Osteogenic differentiation of mesenchymal stem cells : Identification of potential regulators

Abstract

Osteogenic differentiation of mesenchymal stem cells (MSC) is a tightly regulated process, inducing large changes in cell phenotype and gene expression. Characterization of these expressional changes may be important in the understanding of stem cell biology and bone development, and further gain insight into the development of diseases such as cancer.

Conditions for osteogenic differentiation of the immortalized mesenchymal stem cell line iMSC#3 as well as primary human MSCs were optimized.

Gene expression profiles from osteogenic and adipogenic differentiations of iMSC#3 were used to select a subset of genes being specifically regulated during osteogenic differentiation. Expression patterns of selected genes were analyzed with quantitative PCR in both iMSC#3 and primary human MSCs, resulting in the selection of two genes, WNT1 induced secreted protein (WISP) 1 and WISP2, with high upregulation during osteogenic differentiation. Both genes encode proteins of the CYR61/CTGF/NOV (CCN)-family, a protein family having a broad range of functions including regulation of proliferation, differentiation, adhesion, migration, apoptosis and extracellular matrix production. Little is known about the function of these genes and their relation to osteogenic differentiation. To determine if WISP1 and WISP2 were regulators of osteogenic differentiation, siRNA was used to reduce gene expression during differentiation. Knocking down expression of these genes during osteogenic differentiation abolished the calcium mineralization, implicating these genes to be central regulators during in vitro osteogenic differentiation of MSCs.