MiR-133a Modulates Osteogenic Differentiation of Vascular Smooth Muscle Cells

Abstract

Arterial calcification is a key pathologic component of vascular diseases such as atherosclerosis, coronary artery disease, and peripheral vascular disease. A hallmark of this pathological process is the phenotypic transition of vascular smooth muscle cells (VSMCs) to osteoblast-like cells. Several studies have demonstrated that microRNAs (miRNAs) regulate osteoblast differentiation, but it is unclear whether miRNAs also regulate VSMC-mediated arterial calcification. In the present study, we sought to characterize the role of miR-133a in regulating VSMC-mediated arterial calcification. Northern blotting analysis of VSMCs treated with β-glycerophosphate demonstrated that miR-133a was significantly decreased during osteogenic differentiation. Overexpression of miR-133a inhibited VSMC transdifferentiation into osteoblast-like cells as evidenced by a decrease in alkaline phosphatase activity, osteocalcin secretion, Runx2 expression, and mineralized nodule formation. Conversely, the knockdown of miR-133a using an miR-133a inhibitor promoted osteogenic differentiation of VSMCs by increasing alkaline phosphatase activity, osteocalcin secretion, and Runx2 expression. Runx2 was identified as a direct target of miR-133a by a cotransfection experiment in VSMCs with luciferase reporter plasmids containing wild-type or mutant 3'-untranslated region sequences of Runx2. Furthermore, the pro-osteogenic effects of miR-133a inhibitor were abrogated in Runx2-knockdown cells, and the inhibition of osteogenic differentiation by pre-miR-133a was reversed by overexpression of Runx2, providing functional evidence that the effects of miR-133a in osteogenic differentiation were mediated by targeting Runx2. These results demonstrate that miR-133a is a key negative regulator of the osteogenic differentiation of VSMCs.

A current understanding of vascular calcification in CKD

“…When VSMC overexpress miR-133a in the presence of ␤-glycerophosphate (which induces osteogenic differentiation), these cells had decreased alkaline phosphatase activity, osteocalcin secretion, and Runx2 expression along with decreased evidence of mineralization. Similarly, inhibition of miR-133a was found to increase alkaline phosphatase activity, osteocalcin secretion, and Runx2 expression (61). Similarly, miR-204 has been shown to suppress the osteoblastic differentiation of mesenchymal progenitor cells by downregulating Runx2.…”

Roles of the canonical myomiRs miR-1, -133 and -206 in cell development and disease

“…Patients with aortic stenosis surgery [194] Apelin treatment reduces elevated circulating miRs Elevated miR-133a, miR-208 and miR-1 reduced High-fat diet elevated miRs and increased left ventricular diastolic and systolic diameters, and wall thickness Obesity-associated cardiac dysfunction in mouse model [195] NAC treatment Expressed miR-1, miR-499, miR-133a, and miR-133b were strongly depressed in the diabetic cardiomyocytes NAC restored expression of miR-499, miR-1, miR-133a, and miR-133b significantly in the myocardium Diabetic rat hearts [196] Myocardial junctin elevated miR-1 targets junctin NAC reduces junction levels Development of diabetic cardiomyopathy in rat hearts [196] CAD associated ischemic heart failure miR-133 expression decreased with increased severity of heart failure Patients with CAD [197] Runx2 miR-133a targets Runx2 Transition of VSMCs to osteoblast-like cells [198] Increased alkaline phosphatase activity, osteocalcin secretion and Runx2 expression miR-133a was decreased during osteogenic differentiation…”

“…Transition of VSMCs to osteoblast-like cells [198] Circulating miR-133a and 208a levels Cardiac muscle-enriched microRNAs (miR-133a, miR-208a) elevated…”

Vascular calcification: the role of microRNAs

“…Figure 1) miR-30b and miR-30c are downregulated in VSMCs from calcified coronary arteries. These miRs target RUNX2, a protein that plays a significant role in the transdifferentiation of VSMCs to osteoblasts (7,18). RUNX2 regulates osteocalcin, receptor activator of nuclear factor κ-B ligand (RANKL) and osteopontin, which in turn modulate bone matrix formation (4).…”

“…It appears to protect against vascular calcification. More specifically, miR-133a is downregulated in VSMCs that have transitioned to osteoblast-like cells (18). miR-133a binds to the 3′ UTR area of RUNX2, leading to reduced production of osteocalcin and ALP and ultimately protecting the VSMCs from transforming into osteoblast-like cells (18).…”