Mesenchymal stem cells-derived vascular smooth muscle cells release abundant levels of osteoprotegerin

Abstract

Although several studies have shown that the serum levels of osteoprotegerin (OPG) are significantly elevated in patients affected with atherosclerotic lesions in coronary and peripheral arteries, the cellular source and the role of OPG in the physiopathology of atherosclerosis are not completely defined. Therefore, we aimed to investigate the potential contribution of mesenchymal stem cells in the production/release of OPG. OPG was detectable by immunohistochemistry in aortic and coronary atherosclerotic plaques, within or in proximity of intimal vascular smooth muscle cells (SMC). In addition, bone marrow mesenchymal stem cell (MSC)-derived vascular SMC as well as primary aortic SMC released in the culture supernatant significantly higher levels of OPG with respect to MSC-derived endothelial cells (EC) or primary aortic EC. On the other hand, in vitro exposure to full-length human recombinant OPG significantly increased the proliferation rate of aortic SMC cultures, as monitored by bromodeoxyuridine incorporation. Taken together, these data suggest that OPG acts as an autocrine/paracrine growth factor for vascular SMC, which might contribute to the progression of atherosclerotic lesions.

Keywords: atherosclerosis.; mesenchymal stem cells; osteoprotegerin; smooth muscle cells.

Figure 1

OPG expression in SMC of human atherosclerotic plaques. (A) immunohistochemical detection of OPG in samples of human ascending aortas with or without (normal) atherosclerotic plaque. FC, Fibrous cap; LC, Lipidic core. (B) comparison of immunohistochemical staining of a plaque performed with α-smooth muscle actin (SMA) and OPG Abs indicates a good correspondence between the elongated cells expressing OPG and the SMC of atherosclerotic fibrous cap (FC). Scale bar: 100 µm. Representative pictures are shown.

Figure 2

High levels of OPG are released by BM MSC-derived SMC and primary aortic SMC. BM MSC were induced to differentiate into endothelial (MSC-Endo) and SMC (MSC-SMC). Culture differentiation was monitored by immunofluorescence analysis of lineage-specific markers: von Willebrand Factor for endothelial cells and α-SMA for myocytes (both in green). DAPI was used in all fluorescent images to label nuclei in blue. Original magnification 40×. Release of OPG in culture supernatants was measured by ELISA at different time-points of culture. OPG released by aortic EC and SMC cultures is also shown. Measurements were done in duplicate and normalized for cell number. Data is expressed as means α SD of results from three independent experiments.