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Thrombomodulin Functional Domains Support Osteoblast Differentiation and Bone Healing in Diabetes in Mice
Chung-Hwan Chen1,2,3,4,5,6, Chao-Han Lai7, Yi-Kai Hong8
1Orthopaedic Research Center, Kaohsiung Medical University, Kaohsiung, Taiwan.
Abstract:
Thrombomodulin (TM) is a transmembrane glycoprotein that contains five functional domains. Soluble TM (sTM), comprising extracellular domains TMD1 (lectin-like), TMD2 (epidermal growth factor [EGF]-like repeat containing), and TMD3 (serine-threonine rich), can be shed from cells by the intramembrane protease rhomboid-like-2 (RHBDL2). TM is expressed by osteoblasts, yet its role there has not been determined. Herein we aimed to investigate the properties of TM and its domains in osteoblast function and bone repair following injury in diabetes. In response to a scratch injury of cultured osteoblast-like MG63 cells, expression of TM and RHBDL2 was enhanced, with increased release of sTM. Conditioned media from the injured cells promoted osteoblast migration, an effect that was lacking with conditioned media from MG63 cells in which TM was silenced by shRNA. Exogenous recombinant TMD1 had no effect on osteoblast activities or on bone repair in vivo. However, TM domains 2 and 3 (TMD2/3), induced MG63 cell migration, proliferation and mineralization in vitro, and when locally administered in mice, improved in vivo healing of injured calvarium. This beneficial effect of TMD2/3, mediated via fibroblast growth factor receptor (FGFR)/ERK signaling pathways, was also observed in vitro under high glucose conditions where endogenous TM expression was reduced, and in vivo in diabetic mice following tibia fracture or calvarium injury, where the osteoblastic response and healing were otherwise dampened. Taken together, osteoblast TM participates in bone healing, and recombinant TMD2/3 holds promise as a novel therapy for diabetic bone defect healing. © 2020 American Society for Bone and Mineral Research.
Insights
Thrombomodulin (TM) domains 2 and 3 (TMD2/3) enhance osteoblast function and promote bone healing, offering a potential therapy for diabetic bone defects.
Area of Science:
- Biochemistry
- Cell Biology
- Orthopedics
Background:
- Thrombomodulin (TM) is a transmembrane glycoprotein with known roles in coagulation.
- The function of TM in osteoblasts and its role in bone repair, especially in diabetes, remain unclear.
- Soluble TM (sTM) can be shed from cells, and its specific domains may possess distinct biological activities.
Purpose of the Study:
- To investigate the role of osteoblast Thrombomodulin (TM) and its domains in osteoblast function and bone repair.
- To determine the therapeutic potential of TM domains in healing bone defects, particularly in the context of diabetes.
- To elucidate the signaling pathways involved in TM-mediated effects on osteoblasts.
Main Methods:
- Utilized cultured osteoblast-like MG63 cells subjected to scratch injury to assess TM and RHBDL2 expression and sTM release.
- Employed shRNA to silence TM expression and assessed the impact on conditioned media effects.
- Administered recombinant TM domains (TMD1, TMD2/3) to cultured cells and in vivo mouse models (calvarium and tibia fracture) to evaluate effects on migration, proliferation, mineralization, and bone healing.
- Investigated signaling pathways including fibroblast growth factor receptor (FGFR)/ERK.
Main Results:
- Scratch injury to MG63 cells increased TM and RHBDL2 expression and sTM release.
- Conditioned media from injured cells promoted osteoblast migration, an effect dependent on TM.
- Recombinant TMD2/3, but not TMD1, enhanced osteoblast migration, proliferation, and mineralization in vitro.
- Local administration of TMD2/3 improved calvarium bone healing in mice.
- TMD2/3 effects were mediated via FGFR/ERK signaling and were effective in high glucose conditions and in diabetic mice with bone injuries.
Conclusions:
- Osteoblast-expressed Thrombomodulin (TM) plays a significant role in bone healing processes.
- Recombinant TM domains 2 and 3 (TMD2/3) demonstrate therapeutic potential for enhancing bone repair.
- TMD2/3 offers a promising novel therapeutic strategy for treating bone defects in diabetic patients, addressing impaired healing.
- The FGFR/ERK pathway is implicated in the beneficial effects of TMD2/3 on osteoblast function and bone repair.
