Abstract
PurposeThis study aimed to investigate the effects of miR-29b on renal interstitial fibrosis in the obstructed kidney of mouse with unilateral ureteral obstruction (UUO) via inhibiting phosphatidylinositol 3-kinase/protein kinaseB (PI3K/AKT) signaling pathway.MethodsAdult male CD-1 mice were intraperitoneally injected with vehicle or PI3K inhibitor LY294002 (3 mg/kg, 30 mg/kg) daily for 1 or 2 weeks after performing UUO or sham operation. The mice were sacrificed on days 7 and 14 after surgery. The rat proximal tubular epithelial cell (TEC) line NRK-52E was cultured in DMEM and treated with various concentrations angiotensin II (AngII). Obstructed and sham mouse kidneys were analyzed via HE, Masson and immunohistochemistry to assess the degree of renal fibrosis. Real-time quantitative polymerase chain reaction assays (RT-PCR) were performed to investigate changes in the levels of expression of miR-29b and Western blot was used to analyze the activation of PI3K/AKT signaling and expression of E-cadherin, α-smooth muscle actin (α-SMA).ResultsHistologic analyses of obstructed kidney revealed that LY294002 attenuated the degree of renal fibrosis. In this study, loss of miR-29b accompanied with increased epithelial–mesenchymal transition (EMT) was observed in renal tubules of mice after UUO and cultured NRK-52E cells exposed to AngII. LY294002 also prominently decreased phosphorylation of AKT in vivo and vitro. By RT-PCR and Western blot analysis, LY294002 blocked the PI3K/AKT-induced loss of E-cadherin expression and de novo increase of the expression of α-SMA in a time- and dose-dependent manner. The overexpression of miR-29b markedly reversed the phenotype induced by AngII in NRK-52E cells and the downregulation miR-29b expression with an miR-29b inhibitor resulted in enhanced EMT. In addition, the PI3K/AKT signaling pathway was found to be suppressed in the presence of overexpression of miR-29b by direct hybridization with 3′-untranslated region (3′-UTR) of PIK3R2.ConclusionOur findings suggested that miR-29b significantly prevented tubulointerstitial injury in mouse model of UUO by attenuating renal tubular epithelial cell–mesenchymal transition via repressing PI3K/AKT signaling pathway.
Highlights
Renal tubulointerstitial fibrosis is a morphological hallmark of progressive chronic kidney disease (CKD), and is closely related to the loss of renal function [1, 2]
Our findings suggested that miR-29b significantly prevented tubulointerstitial injury in mouse model of ureteral obstruction (UUO) by attenuating renal tubular epithelial cell–mesenchymal transition via repressing phosphatidylinositol 3-kinase (PI3K)/AKT signaling pathway
The immunohistochemistry results of α-smooth muscle actin (α-SMA), a marker of myofibroblasts, showed that the total integral optical density (IOD) expression of α-SMA in the kidney tissues from mice having increased after UUO (Fig. 2a)
Summary
Renal tubulointerstitial fibrosis is a morphological hallmark of progressive chronic kidney disease (CKD), and is closely related to the loss of renal function [1, 2]. It has been confirmed that a considerable proportion of interstitial myofibroblasts are derived from tubular epithelial cells of the affected kidneys via epithelial–mesenchymal transition (EMT) [6,7,8]. The EMT process constitutes epithelial cells losing cell–cell contacts, apical–basal. A number of studies have shown that miRNAs play important roles in a variety of biological processes including EMT [10]. The miR-29 family consists of three members, which are encoded by two different genomic loci in both human and rodent genomes. They all bind to the same set of target genes, because the members have the same binding seed sequence [13]. The biological roles of miR-29b in renal fibrosis are poorly understood
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
