Bone development and fracture repair include the coordination of multiple events such as migration, differentiation, and activation of multiple cell types and tissues. Angiogenesis plays a critical role in bone development and bone fracture repair. Angiopoietin 1 (Ang1) is an endothelial growth fact...
Bone development and fracture repair include the coordination of multiple events such as migration, differentiation, and activation of multiple cell types and tissues. Angiogenesis plays a critical role in bone development and bone fracture repair. Angiopoietin 1 (Ang1) is an endothelial growth factor essential for embryonic vascular development that generates a stable and functional vasculature via binding to the tyrosine kinase with Ig and epidermal growth factor homology domain 2 (Tie2) receptor and induces Tie2 phosphorylation. Recently, cartilage oligomeric matrix protein (COMP)-Ang1 was developed by replacing the N-terminal portion of Angiopeotin 1 with the short coiled-coil domain of cartilage oligomeric matrix protein, a chimeric protein of angiopoietin-1. COMP-Ang1 is known to have more soluble, stable and potent than did the native Ang1. The periodontal ligament cells (PDLCs) have different tendencies for osteogenesis, chondrogenesis, adipogenesis, or proliferation. The PDLCs have osteoblast-like properties, including the capacity to form mineralized nodules in vitro. A diversity of cellular characteristics can be controlled in part by the addition of cytokines, growth factors, or drugs. Therefore, understanding the exact roles of these factors and identifying the signal transduction pathways involved are to be the crucial events in controlling a balanced cellular differentiation. The main aims of this study were to evaluate the osteogenic capacity of COMP-Ang1 in bone tissue and to determine the osteogenic activity of COMP-Ang1 in vitro by estimating its effect on osteoblast differentiation using human PDLCs. In order to compare the phenotypes of wild type (WT) and COMP-Ang1 tissue-specific inducible (MT) mice in bones, this study used ovariectomy osteoporosis and calvarial bone defect models. The osteogenic capacity of COMP-Ang1 in bone tissue in vivo was also investigated using an 8 mm critical-sized calvarial defect model. Bone structure was evaluated using micro-computed tomography (CT) software, which calculates morphometirc parameters from a selected volume of interest (VOI). To confirm whether COMP-Ang1 regulates the expression of angiogenic and osteogenic genes, this study measured the mRNA expression levels in the scaffolds inserted in a rat calvarial defect model. In vitro differentiation potential of COMP-Ang1 recombinant protein was evaluated. In addition, this study evaluated the effect of COMP-Ang1 on migration using human PDLCs. As the results, MT mice did not show any increase in bone formation, compared to WT at a significant level. In contrast, the values of micro-CT revealed that MT group resulted in a significant increase in new bone formation, compared to control group in rats. Further, the expression levels of osteogenic genes, such as Runx2, Osx, and Bmp-2, and angiogenic genes, such as Pecam1 and Lyve1, were significantly up-regulated in the COMP–Ang1 group. Stimulating rat bone marrow derived cells (BMCs) with COMP-Ang1 up-regulated the expression of Runx2, Osx, and a stromal cell-derived factor (SDF-1) receptor, CXCR4 in a dose dependent manner. It was found that human PDLCs were migrated into COMP-Ang1-loaded scaffold and recruited through COMP-Ang1-mediated up-regulation of the SDF-1. Quantitative real-time RT–PCR revealed that the genes specific for c-Jun, FosB, and Fra1 were significantly induced by treatment with COMP-Ang1. These results suggest that COMP–Ang1 accelerates new bone formation through stimulation of osteogenesis and angiogenesis as well as the migration of progenitor cells. Collectively, these results demonstrate that COMP-Ang1 has a striking ability to stimulate bone formation, suggesting that COMP-Ang1 may be a useful as a therapeutic protein for bone repair.
Bone development and fracture repair include the coordination of multiple events such as migration, differentiation, and activation of multiple cell types and tissues. Angiogenesis plays a critical role in bone development and bone fracture repair. Angiopoietin 1 (Ang1) is an endothelial growth factor essential for embryonic vascular development that generates a stable and functional vasculature via binding to the tyrosine kinase with Ig and epidermal growth factor homology domain 2 (Tie2) receptor and induces Tie2 phosphorylation. Recently, cartilage oligomeric matrix protein (COMP)-Ang1 was developed by replacing the N-terminal portion of Angiopeotin 1 with the short coiled-coil domain of cartilage oligomeric matrix protein, a chimeric protein of angiopoietin-1. COMP-Ang1 is known to have more soluble, stable and potent than did the native Ang1. The periodontal ligament cells (PDLCs) have different tendencies for osteogenesis, chondrogenesis, adipogenesis, or proliferation. The PDLCs have osteoblast-like properties, including the capacity to form mineralized nodules in vitro. A diversity of cellular characteristics can be controlled in part by the addition of cytokines, growth factors, or drugs. Therefore, understanding the exact roles of these factors and identifying the signal transduction pathways involved are to be the crucial events in controlling a balanced cellular differentiation. The main aims of this study were to evaluate the osteogenic capacity of COMP-Ang1 in bone tissue and to determine the osteogenic activity of COMP-Ang1 in vitro by estimating its effect on osteoblast differentiation using human PDLCs. In order to compare the phenotypes of wild type (WT) and COMP-Ang1 tissue-specific inducible (MT) mice in bones, this study used ovariectomy osteoporosis and calvarial bone defect models. The osteogenic capacity of COMP-Ang1 in bone tissue in vivo was also investigated using an 8 mm critical-sized calvarial defect model. Bone structure was evaluated using micro-computed tomography (CT) software, which calculates morphometirc parameters from a selected volume of interest (VOI). To confirm whether COMP-Ang1 regulates the expression of angiogenic and osteogenic genes, this study measured the mRNA expression levels in the scaffolds inserted in a rat calvarial defect model. In vitro differentiation potential of COMP-Ang1 recombinant protein was evaluated. In addition, this study evaluated the effect of COMP-Ang1 on migration using human PDLCs. As the results, MT mice did not show any increase in bone formation, compared to WT at a significant level. In contrast, the values of micro-CT revealed that MT group resulted in a significant increase in new bone formation, compared to control group in rats. Further, the expression levels of osteogenic genes, such as Runx2, Osx, and Bmp-2, and angiogenic genes, such as Pecam1 and Lyve1, were significantly up-regulated in the COMP–Ang1 group. Stimulating rat bone marrow derived cells (BMCs) with COMP-Ang1 up-regulated the expression of Runx2, Osx, and a stromal cell-derived factor (SDF-1) receptor, CXCR4 in a dose dependent manner. It was found that human PDLCs were migrated into COMP-Ang1-loaded scaffold and recruited through COMP-Ang1-mediated up-regulation of the SDF-1. Quantitative real-time RT–PCR revealed that the genes specific for c-Jun, FosB, and Fra1 were significantly induced by treatment with COMP-Ang1. These results suggest that COMP–Ang1 accelerates new bone formation through stimulation of osteogenesis and angiogenesis as well as the migration of progenitor cells. Collectively, these results demonstrate that COMP-Ang1 has a striking ability to stimulate bone formation, suggesting that COMP-Ang1 may be a useful as a therapeutic protein for bone repair.
주제어
#Angiopoietin1, COMP-Ang1, hPDLCs, scaffold, bone regeneration, osteogenesis
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