biomarkers in multisystem autoimmune disorders, including systemic vasculitides. [11][12][13][14][15][16] However, there is still a lack of information regarding miRNA expression in IgAV.…”