Reticulocalbin 2 enhances osteogenic differentiation of human vascular smooth muscle cells in diabetic conditions

Aim: Diabetes accelerates pro-atherogenic and pro-osteogenic phenotypes of vascular smooth muscle cells (VSMCs), an important process for vascular calcification. Reticulocalbin 2 (RCN2) is a candidate gene for atherosclerosis and involved in vascular remodeling in hypertension. However, the role of RCN2 in VSMCs calcification under diabetic conditions is unclear.

Materials and methods: Expression of RCN2 and Runt-related transcription factor 2 (Runx2) in femoropopliteal arterial plaques was compared between type 2 diabetes mellitus (DM) and non-DM patients using immunohistochemical staining (IHCS). Human aortic VSMCs (HAVSMCs) were analyzed under RCN2 gene knockdown and overexpression conditions. Alizarin red staining and intracellular calcium deposition quantification were used to observe calcification induced in vitro under normal glucose or high glucose combined with β-glycerol phosphoric acid conditions. The cells were investigated for gene modulation of osteogenic differentiation markers using Western blotting.

Key findings: The expression of RCN2 and Runx2 in femoropopliteal artery plaques was significantly higher in DM than in non-DM patients. In addition, a significant positive correlation was observed between RCN2 and Runx2 levels. RCN2 was highly expressed when HAVSMCs were treated with high glucose and the expression levels correlated with the calcification characteristics. RCN2 upregulated osteogenic transformation markers Runx2 and Osterix in HAVSMCs and downregulated contractile phenotype markers α-SMA and SM22α.

Significance: The results from this study indicate RCN2 is a major factor in mediating the calcification process of HAVSMCs in diabetic conditions. Thus, RCN2 may serve as a future therapeutic target for vascular calcification in diabetes.