Abstract 1418: Receptor For Advanced-glycation End Products Inhibits Myocardin Function And Induces Msx2-dependent Osteoblastic Differentiation Of Vascular Smooth Muscle Cells

Abstract

As an activator of diverse signaling pathways in the vascular cells, receptor for advanced-glycation end products (RAGE) has been related to atherosclerosis initiation and progression in the patients with diabetes and chronic renal failure (CRF). However, little is known about the efects of RAGE activation on the differentiation program of vascular smooth muscle cells (VSMC). We first examined effects of RAGE on expression of myosin heavy chain (MHC), a definitive VSMC marker. RT-PCR analyses showed that expression of MHC was diminished in VSMC transduced with adenovirus expressing RAGE. A RAGE ligand HMGB1, but not another ligand AGE2, enhanced RAGE-dependent MHC repression. Activation of HMGB1-RAGE pathway induced the endogenous RAGE expression, suggesting positive feedback loop in atherosclerotic lesion. In mesenchymal 10T1/2 cells, HMGB1-RAGE inhibited an induction of MHC gene expression by myocardin, a potent positive regulator of VSMC differentiation. We next examined the effects of RAGE on osteogenic conversion of VSMC by monitoring the expression and activity of alkaline phosphatase (ALP). RT-PCR analysis showed that expression of ALP was up-regulated in VSMC when RAGE was overexpressed by adenovirus. The degree of the ALP induction was dependent on the concentration of fetal bovine serum, suggesting that fetal bovine serum contains RAGE ligands. The induction of ALP was accompanied by ALP activity and an inducible expression of osteogenic transcription factors, Msx2 and Runx2 as well as an increase in osteopontin and bone morphogenic protein 2 (BMP2) expression. We further found that addition of BMP2 and RAGE synergistically enhanced expression of ALP mRNA and ALP activity. Immunohistochemistry revealed that RAGE antigen is present in calcified atherosclerotic plaque in a distribution overlapping with osteoblastic markers and calcium deposition detected by Kossa staining in human carotid artery. These findings suggest that RAGE induction in VSMC contributes to the development of calcifying atherosclerotic plaque characterizing the diabetes- and CRF-associated vascular lesion by repressing myocardin-induced VSMC differentiation and inducing Msx2-dependent osteogenic differentiation.