Assessment of gene expression during osteoblast induction of human mesenchymal stem cells

Abstract

The aim of this study was to define transcriptional changes that occur during dexamethasone-induced in vitro osteoblastic differentiation using human mesenchymal stem cells (hMSCs). Bone marrow derived hMSCs from three individual donors were grown in DMEM containing 10% fetal bovine serum and antibiotics (basal media, BM). At confluence (Day 0), cells were grown in BM or BM plus ascorbic acid, beta-glycerophosphate and dexamethasone (OS). At 0, 3, 7 and 14 days, total RNA was isolated from hMSCs, and 32ρ-labelled probes were synthesized. Atlas 1.2 gene arrays (Clontech) were hybridized with donor-specific cDNA probes. Acquired data were analyzed using GeneSpring Software (Silicon Genetics) enabling normalization and averaging of donor-specific data sets at each time point. Only 8, 31, and 27 genes were upregulated more than 3-fold at 3, 7 and 14 days, respectively. Over 50 genes were downregulated at each time point. Interestingly, the results of this study did not reflect commonly identified genes that are frequently considered in cellular differentiation along the osteoblastic lineage. Instead, some interesting genes were upregulated. Seven days exposure to OS media resulted in the relatively high induction of several genes, including growth factors and receptors which affect adipocyte, chondrocyte and osteoblast differentiation. IGF II and FGFR 3, FGFR 2 were upregulated in the middle of 14 days' induction. Examination of gene expression at day 14 also revealed a number of receptors (LEPR) and transcription factors (ROR alpha) were upregulated during 14 days. These could be of importance to the process of early stage differentiation.