Vascular calcification is the formation of bone-like structures in the vessel wall. The pathology is characterised by osteogenic differentiation of vascular smooth muscle cells (VSMCs) and mineralisation of the extracellular matrix. Tumour necrosis factor-stimulated gene-6 (TSG-6) is an inflammation-associated protein that has been implicated in the inhibition of bone formation and thus, we hypothesised, may also influence vascular calcification.
Confluent VSMCs were cultured with 3 mM beta-glycerophosphate (BGP) to induce mineralisation. Calcification was confirmed by staining with alizarin red. Osteogenic differentiation was demonstrated by the up-regulation of osteogenic markers (e.g. Runx2 and Msx2) and down-regulation of smooth muscle cell markers (e.g. aSMA) by qPCR. TSG-6 mRNA expression is significantly up-regulated (6-fold, p < 0.0005) during the early stages of BGP-induced VSMC mineralisation, in comparison to control. TSG-6 was knocked-down using siRNA and reduced expression (>86%) confirmed using qPCR and western blotting. Loss of TSG-6 expression significantly increased mineralisation (1.6-fold, p < 0.001) and accelerated osteogenic differentiation. Knock-down of basal TSG-6 expression in VSMCs promoted a mature smooth muscle phenotype, as shown by up-regulation of contractile markers (smooth muscle myosin heavy chain, aSMA, SM22a and calponin) using qPCR and western blotting. On-going studies aim to determine the mechanism by which these effects occur.
In conclusion, TSG-6 is a novel regulator of vascular calcification, which can modulate both mineralisation and osteogenic differentiation of VSMCs. Our data also suggest that basal TSG-6 expression is crucial in maintaining the VSMC synthetic state, as loss of basal TSG-6 promotes a mature, contractile VSMC phenotype.
- vascular calcification
- smooth muscle cells
Statistics from Altmetric.com
If you wish to reuse any or all of this article please use the link below which will take you to the Copyright Clearance Center’s RightsLink service. You will be able to get a quick price and instant permission to reuse the content in many different ways.