?-Catenin and BMP-2 synergize to promote osteoblast differentiation and new bone formation

Abstract

Mutations of critical components of the Wnt pathway profoundly affect skeletal development and maintenance, probably via modulation of β-catenin signaling. We tested the hypothesis that β-catenin is involved in mesenchymal lineage allocation to osteogenic cells using a β-catenin mutant with constitutive transcriptional activity (ΔN151). Although this stable β-catenin had no effects by itself on osteogenic differentiation of multipotent embryonic cell lines, it synergized with bone morphogenetic protein-2 (BMP-2) resulting in dramatic stimulation of alkaline phosphatase activity, osteocalcin gene expression, and matrix mineralization. Likewise, ΔN151 and BMP-2 synergistically stimulated new bone formation after subperiosteal injection in mouse calvaria in vivo. Conversely, ΔN151 prevented adipogenic differentiation from pre-adipocytic or uncommitted mesenchymal cells in vitro. Intriguingly, the synergism with BMP-2 on gene transcription occurred without altering expression of Cbfa1/Runx2, suggesting actions independent or downstream of this osteoblastspecific transcription factor. Thus, β-catenin directs osteogenic lineage allocation by enhancing mesenchymal cell responsiveness to osteogenic factors, such as BMP-2, in part via Tcf/Lef dependent mechanisms. In vivo, this synergism leads to increased new bone formation. Keywords cell-cell adhesion; mesenchymal differentiation; bone formation; adipogenesisIn adult life, the skeleton is constantly remodeled to replace aging tissue and repair injuries by successive phases of osteoclast bone resorption and osteoblast mediated bone formation. Therefore, a continuous supply of bone forming cells is required to maintain bone homeostasis. Bone marrow mesenchymal stem cells are the source of osteoprogenitors in adult life, although these cells can also differentiate into adipocytes and myocytes in the presence of appropriate stimuli [Pittenger et al., 1999]. Lineage allocation of mesenchymal stem cells to either osteogenic or adipogenic cells must be kept in a critical balance, and shifts that may favor one lineage over the other may have important consequences for an individual's ability to produce sufficient numbers of bone forming cells. An osteogenic to adipogenic shift may explain the pathogenesis of age-dependent bone loss, which is associated with a reduced potential of bone marrow to produce osteogenic cells in the face of an increased number of adipocytes [Jilka et al., 1996;Gimble and Nuttall, 2004]. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author ManuscriptThe osteogenic differentiation program is under the control of hormonal and local factors converging onto a finite number of transcriptional regulators that ultimately determine the fate of cells committing to the osteogenic lineage [Karsenty and Wagner, 2002]. An emerging body of work demonstrates that the Wnt signaling system is one of the most important local regulators of bone formation, presumably via activation of the β-catenin signaling system. Wnts are a family of secreted protei...

Wnt signaling and skeletal development

“…It was shown that stable β-catenin is able to induce ALP activity in C3H10T1/2 cells by transient transfection [53], enhance osteoblastogenesis (by retroviral infection) in both C3H10T1/2 and ST2 cells [89], and dramatically stimulate osteoblastogenesis by synergizing with BMP-2 in C3H10T1/2 cells via retroviral infection [92]. A study showed that retroviral expression of a stabilized form of β-catenin in C3H10T1/2 cells induced expression of ALP mRNA and protein (but not osteocalcin expression) in non-differentiated medium [93].…”

“…A study showed that retroviral expression of a stabilized form of β-catenin in C3H10T1/2 cells induced expression of ALP mRNA and protein (but not osteocalcin expression) in non-differentiated medium [93]. Another study showed that constitutively active β-catenin has no effect on basal ALP activity in either C3H10T1/2 or C2C12 cells [92]. Deletion of β-catenin by using adenovirus expressing the Cre recombinase in calvarial osteoblasts resulted in delayed and diminished expression of osteocalcin as well as in significantly reduced mineralization [94].…”

Housekeeping gene stability influences the quantification of osteogenic markers during stem cell differentiation to the osteogenic lineage

“…To address this issue, this work aimed to study the stability of two of the most commonly used housekeeping genes in osteogenic differentiation studies of stem cells: ACTB and GAPDH (Vandesompele et al 2002;Cho et al 2005;Mbalaviele et al 2005), in hBMSCs undergoing osteogenic differentiation. The results were compared with the stability of the gene ribosomal protein L13A (RPL13A), which has been shown to have stable expression levels in bone marrow tissues (Vandesompele et al 2002).…”

Synergistic and Multidimensional Regulation of Plasminogen Activator Inhibitor Type 1 Expression by Transforming Growth Factor Type β and Epidermal Growth Factor

“…EGF and TGF␤ synergize to regulate several important cellular processes, including colony formation in soft agar (24), the epithelial to mesenchymal transition (25), and cell movement during tissue remodeling (26). The ability to tightly control gene expression by the synergistic interaction of extracellular regulators, such as growth factors, is likely to be an important aspect of gene regulation in vivo and has already been demonstrated for some systems (27)(28)(29).…”