Downregulation of KLF9 alleviates tubulointerstitial fibrosis by modulating FABP4-mediated lipid accumulation

Abstract

Tubulointerstitial fibrosis (TIF) is a significant determinant in the pathogenesis of chronic kidney disease (CKD) and is commonly concurrent with lipid infiltration in the renal tubules. Nonetheless, the precise regulatory mechanism of this phenomenon remains incompletely understood. This research sought to uncover the involvement and underlying mechanism of KLF9 in the accumulation of lipids linked to TIF. As renal fibrosis models, TGF-β1 treated HK-2 cells and a unilateral ureteral obstruction (UUO) mouse model were utilized. Histopathological analysis of kidney tissues were evaluated by hematoxylin eosin (HE), periodic acid schiff (PAS), and Masson's trichrome staining. The levels of KLF9 protein and mRNA were quantified through western blot and real-time quantitative PCR, respectively, while triglyceride (TG) levels and lipid accumulation were evaluated using a TG assay kit and Oil Red O staining, respectively. The Pearson correlation coefficient was employed to assess the relationship between KLF9 levels and lipid accumulation. To elucidate the mechanisms underlying KLF9's regulation of lipid accumulation in TIF, luciferase reporter assays, chromatin immunoprecipitation (ChIP), and rescue experiments were performed. This research identified a significant increase in KLF9 expression in TIF, correlating with lipid accumulation. The inhibition of KLF9 in HK-2 cells significantly mitigated TGF-β1 triggered fibrosis and lipid accumulation. Subsequent animal studies corroborated these findings, showing that downregulating KLF9 mitigated fibrosis and lipid accumulation. The expression level of FABP4 was considerably higher in TIF models both in vitro and in vivo. Mechanistically, KLF9 bound to the FABP4 promoter region and positively regulated the expression of FABP4. The KLF9-FABP4 pathway regulated lipid synthesis and promoted lipid accumulation, which in turn promotes the progression of TIF. Our study has unveiled the involvement of KLF9 in driving FABP4 expression at the transcriptional level, culminating in lipid accumulation and subsequent fibrosis in TIF. These findings propose that targeting lipid deposition as a therapeutic strategy may hold promise for addressing TIF.