SETD8 cooperates with MZF1 to participate in hyperglycemia-induced endothelial inflammation via elevation of WNT5A levels in diabetic nephropathy

Abstract

Objective: Diabetic nephropathy (DN) is regarded as the main vascular complication of diabetes mellitus, directly affecting the outcome of diabetic patients. Inflammatory factors were reported to participate in the progress of DN. Wingless-type family member 5 (WNT5A), myeloid zinc finger 1 (MZF1), and lysine methyltransferase 8 (SETD8) have also been reported to elevate inflammatory factor levels and activate the nuclear factor kappa B (NF-κB) pathway to induce endothelial dysfunction. In the current study, it was assumed that MZF1 associates with SETD8 to regulate WNT5A transcription, thus resulting in hyperglycemia-induced glomerular endothelial inflammation in DN. Methods: The present study recruited 25 diagnosed DN patients (type 2 diabetes) and 25 control participants (nondiabetic renal cancer patients with normal renal function, stage I–II) consecutively. Moreover, a DN rat and cellular model was constructed in the present study. Immunohistochemistry, Western blot, and quantitative polymerase chain reaction (qPCR) were implemented to determine protein and messenger RNA (mRNA) levels. Coimmunoprecipitation (CoIP) and immunofluorescence were implemented in human glomerular endothelial cells (HGECs). Chromatin immunoprecipitation assays and dual luciferase assays were implemented to determine transcriptional activity. Results: The results of this study indicated that levels of WNT5A expression, p65 phosphorylation (p-p65), and inflammatory factors were all elevated in DN patients and rats. In vitro, levels of p-p65 and inflammatory factors increased along with the increase of WNT5A expression in hyperglycemic HGECs. Moreover, high glucose increased MZF1 expression and decreased SETD8 expression. MZF1 and SETD8 inhibit each other under the stimulus of high glucose, but cooperate to regulate WNT5A expression, thus influencing p-p65 and endothelial inflammatory factors levels. Overexpression of MZF1 and silencing of SETD8 induced endothelial p-p65 and inflammatory factors levels, which can be reversed by si-WNT5A. Mechanistic research indicated that MZF1, SETD8, and its downstream target histone H4 lysine 20 methylation (H4K20me1) all occupied the WNT5A promoter region. sh-SETD8 expanded the enrichment of MZF1 on WNT5A promoter. Our in vivo study proved that SETD8 overexpression inhibited levels of WNT5A, p-p65 expression, and inflammatory factors in DN rats. Conclusions: MZF1 links with SETD8 to regulate WNT5A expression in HGECs, thus elevating levels of hyperglycemia-mediated inflammatory factors in glomerular endothelium of DN patients and rats. Trial registration ChiCTR, ChiCTR2000029425. 2020/1/31, http://www.chictr.org.cn/showproj.aspx?proj=48548