miR-132-3p and KLF7 as novel regulators of aortic stiffening-associated EndMT in type 2 diabetes mellitus

Abstract

Background The prevalence of diabetes mellitus has risen considerably and currently affects more than 422 million people worldwide. Cardiovascular diseases including myocardial infarction and heart failure represent the major cause of death in type 2 diabetes (T2D). Diabetes patients exhibit accelerated aortic stiffening which is an independent predictor of cardiovascular disease and mortality. We recently showed that aortic stiffness precedes hypertension in a mouse model of diabetes (db/db mice), making aortic stiffness an early contributor to cardiovascular disease development. Elucidating how aortic stiffening develops is a pressing need in order to halt the pathophysiological process at an early time point. Methods To assess EndMT occurrence, we performed co-immunofluorescence staining of an endothelial marker (CD31) with mesenchymal markers (α-SMA/S100A4) in aortic sections from db/db mice. Moreover, we performed qRT-PCR to analyze mRNA expression of EndMT transcription factors in aortic sections of db/db mice and diabetic patients. To identify the underlying mechanism by which EndMT contributes to aortic stiffening, we used aortas from db/db mice and diabetic patients in combination with high glucose-treated human umbilical vein endothelial cells (HUVECs) as an in vitro model of diabetes-associated EndMT. Results We demonstrate robust CD31/α-SMA and CD31/S100A4 co-localization in aortic sections of db/db mice which was almost absent in control mice. Moreover, we demonstrate a significant upregulation of EndMT transcription factors in aortic sections of db/db mice and diabetic patients. As underlying regulator, we identified miR-132-3p as the most significantly downregulated miR in the micronome of db/db mice and high glucose-treated HUVECs. Indeed, miR-132-3p was also significantly downregulated in aortic tissue from diabetic patients. We identified Kruppel-like factor 7 (KLF7) as a target of miR-132-3p and show a significant upregulation of KLF7 in aortic sections of db/db mice and diabetic patients as well as in high glucose-treated HUVECs. We further demonstrate that miR-132-3p overexpression and KLF7 downregulation ameliorates EndMT in high glucose-treated HUVECs. Conclusions We demonstrate for the first time that EndMT contributes to aortic stiffening in T2D. We identified miR-132-3p and KLF7 as novel EndMT regulators in this context. Altogether, this gives us new insights in the development of aortic stiffening in T2D.

Non-coding RNA-mediated endothelial-to-mesenchymal transition in human diabetic cardiomyopathy, potential regulation by DNA methylation

“…EndMT has also been found to occur in calcific aortic valve disease, where it precedes EC osteogenesis [ 52 , 53 ], and in diabetes-related aortic stiffening [ 35 ]. These instances of EndMT have been shown in both experimental models and in humans [ 35 , 51 , 52 ]. We and others have shown that hyperglycemia-induced EndMT occurs and contributes to DCM both in vitro and in vivo, but these findings have never been recapitulated in human diabetic patients [ 8 , 9 , 16 , 19 , 37 ].…”

“…HDACs 3 and 9 have been shown to induce EndMT in atherosclerotic plaques [ 32 , 33 ], and DNMT1 has been found to promote EndMT in diabetic retinopathy by hypermethylating the lncRNA MEG334. Recently, miR-132-3p—downregulated in diabetes—has been reported to regulate EndMT in the aorta via regulation of Kruppel-like factor 7 [ 35 ]. We have previously identified lncRNA ZFAS1 and miRNAs 9, 146a, and 200b as regulators of fibrosis and EndMT in DCM and other diabetic complications in cell culture and in animal models [ 8 , 9 , 18 , 36 – 38 ].…”

Screening of the Key Genes and Signaling Pathways for Schizophrenia Using Bioinformatics and Next Generation Sequencing Data Analysis

“…RUNX1 [186], hsa-mir-132-3p [606] and HOXA5 [607] was an important therapeutic targets of multiple sclerosis. RUNX1 [81], CAV1 [241], STAT6 [239], CDKN2A [226], SYK (spleen associated tyrosine kinase) [148], hsa-mir-132-3p [608], hsa-mir-10b-5p [609], hsa-mir-155-5p [610], hsa-mir-191-5p [611], SRY (sex determining region Y) [612], PAX2 [613] and PDX1 [614] were the potential molecular targets of the drugs for treating diabetes mellitus. RUNX1 [395], CDKN2A [403], HSPA2 [423], hsa-mir-132-3p [615], hsa-mir-155-5p [610] and USF2 [616] have been reported to be expressed in Alzheimer’s disease.…”

Krüpple-like factors in cardiomyopathy: emerging player and therapeutic opportunities

“…Although the underlying mechanism of some KLFs in diabetes remains unclear ( 44 ), studies have acknowledged the essential role KLFs play in many types of diabetes ( 71 , 89 , 119 – 124 ) ( Table 3 ). For example, it was reported that KLF7 can regulate insulin sensitivity and susceptibility to type 2 diabetes by lowering adiponectin and leptin levels ( 126 , 127 ), and can negatively regulate miR-132-3p to aggravate the transition of Human Umbilical Vein Endothelial Cells (HUVECs) to a mesenchymal state after high glucose exposure ( 128 ). Similarly, KLF14 has been implicated in increasing susceptibility to type 2 diabetes by regulating key genes associated with insulin resistance ( 122 , 123 ).…”