Novel markers of osteogenic and adipogenic differentiation of human bone marrow stromal cells identified using a quantitative proteomics approach

Abstract

Today, the tool that is most commonly used to evaluate the osteogenic differentiation of bone marrow stromal cells (BMSCs) in vitro is the demonstration of the expression of multiple relevant markers, such as ALP, RUNX2 and OCN. However, as yet, there is no single surface marker or panel of markers which clearly defines human BMSCs (hBMSCs) differentiating towards the osteogenic lineage. The aim of this study was therefore to examine this issue. Stable isotope labeling by amino acids in cell culture (SILAC)-based quantitative proteomics was utilized to investigate differently expressed surface markers in osteogenically differentiated and undifferentiated hBMSCs. Labeled membrane proteins were analyzed by mass spectrometry (MS) and 52 proteins with an expression ratio above 2, between osteogenically differentiated and undifferentiated cells, were identified. Subsequent validation, by flow cytometry and ELISA, of the SILAC expression ratios for a number of these proteins and investigations of the lineage specificity of three candidate markers were performed. The surface markers, CD10 and CD92, demonstrated significantly increased expression in hBMSCs differentiated towards the osteogenic and adipogenic lineages. In addition, there was a slight increase in CD10 expression during chondrogenic differentiation. Furthermore, the expression of the intracellular protein, crystalline-αB (CRYaB), was only significantly increased in osteogenically differentiated hBMSCs and not affected during differentiation towards the chondrogenic or adipogenic lineages. It has been concluded from the present results that CD10 and CD92 are potential markers of osteogenic and adipogenic differentiation and that CRYaB is a potential novel osteogenic marker specifically expressed during the osteogenic differentiation of hBMSCs in vitro.

Incorporation of aligned PCL–PEG nanofibers into porous chitosan scaffolds improved the orientation of collagen fibers in regenerated periodontium

“…ALP and Bglap (OCN) are commonly used as early and late markers of osteogenic differentiation [50]. In our study, increased ligamentogenesis and partially decreased osteogenesis of progenitor cells mediated by aligned surface topography was noted.…”

Cardiac Fibroblasts Adopt Osteogenic Fates and Can Be Targeted to Attenuate Pathological Heart Calcification

“…In contrast, genes that were minimally expressed in cardiac fibroblasts were induced in a specific temporal manner during the course of osteogenic differentiation (Figure 1C) and included sets of genes known to regulate inflammation, extracellular matrix proteins and cell metabolism (Figure S1A). Next, we created an osteogenic signature based on a set of 37 genes that are induced during osteogenic differentiation (Chen et al, 2012);(Choi et al, 2010);(Graneli et al, 2014);(Harkness et al, 2011);(Hoshiba et al, 2009);(Liu et al, 2013);(Miguez et al, 2014);(Nora et al, 2012);(Olivares-Navarrete et al, 2011) (Figure S1B). We used the mean fold change in expression of this set of genes to quantitatively determine an osteogenic signature and observed that compared to control cardiac fibroblasts, cardiac fibroblasts subjected to osteogenic differentiation progressively adopted an osteogenic signature (Figure 1D).…”

“…1685 genes with a FDR < 0.05 and a minimum 2 fold (log 2 ) change were subsequently clustered and visualized. A set of osteogenic genes known to be induced during bone formation or osteogenic differentiation was compiled from the literature to determine an osteogenic signature (Chen et al, 2012); (Choi et al, 2010); (Graneli et al, 2014); (Harkness et al, 2011); (Hoshiba et al, 2009); (Liu et al, 2013); (Miguez et al, 2014); (Nora et al, 2012); (Olivares-Navarrete et al, 2011). All RNA-seq analysis were performed with ≥ 2 biological replicates.…”

Cytotoxicity and osteogenic potential of silicate calcium cements as potential protective materials for pulpal revascularization

“…It is interesting that OCN was upregulated in the present study after hDPSCs were exposed to MTA Angelus and Bodentine, while Runx2 was neither upregulated nor downregulated. Because the process of osteogenic differentiation from mesenchymal stem cells occurs in different phases [54,55], each of which is characteristized by a particular pattern of osteogenic marker genes, it is possible that MTA Angelus and Biodentine may upregulate other Runx-2 dependent or Runx-2 independent transcription genes, such as osterix, Wnt/b-catenin signaling, homeobox proteins DLX-3, DLX-5, MSX-2, or other less well-characterized downstream targets [52,55-60]. These factors have not been examined in the present work and require further investigation.…”