Abstract 443: Constitutive Bone Morphogenetic Protein 2-dependent Osteogenic Signaling in M1 Macrophages

Abstract

Atherosclerosis is the leading cause of carotid and coronary artery disease. Calcification often complicates atherosclerotic plaques and contributes to plaque instability.Mechanisms calcification are somewhat reminiscent of bone formation, involving interplay between endothelium, smooth muscle cells and plaque macrophages. Macrophages contribute to vascular calcification through signals that stimulate the osteogenic program in smooth muscle cells and/or by becoming osteoclast-like cells. Yet, the specific roles of polarized M1 (pro-inflammatory) and M2 (anti-inflammatory) macrophages, predominant in plaques, in lesion calcification remain poorly understood. Recent studies from our laboratory show that the calcium-permeable channel Transient Receptor Potential Canonical 3 (TRPC3) is a key component of mechanisms linked to ER stress signaling in M1, but not M2 macrophages. Because ER stress has a recognized effect in inducing vascular calcification, we speculated that TRPC3, by virtue of its roles in M1 macrophages, might have an effect on the osteogenic potential of these cells. To address this question we utilized bone marrow derived macrophages from mice with macrophage-specific loss of TRPC3 function (MacTRPC3KO) or from their littermate controls (TRPC3 lox/lox ) and polarized in vitro to the M1 and M2 phenotypes. Osteogenic proteins and factors along with signaling mechanisms from the two groups were examined under basal and ER stress conditions. The results showed reduced expression of BMP-2 and Runx-2 at mRNA level in MacTRPC3KO M1 macrophages but not at the protein level between the groups. We also examined the phosphorylation status of SMAD1/5 in M1 macrophages, which is an early indicator of signaling downstream the BMP-2 receptor. Although no differences were observed between groups, SMAD1/5 was significantly phosphorylated, even under basal conditions. BMP-2 levels in supernatants from cultures of M1 macrophages was also significantly elevated regardless of the treatment condition. Notably, phosphorylation levels of SMAD1/5 were markedly reduced when cells were exposed to the BMP-2 receptor blocker dorsomorphin. These results indicate that BMP-2 is involved in activating osteogenic signaling in M1 macrophages.