TGF beta-2 signaling is essential for osteoblast migration and differentiation during fracture healing in medaka fish

摘要

TGF beta is known as a canonical coupling factor based on its effects on bone formation and bone resorption. There are 3 different isoforms of it related to bone metabolism in mammals. TGF beta function in vivo is complicated, and each isoform shows a different function. Since TGF beta s are secreted during inflammation accompanied by the release of latent TGF beta from inside of the bones where they are stored in the extracellular matrix, TGF beta function is potentially related to fracture healing. Although a few reports examined the TGF beta expression during fracture healing, the function of TGFO in this process is poorly understood. To investigate TGF beta function during fracture healing in vivo, we used the fracture healing model of the medaka fish, which enabled us to observe the behavior and function of living cells in response to a bone-specific injury. RNA in-situ hybridization analysis showed that only tgf beta-2 of the 4 TGF13 isoforms in medaka was expressed in the bone-forming region. To examine the TGF beta-2 function for bone formation by osteoblasts, we used a medaka transgenic line, Tg (type X collagen: GFP); and the results revealed that type X collagen-positive immature osteoblasts migrated to the fracture site and differentiated toosterix-positive osteoblasts. However, only a few type X collagen-positive osteoblasts exhibited BrdU incorporation after the fracture. Then we inhibited TGFS signaling by using a chemical TGF beta receptor kinase inhibitor (5B431542), and demonstrated that inhibition of TGF13 strongly impaired osteoblast migration and differentiation. In addition, this TGF beta inhibitor reduced the RANKL expression and caused a delay of osteoclast differentiation. Our findings thus demonstrated that TGF beta-2 functioned specifically during fracture healing to stimulate the migration of osteoblasts as well as the differentiation of osteoblasts and osteoclasts. (C) 2016 Elsevier Inc. All rights reserved.