Osteoblast differentiation and a transcriptional factor, Cbfa1

Cbfa1 is a member of runt-domain gene family. Cbfa1-deficient mice lack of bone formation and introduction of Cbfa1 into non-osteoblastic cells induced the expression of osteoblast specific genes. Cbfa1-deficient calvarial cells were composed of immature mesenchymal cells and can differentiated into adipocytes and chondrocytes. Based on these findings, Cbfa1 has been identified as a master regulatory gene of the osteoblast differentiation and it has been suggested that Cbfa1 plays an essential role in determining the lineage of multipotential mesenchymal precursor cells.     

促进成骨细胞分化和骨生成的多重信号通路及相关节点蛋白

胚胎时期骨形成、出生后骨塑建及成人时期的骨重建过程中,均伴随着骨质生成现象。成骨细胞是骨质形成的主要功能细胞,负责骨基质的合成、分泌和矿化。成骨细胞分化和骨生成是受多种信号蛋白和转录因子调节的多步骤分子事件。本文主要对参与成骨细胞分化和骨生成的多重信号通路进行综述,包括BMPs通路、Wnt—B—Catenin通路、MAPK通路、Hedgehog通路、NF—KB通路、PTH／PTHrP通路及Insulin Receptor通路等,并介绍了几种相关节点蛋白,包括Runx2、Osterix和同源结构域蛋白等。当前,骨质疏松等骨相关疾病已经成为危害人类健康的突出问题,对调节成骨分化和骨生成信号通路及相关节点蛋白的研究能为开发预防和治疗骨相关疾病药物据供恩足箨和箫田备．     

Bone morphogenetic proteins stimulate angiogenesis through osteoblast-derived vascular endothelial growth factor A

During bone formation and fracture healing there is a cross-talk between endothelial cells and osteoblasts. We previously showed that vascular endothelial growth factor A (VEGF-A) might be an important factor in this cross-talk, as osteoblast-like cells produce this angiogenic factor in a differentiation-dependent manner. Moreover, exogenously added VEGF-A enhances osteoblast differentiation. In the present study we investigated, given the coupling between angiogenesis and bone formation, whether bone morphogenetic proteins (BMPs) stimulate osteoblastogenesis and angiogenesis through the production of VEGF-A. For this we used the murine preosteoblast-like cell line KS483, which forms mineralized nodules in vitro, and an angiogenesis assay comprising 17-d-old fetal mouse bone explants that have the ability to form tube-like structures in vitro. Treatment of KS483 cells with BMP-2, -4, and -6 enhanced nodule formation, osteocalcin mRNA expression, and subsequent mineralization after 18 d of culture. This was accompanied by a dose-dependent increase in VEGF-A protein levels throughout the culture period. BMP-induced osteoblast differentiation, however, was independent of VEGF-A, as blocking VEGF-A activity by a VEGF-A antibody or a VEGF receptor 2 tyrosine kinase inhibitor did not affect BMP-induced mineralization. To investigate whether BMPs stimulate angiogenesis through VEGF-A, BMPs were assayed for their angiogenic activity. Treatment of bone explants with BMPs enhanced angiogenesis. This was inhibited by soluble BMP receptor 1A or noggin. In the presence of a VEGF-A antibody, both unstimulated and BMP-stimulated angiogenesis were arrested. Conditioned media of KS483 cells treated with BMPs also induced a strong angiogenic response, which was blocked by antimouse VEGF-A but not by noggin. These effects were specific for BMPs, as TGF beta inhibited osteoblast differentiation and angiogenesis while stimulating VEGF-A production. These findings indicate that BMPs stimulate angiogenesis through the production of VEGF-A by osteoblasts. In conclusion, VEGF-A produced by osteoblasts in response to BMPs is not involved in osteoblast differentiation, but couples angiogenesis to bone formation.     

Osteoblasts and bone formation

Bone is constantly being remodelled in a dynamic process where osteoblasts are responsible for bone formation and osteoclasts for its resorption. Osteoblasts are specialized mesenchymal cells that undergo a process of maturation where genes like core-binding factor alpha1 (Cbfa1) and osterix (Osx) play a very important role. Moreover, it was found recently that Wnt/ beta-catenin pathway plays a part on osteoblast differentiation and proliferation. In fact, mutations on some of the proteins involved in this pathway, like the low-density lipoprotein receptor related protein 5/6 (LRP5/6) lead to bone diseases. Osteoblast have also a role in regulation of bone resorption through receptor activator of nuclear factor-kappaB (RANK) ligand (RANKL), that links to its receptor, RANK, on the surface of pre-osteoblast cells, inducing their differentiation and fusion. On the other hand, osteoblasts secrete a soluble decoy receptor (osteoprotegerin, OPG) that blocks RANK/RANKL interaction by binding to RANKL and, thus, prevents osteoclast differentiation and activation. Therefore, the balance between RANKL and OPG determines the formation and activity of osteoclasts. Another factor that influences bone mass is leptin, a hormone produced by adipocytes that have a dual effect. It can act through the central nervous system and diminish osteoblasts activity, or can have an osteogenic effect by binding directly to its receptors on the surface of osteoblast cells.     

Extracellular matrix-associated bone morphogenetic proteins are essential for differentiation of murine osteoblastic cells in vitro

Osteoblastic differentiation is an essential part of bone formation that compensates resorbed bone matrix to maintain its structural integrity. Cells in an osteoblast lineage develop differentiated phenotypes during a long-term culture in vitro. However, intrinsic mechanisms whereby these cells differentiate into mature osteoblasts are yet unclear. Bone morphogenetic proteins (BMPs) stimulate osteoblastic differentiation and bone formation. We demonstrate that mouse osteoblastic MC3T3-E 1 cells constitutively expressed messenger RNAs (mRNAs) for BMP-2 and BMP-4 and accumulated BMPs in collagen-rich extracellular matrices. BMPs associated with the extracellular matrices were involved in the induction of osteoblastic differentiation of nonosteogenic mesenchymal cells as well as cells in the osteoblast lineage. MC3T3-E1 cells constitutively expressed type IA and type II BMP receptors. When a kinase-deficient type IA BMP receptor was stably transfected to MC3T3-E 1 cells to obliterate BMP-2/4 signaling, these cells not only failed to respond to exogenous BMP-2 but lost their capability of differentiation into osteoblasts that form mineralized nodules. These observations strongly suggest that endogenous BMP-2/4 accumulated in extracellular matrices are essential for the osteoblastic differentiation of cells in the osteoblast lineage. Therefore, the regulatory mechanism of BMP-2/4 actions in osteoblastic cells is a principal issue to be elucidated for better understanding of pathogenesis of bone losing diseases such as osteoporosis.