Flow-Dependent Regulation of Krüppel-Like Factor 2 Is Mediated by MicroRNA-92a

Abstract

Background Upregulated by atheroprotective flow, the transcription factor Krüppel-like factor 2 (KLF2) is crucial for maintaining endothelial function. MicroRNAs (miRNAs) are non-coding small RNAs that regulate gene expression at the post-transcriptional level. We examined the role of miRNAs, particularly miR-92a, in the atheroprotective flow-regulated KLF2. Methods and Results Dicer knockdown increased the level of KLF2 mRNA in human umbilical vein endothelial cells (HUVECs), suggesting that KLF2 is regulated by miRNA. In silico analysis predicted that miR-92a could bind to the 3’ untranslated region (3’UTR) of KLF2 mRNA. Overexpression of miR-92a precursor (pre-92a) decreased the expression of KLF2 and the KLF2-regulated endothelial nitric oxide synthase (eNOS) and thrombomodulin (TM) at mRNA and protein levels. A complementary finding is that miR-92a inhibitor (anti-92a) increased the mRNA and protein expression of KLF2, eNOS, and TM. Subsequent studies revealed that atheroprotective laminar flow downregulated the level of miR-92a to induce KLF2 and the level of this flow-induced KLF2 was reduced by pre-92a. Furthermore, miR-92a level was lower in HUVECs exposed to the atheroprotective pulsatile shear flow (PS) than under atheroprone oscillatory shear flow. Anti-Ago1/2 immunopreciptation coupled with RT-PCR revealed that PS decreased the functional targeting of miR-92a/KLF2 mRNA in HUVECs. Consistent with these findings, mouse carotid arteries receiving pre-92a exhibited impaired vasodilatory response to flow. Conclusions Atheroprotective flow patterns decrease the level of miR-92a, which in turn increases KLF2 expression to maintain endothelial homeostasis.

DNA Methyltransferase 1–Dependent DNA Hypermethylation Constrains Arteriogenesis by Augmenting Shear Stress Set Point

“…HUVECs are both phenotypically consistent and one of the most extensively used cell culture models for study of flow‐mediated EC signaling, including numerous studies of flow responses of the arterial system17, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 (eg, atheroprone versus atheroprotective waveforms) and DNMT1‐dependent DNA methylation 17, 18. For each set of experimental comparisons, cells were used from the same cell line between subculture passages 2 to 3.…”

FBW7 regulates endothelial functions by targeting KLF2 for ubiquitination and degradation

“…Similarly, the antiinflammation drugs statins also induce KLF2 expression via MEF2 activation [18]. Recent evidence shows that miRNA-92a regulates KLF2 mRNA in endothelial cells [19]. Moreover, proteasome inhibitors can also induce the expression of KLF2 mRNA in the endothelium and thus increase the expression of Thrombomodulin [20].…”

IGF-1 deficiency in a critical period early in life influences the vascular aging phenotype in mice by altering miRNA-mediated post-transcriptional gene regulation: implications for the developmental origins of health and disease hypothesis

“…miRNA-dependent pathways have been shown to regulate multiple aspects of cellular physiology relevant for vascular aging, including angiogenesis (Kuehbacher et al 2007;Suarez et al 2007Suarez et al , 2008Yang et al 2005), structural integrity of the vessels, replicative senescence (Menghini et al 2009;Vasa-Nicotera et al 2011), mechanotransduction (Wu et al 2011), NO production (Suarez et al 2007;Wu et al 2011), endothelial apoptosis (Asada et al 2008), and inflammation (Suarez et al 2007). Among the miRNAs whose expression is regulated by developmental IGF-1 deficiency, upregulation of miR-125a-5p has been linked to impaired angiogenesis and endothelial dysfunction (Che et al 2014), endothelial apoptosis (Svensson et al 2014), and dysregulation of endothelial tight junctions (Reijerkerk et al 2013).…”