Heparan sulfate regulates the anabolic activity of MC3T3‐E1 preosteoblast cells by induction of Runx2

Abstract

The transcription factor Runx2 can be controlled by a number of upstream regulators involved in intracellular signaling, including the activation ERK1/2 signaling by fibroblast growth factor-2 (FGF-2). FGFs interact with their cell surface receptors (FGFRs) through an obligate cross-binding interaction with heparan sulfate proteoglycan (HSPG) co-receptors; exogenous HS sugar chains have been shown to potently modulate changes in cell phenotype depending on the stage of tissue differentiation when the HS is harvested, suggesting that HS chain structure and function varies depending on the stage of cell maturity. This study examined the potential of bone-derived heparan sulfate (HS), harvested from differentiating osteoblasts, for the enhancement of preosteoblast growth and differentiation. HS was harvested from conditioned media, cell surface and matrix compartments of postconfluent (differentiating) MC3T3-E1 osteoblasts and dosed back onto preconfluent MC3T3-E1 cells. We show that HS can increase the expression Runx2, ALP, and OPN in preosteoblast cells, suggesting the potential for exogenous HS to shift cells from proliferative to differentiative phenotypes. In line with their structural differences, only HS released into the media was found to co-stimulate the mitogenic effect of FGF-2, whilst exogenous application of all the HSs together with FGF-2 served to increase the expression of OPN. Only the application of cell surface-derived HS triggered a synergistic increase in FGFR1 expression together with FGF-2, although all three HS preparations could trigger transient increases in PI3K, ERK1/2, and stat3 phosphorylation levels. These findings demonstrate that the compartmentally distinct HS species expressed by differentiating MC3T3-E1 cells act in complex ways to coordinate the extracellular conditions that lead to osteoblast differentiation, with the cell surface species coordinating the FGF response. J. Cell. Physiol. 210: 38-50, 2007. ß 2006 The orderly progression of bone stem cells through their precursor stages into mature phenotypes requires a systematic cascade of tissue-specific transcription factors to coordinate the process. Of these transcription factors, Runx2, a member of the runt-related family of transcription factors, has been shown to be essential for both skeletal patterning during embryogenesis and the progression of osteoblast differentiation (Ducy et al

Synergism between Wnt3a and Heparin Enhances Osteogenesis via a Phosphoinositide 3-Kinase/Akt/RUNX2 Pathway

“…Heparan sulfate proteoglycans, which are abundant on the cell surface and in the extracellular matrix, have emerged as major co-regulators controlling signaling pathways in osteogenic cells (44). Heparan sulfates increase the expression of bone-related biomarkers and maintain BMP signaling, and studies have suggested that this family of sugar-based compounds orchestrates the transition from proliferation to differentiation (3,33,44). Supporting this view, our study demonstrates that heparin strongly synergizes with Wnt3a and selectively stimulates the osteogenic but not mitogenic potential of Wnt3a.…”

“…At present, the in vivo efficacy of bone anabolic factors (e.g. BMP2, 3 parathyroid hormone, and Wnts) is limited, and new molecular approaches to improve the effectiveness and selectivity of osteo-inductive extracellular ligands are desirable. Results from our group and others suggest that glycosaminoglycans (GAGs) and in particular heparan sulfate (HS) and heparin are potent co-stimulators of osteogenic signaling pathways, including BMP2-Smads and FGF-FGFR-MAPK (1-7).…”

“…Heparin-growth factor interactions control the activities of susceptible ligands and in particular their interactions with cognate receptors, so dictating mesenchymal lineage progression and cell behavior. Heparin (and the less sulfated heparan sulfates) facilitates the binding of FGF2 to FGFRs, so augmenting osteoblast proliferation and differentiation (1)(2)(3)(4)(5). These GAGs also enhance the activity of BMPs, osteoinductive growth factors that have been clinically approved for spinal fusion, in part by protecting them from enzymatic attack or structural antagonists (6,7).…”

Fibroblast growth factor signaling in skeletal development and disease

“…Early studies showed that some HSPGs, such as syndecans, expressed in the bone environment affect the osteoblast response to FGF2 during osteogenesis (Molteni et al 1999a). In vitro data indicate that both cell surface and secreted HSPGs can further amplify FGF2 signaling and osteoblast differentiation (Jackson et al 2007). Notably, the HSPG glypican-3 was found to be essential for Runx2 expression and subsequent osteoblast differentiation in vitro ).…”

Comparative Assessment of the Effects of Gender-specific Heparan Sulfates on Mesenchymal Stem Cells

“…We have previously shown that HS helps control the osteogenic differentiation of preosteoblast MC3T3-E1 cells (12) and that exogenous application of such HS to cultures of rat bone marrow stem cells can stimulate their proliferation, which then leads to an increased expression of osteogenic markers and enhanced bone nodule formation (13). In recent years human mesenchymal stem cells (hMSCs) have been shown to be an important source of cells for tissue engineering applications.…”