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Can valvular interstitial cells become true osteoblasts? A side-by-side comparison.

ABSTRACT: Aortic valve calcification is believed to involve the differentiation of valvular interstitial cells (VICs) into either a myofibroblastic or an osteoblast-like phenotype. Despite purported similarities between diseased VICs and osteoblasts, few studies have directly compared VICs and osteoblasts in side-by-side experiments. In the present study, VICs were compared against multiple osteoblastic cell types at different stages of differentiation. These findings may help to resolve whether VICs progress through a myofibroblastic phenotype before reaching an osteoblast-like stage.Three cell types representing a range of osteoblastic lineage commitment and differentiation were used in the phenotypic comparison against VICs. Specifically, VICs, embryonic fibroblasts (C3H10T1/2), pre-osteoblasts (MC3T3-E1), and mature primary osteoblasts were cultured on tissue-culture polystyrene in control or mineralization medium, and harvested for qPCR, DNA, and protein analysis at time points ranging from one to eight days.Culture of VICs in mineralization medium decreased the expression of alpha-smooth muscle actin (alpha-SMA; a myofibroblast marker), with no peak in alpha-SMA gene or protein expression in mineralization medium at any time point. The application of a mineralization medium led to increased expression levels of alkaline phosphatase (ALP; an early mineralization marker) for all cell types, although the magnitude of the increase in ALP was drastically smaller for VICs than for the osteogenic cell types. Only the osteogenic cell types demonstrated an appreciable increase in osteocalcin (an indicator of later-stage mineralization).While the addition of mineralization medium generally increased the expression of osteogenic markers and decreased the expression of myofibroblastic markers, VICs displayed different levels and patterns of expression than the osteoblastic cell types used for comparison. Additionally, the lack of an alpha-SMA increase at any point after the addition of mineralization medium to VICs indicated that these cells may not need to progress through a myofibroblastic stage before reaching an osteoblast-like gene expression profile.

SUBMITTER: Monzack EL

PROVIDER: S-EPMC3285463 | biostudies-literature | 2011 Jul

REPOSITORIES: biostudies-literature

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Can valvular interstitial cells become true osteoblasts? A side-by-side comparison.

Monzack Elyssa L EL Masters Kristyn S KS

The Journal of heart valve disease 20110701 4

<h4>Background and aim of the study</h4>Aortic valve calcification is believed to involve the differentiation of valvular interstitial cells (VICs) into either a myofibroblastic or an osteoblast-like phenotype. Despite purported similarities between diseased VICs and osteoblasts, few studies have directly compared VICs and osteoblasts in side-by-side experiments. In the present study, VICs were compared against multiple osteoblastic cell types at different stages of differentiation. These findin ...[more]

PMID: 21863660

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Project description:The development of a substance or inhibitor-based treatment strategy for the prevention of aortic valve stenosis is a challenge and a main focus of medical research in this area. One strategy may be to use the tankyrase inhibitor XAV-939, which leads to Axin stabilisation and subsequent destruction of the ?-catenin complex and dephosphorylation of ?-catenin. The dephosphorylated active form of ?-catenin (non-phospho-?-catenin) then promotes nuclear transcription that leads to osteogenesis. The aims of the present study were to develop an experimental system for inducing in vitro calcification of human aortic valvular interstitial cells (VICs) to investigate the potential anti-calcific effect of XAV-939 and to analyse expression of the Wnt signalling proteins and Sox9, a chondrogenesis regulator, in this model. Calcification of human VIC cultures was induced by cultivation in an osteogenic medium and the effect of co-incubation with 1?M XAV-939 was monitored. Calcification was quantified when mineral deposits were visible in culture and was histologically verified by von Kossa or Alizarin red staining and by IR-spectroscopy. Protein expression of alkaline phosphatase, Axin, ?-catenin and Sox9 were quantified by western blotting. In 58% of the VIC preparations, calcification was induced in an osteogenic culture medium and was accompanied by upregulation of alkaline phosphatase. The calcification induction was prevented by the XAV-939 co-treatment and the alkaline phosphatase upregulation was suppressed. As expected, Axin was upregulated, but the levels of active non-phospho-?-catenin were also enhanced. Sox9 was induced during XAV-939 treatment but apparently not as a result of downregulation of ?-catenin signalling. XAV-939 was therefore able to prevent calcification of human VIC cultures, and XAV-939 treatment was accompanied by upregulation of active non-phospho-?-catenin. Although XAV-939 does not downregulate active ?-catenin, treatment with XAV-939 results in Sox9 upregulation that may prevent the calcification process.

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Can valvular interstitial cells become true osteoblasts? A side-by-side comparison.

Publications

Can valvular interstitial cells become true osteoblasts? A side-by-side comparison.

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