Gene expression profiling of human mesenchymal stem cells derived from bone marrow during expansion and osteoblast differentiation

Abstract

BackgroundHuman mesenchymal stem cells (MSC) with the capacity to differentiate into osteoblasts provide potential for the development of novel treatment strategies, such as improved healing of large bone defects. However, their low frequency in bone marrow necessitate ex vivo expansion for further clinical application. In this study we asked if MSC are developing in an aberrant or unwanted way during ex vivo long-term cultivation and if artificial cultivation conditions exert any influence on their stem cell maintenance. To address this question we first developed human oligonucleotide microarrays with 30.000 elements and then performed large-scale expression profiling of long-term expanded MSC and MSC during differentiation into osteoblasts.ResultsThe results showed that MSC did not alter their osteogenic differentiation capacity, surface marker profile, and the expression profiles of MSC during expansion. Microarray analysis of MSC during osteogenic differentiation identified three candidate genes for further examination and functional analysis: ID4, CRYAB, and SORT1. Additionally, we were able to reconstruct the three developmental phases during osteoblast differentiation: proliferation, matrix maturation, and mineralization, and illustrate the activation of the SMAD signaling pathways by TGF-β2 and BMPs.ConclusionWith a variety of assays we could show that MSC represent a cell population which can be expanded for therapeutic applications.

Stiffening hydrogels to probe short- and long-term cellular responses to dynamic mechanics

“…Our work shows that there is an inherent heterogeneity to the hMSC population, even though the starting hMSCs exhibited high surface expression for mesenchymal markers (>90%) and were negative for hematopoietic markers ( < 10%) [48][49][50] . Thus, we believe there was epigenetic heterogeneity to the cells, which led to differences in the rate of differentiation on the hydrogel substrates, leading to differential populations based on the timing of substrate stiffening, as committed cells did not respond to the mechanical changes.…”

“…The hMSCs were reported to be positive for mesenchymal surface markers (>90% CD105, CD166, CD29 and CD44) and negative for hematopoietic markers ( < 10% CD14, CD34 and CD45). To ensure the undifferentiated state of hMSCs, we used early passage cells (passage 3) that are previously reported to maintain these same levels of surface markers [48][49][50] . In this study, hMSCs at passage 3 were seeded onto MeHA substrates at 5×10 3 cells per cm 2 .…”

Characterization of the growth modulatory activities of osteoblast conditioned media on cord blood progenitor cells

“…On one hand, OCM contains large cell-based elements lost by 0.2 lM filtration that collaborate with soluble factors in promoting the growth of most CB cells including clonogenic progenitors and immature subpopulations. Osteoblast and M-OSTs secrete a wide array of matrix materials and ECM proteins such as collagen I, fibronectin, osteocalcin and osteopontin (Kulterer et al 2007;Neve et al 2011). ECM proteins have been implicated in the regulation of homing, survival, differentiation and proliferation of hematopoietic cells (Yokota et al 1998;Voermans et al 1999;Sagar et al 2006).…”

Histogenetic Characterization of Giant Cell Tumor of Bone

“…A 20% to 27% binuclear cell component [27] was present in SC but not in hMSC. Under basal conditions, SC and hMSC showed similar levels of ALP activity, but SC did not express Cbfa1, an early marker of osteoblastic differentiation that is always expressed in hMSC [26]. Cbfa1 is a member of the RUNX family of transcription factors with a highly restricted tissue expression pattern in bone.…”

“…Interestingly, ICAM-1 plays a pivotal role in osteoclastogenesis, because osteoblasts expressing ICAM-1 support bone resorption activity [44,45]. mRNA semiquantitative analysis for ICAM-1 suggests this adhesion molecule is expressed in tissue samples of GCT, and SC tend to show a higher expression of ICAM-1 as compared with both undifferentiated hMSC (T0) and differentiated osteoblasts (T1) [26]. The hypothesis that SC are similar to the bone lining cell phenotype deserves additional investigation.…”