KLF11 deficiency exacerbates renal damage in experimental unilateral ureteral obstruction

Abstract

KLFs (Kruppel-like factors) comprise a family of over 17 distinct transcription factors that have been shown to regulate cell metabolism and renal function, including podocyte differentiation and maintenance of the glomerular filtration barrier. In non-renal systems, KLF11 has been shown to regulate fibrosis. However, no previous studies have established a potential role for KLF11 in renal pathophysiology. We sought to test the hypothesis that KLF11 regulates renal fibrosis in unilateral ureteral obstruction, a well-established murine model of renal fibrosis. Mice with homozygous deletion of the KLF11 gene (KO) and wild-type (WT) mice were subjected to unilateral ureteral obstruction (UUO; n=8 per group); sham controls (n=4 per group) were subjected to manipulation of the ureter without ligation. Kidneys were harvested at 9 days following surgery,. Semiquantitative histopathologic analysis of interstitial fibrosis, tubular atrophy, and interstitial inflammation was performed in a blinded fashion. Analysis of gene expression was performed on renal cortex using Pathway Detect RNA arrays and by RNASeq. Histopathologic analysis of sham controls revealed no significant alterations. However, expression of several elements of the TGF-beta signaling pathway was significantly higher in KLF11 KO sham mice, compared to WT mice, including TGF-beta receptors type I and type II, Smad2, Smad 7, Bmpr1B, and Bmpr2R, and the KLF-related transcription factor Sp1 (all p<0.05). In mice subjected to UUO, interstitial fibrosis, tubular atrophy, and interstitial inflammation were significantly greater in KLF11 KO mice compared to WT mice. Interstitial inflammation was associated with influx of alternatively activated macrophages, as assessed by quantitative analysis of CD206+ infiltrating cells (p<0.05); no significant increase in CD3+ T cell numbers were noted. Expression of several members of the TGF-beta signaling pathway were significantly increased in KLF11 KO UUO mice, compared to WT UUO mice, including TGF-beta 1, TGF-beta-2, and TGF-beta3, TGF-beta receptors type I and type II, and Smad 1,2,3,4, and 5 (all p<0.05). Conversely, expression of the transcription factor JunB was markedly increased in WT UUO mice compared to WT sham mice (12.2 -fold; p<0.0001). However, JunB expression was markedly decreased in KLF11 KO mice (both sham and UUO). Although MMP expression was not significantly different in KO versus WT mice, TIMP3 expression was higher in KO mice compared to WT mice. In conclusion, we found that renal damage is exacerbated in KLF11 KO mice subjected to UUO, compared to WT UUO mice and is associated with influx of potentially pro-fibrotic, alternatively activated M2 macrophages. KLF11 appears to be a critical regulator of TGF-beta signaling, as KLF11 deficiency is associated with increased expression of TGF-beta and signaling intermediates of this fibrogenic pathway.