Relaxin Augments BMP-2–Induced Osteoblast Differentiation and Bone Formation

Abstract

Relaxin (Rln), a polypeptide hormone of the insulin superfamily, is an ovarian peptide hormone that is involved in a diverse range of physiological and pathological reactions. In this study, we investigated the effect of Rln on bone morphogenetic protein 2 (BMP-2)-induced osteoblast differentiation and bone formation. Expression of Rln receptors was examined in the primary mouse bone marrow stem cells (BMSCs) and mouse embryonic fibroblast cell line C3H/10T1/2 cells by RT-PCR and Western blot during BMP-2-induced osteoblast differentiation. The effect of Rln on osteoblast differentiation and mineralization was evaluated by measuring the alkaline phosphatase activity, osteocalcin production, and Alizarin red S staining. For the in vivo evaluation, BMP-2 and/or Rln were administered with type I collagen into the back of mice, and after 3 weeks, bone formation was analyzed by micro-computed tomography (mCT). Western blot was performed to determine the effect of Rln on osteoblast differentiation-related signaling pathway. Expression of Rxfp 1 in BMSCs and C3H/10T1/2 cells was significantly increased by BMP-2. In vitro, Rln augmented BMP-2-induced alkaline phosphatase expression, osteocalcin production, and matrix mineralization in BMSCs and C3H/10T1/2 cells. In addition, in vivo administration of Rln enhanced BMP-2-induced bone formation in a dose-dependent manner. Interestingly, Rln synergistically increased and sustained BMP-2-induced Smad, p38, and transforming growth factor-b activated kinase (TAK) 1 phosphorylation. BMP-2-induced Runx 2 expression and activity were also significantly augmented by Rln. These results show that Rln enhanced synergistically BMP-2-induced osteoblast differentiation and bone formation through its receptor, Rxfp 1, by augmenting and sustaining BMP-2-induced Smad and p38 phosphorylation, which upregulate Runx 2 expression and activity. These results suggest that Rln might be useful for therapeutic application in destructive bone diseases.

The Importance of the Prenyl Group in the Activities of Osthole in Enhancing Bone Formation and Inhibiting Bone ResorptionIn Vitro

“…BMP-2 [31] is known to induce mesenchymal cells to differentiate into osteogenic cells, and osteocalcin [32] and OPN [33], often used as the biochemical markers for bone formation, are known also to play important roles in the differentiation process of osteoblasts. As shown in Figures 7(b)–7(d) and Figures 8(b)–8(d), trends of changes of conditioned medium levels during the osteogenic culture time course are similar for BMP-2, osteocalcin, and OPN in both rBMSCs and ROB.…”

Relaxin Activates Peroxisome Proliferator-activated Receptor γ (PPARγ) through a Pathway Involving PPARγ Coactivator 1α (PGC1α)

“…There have been few studies of the role of p38 MAPK in mediating relaxin effects. In human periodontal ligament cells, relaxin regulation of matrix metalloproteinase expression was reduced by a p38 MAPK inhibitor (37), and relaxin potentiated BNP2-mediated p38 MAPK activation in multipotent mesenchymal cells (38). Relaxin induced activation of p38 MAPK in vascular smooth muscle cells, but not HeLa or vascular endothelial cells (39) or renal fibroblasts (40).…”

Relaxin enhances bone regeneration with BMP‐2‐loaded hydroxyapatite microspheres

“…The amount of new bone in defects implanted with the HA microspheres loaded with 0.5 and 1 μg BMP‐2 per defect was significantly higher than that in defects loaded with the as‐prepared microspheres (i.e., with no BMP‐2 loaded) (Figure a). In comparison, relaxin is known to be an enhancer of BMP‐2 and does not have any osteoinductive properties on its own (Moon et al, ). In the present study, loading HA microspheres with relaxin alone (0.05 to 0.25 μg per defect) seemed to produce bone regeneration not statistically different from the as‐prepared microspheres (Figure and b; Table ).…”

“…It is typically known as a pregnancy hormone, in which it has a role in promoting cervical softening to facilitate birth (Bathgate et al, ). Relaxin also plays a part in connective tissue metabolism, collagen turnover, angiogenesis, and tumor metastasis (Bathgate et al, ; Moon et al, ). Relaxin‐2, the only relaxin protein found circulating in the blood, acts equally through the relaxin/insulin‐like family peptide receptors (Rxfp) 1 and 2, although Rxfp2 is known to play a larger role in osteogenesis (Bathgate et al, ; Duarte, Kobayashi, Morita, Kawamoto, & Moriyama, ).…”