Abstract
Cardiovascular diseases are commonly appreciated to represent a chronic inflammatory response of the vascular wall, and its complications cause high mortality in patients. Although the pathophysiological mechanisms underlying cardiovascular diseases have been well-established, new molecules that control the progress of these pathologies are continuously discovered. Recently, the aberrant expression of microRNAs (miRNAs) has been revealed in diverse cardiovascular diseases, and recognized as crucial regulators in the progression of these cardiovascular diseases by regulating the function of endothelial cells (ECs), smooth muscle cells (SMCs) and macrophages. Convincingly, miRNAs regulate several cellular processes involved in the development of these cardiovascular diseases, such as vascular inflammation and proliferation. In this thesis, co-culturing ECs with synthetic SMCs under static condition causes initial increases of four anti-inflammatory miRNAs (146a/708/451/98) in ECs followed by decreases below the basal levels at 7 days; the increases for miR-146a/708 peaked at 24 h and those for miR-451/98 lasted for only 6-12 h. Shear stress (12 dynes/cm2) to co-cultured ECs for 24 h augments these four miRNA expressions. In vivo, these four miRNAs are not expressed in neointimal ECs in injured arteries under flow stagnation, but become highly expressed under physiological levels of flow four weeks after balloon injury. MiR-146a, -708, -451, and -98 target interleukin (IL)-1 receptor-associated kinase-1 (IRAK-1), inhibitor of nuclear factor-κB (NF-κB) kinase subunit-, IL-6 receptor, and conserved helix-loop-helix ubiquitous kinase, respectively, to inhibit NF-κB signaling, which exerts negative feedback control on the biogenesis of these miRNAs. NF-E2-related factor-2 (Nrf-2) is critical for shear-induction of miR-146a in co-cultured ECs. Silencing either Nrf-2 or miR-146a in ECs led to increased neointima formation of injured rat carotid artery under physiological levels of flow. Overexpressing miR-146a in ECs inhibits neointima formation of rat or mouse carotid artery induced by injury or blood flow cessation. Among these miRNAs, the miR-451 was significantly expressed in vascular SMCs of the medial layer in diseased human coronary arteries. In vitro study showed that culturing vascular SMCs on fibrillar collagen increased miR-451 level and suppressed proliferative and anti-inflammatory responses; these effects were diminished by anti-miR-451. Rab5a was inhibited by miR-451 directly targeting to lead to suppression of vascular SMC proliferation and inflammation. In vivo studies on rats indicated that overexpression of miR-451 and lentivirus-mediated Rab5a silencing markedly suppressed the neointima formation induced by balloon injury. The level of circulating miR-451 in blood plasma of patients with coronary artery disease (CAD) and apolipoprotein E knockout mice (ApoE-/-) fed with a high-cholesterol diet was significantly lower than control group. Increase of circulating miR-451 by tail vein injection with agomir-451 in ApoE-/- mice fed with a high-cholesterol diet for three months decreased atherosclerotic lesion formation. Taken together, these findings indicate that EC miR-146a and SMC miR-451, by respectively targeting IRAK-1 and Rab5a, respectively inhibits inflammation and proliferation in cultured EC and SMCs in vitro, and suppresses neointima formation in injured arteries in vivo. These results suggest the miR-451 may serve as potential biomarker for CAD, miR-146a and miR-451 as a potential target for treatment against neointima formation in cardiovascular diseases.