Fouad M Moussa1, Bryson P Cook2, Greg R Sondag1, Matthew DeSanto2, Mark S Obri2, Scott E McDermott3, Fayez F Safadi4. 1. Department of Anatomy and Neurobiology, Northeast Ohio Medical University (NEOMED), College of Medicine, Rootstown, OH, United States of America; Musculoskeletal Research Group, NEOMED, Rootstown, OH, United States of America; School of Biomedical Sciences, Kent State University, Kent, OH, United States of America. 2. Department of Anatomy and Neurobiology, Northeast Ohio Medical University (NEOMED), College of Medicine, Rootstown, OH, United States of America; Musculoskeletal Research Group, NEOMED, Rootstown, OH, United States of America. 3. Department of Orthopaedics, SUMMA Health System, Rebecca D. Considine Research Institute, Akron Children's Hospital, Akron, OH, United States of America. 4. Department of Anatomy and Neurobiology, Northeast Ohio Medical University (NEOMED), College of Medicine, Rootstown, OH, United States of America; Musculoskeletal Research Group, NEOMED, Rootstown, OH, United States of America; School of Biomedical Sciences, Kent State University, Kent, OH, United States of America; Department of Orthopaedics, SUMMA Health System, Rebecca D. Considine Research Institute, Akron Children's Hospital, Akron, OH, United States of America. Electronic address: fsafadi@neomed.edu.
Abstract
BACKGROUND: mir-RNAs play a role in regulating bone homeostasis. In this study we assessed the functional role of mir-RNA 150 in bone homeostasis. We also assess the effects of miR-150 deficiency on osteoblast and osteoclast differentiation and function using in vivo and in vitro approaches. METHODS: Wild type (WT) (C57BL/6J) and miR-150 KO mice were compared for a variety of parameters. Micro-CT imaging was conducted to quantify trabecular bone mass inferior to the distal growth plate of the femur. Von Kossa staining was performed for osteoblast culture mineralization. RT-qPCR, biochemical analysis and bone histomorphometry were utilized for quantification of relevant genes and serum protein measurements. Differentiation and function of osteoblasts and osteoclasts was performed using primarily cultures and assessed the cell autonomous response of mir-RNA-150 on cell differentiation and function. RESULTS: Mir-150 exhibited expression in a variety of tissues and increases progressively with age. Through micro-CT imaging, we found that KO mice presented reduced bone mass at 4, 8, and 16 weeks of age compared to WT mice. Furthermore, histomorphometric analysis revealed increased trabecular separation, decreased bone thickness, and decreased osteoblast number in KO compared to WT mice. Mir-150 deficiency also correlated with higher bone resorption, accompanied with significant increases in CTX-1 serum levels, and a decrease in cell apoptotic rate ex vivo. Additionally, miR-150 KO mice showed increased osteoblast differentiation and decreased osteoclastogenesis ex vivo. Luciferase assay showed increased Osteoactivin/GPNMB expression in miR-150 KO osteoblasts compared to WT cells. CONCLUSION: Our data suggests that miR-150 influences osteoblast and osteoclast functionality and differentiation; specifically, miR-150 serves as a negative regulator for osteoblasts and a positive regulator for osteoclasts by regulating, at least in part, Osteoactivin/GPNMB expression.
BACKGROUND: mir-RNAs play a role in regulating bone homeostasis. In this study we assessed the functional role of mir-RNA 150 in bone homeostasis. We also assess the effects of miR-150 deficiency on osteoblast and osteoclast differentiation and function using in vivo and in vitro approaches. METHODS: Wild type (WT) (C57BL/6J) and miR-150 KO mice were compared for a variety of parameters. Micro-CT imaging was conducted to quantify trabecular bone mass inferior to the distal growth plate of the femur. Von Kossa staining was performed for osteoblast culture mineralization. RT-qPCR, biochemical analysis and bone histomorphometry were utilized for quantification of relevant genes and serum protein measurements. Differentiation and function of osteoblasts and osteoclasts was performed using primarily cultures and assessed the cell autonomous response of mir-RNA-150 on cell differentiation and function. RESULTS: Mir-150 exhibited expression in a variety of tissues and increases progressively with age. Through micro-CT imaging, we found that KO mice presented reduced bone mass at 4, 8, and 16 weeks of age compared to WT mice. Furthermore, histomorphometric analysis revealed increased trabecular separation, decreased bone thickness, and decreased osteoblast number in KO compared to WT mice. Mir-150 deficiency also correlated with higher bone resorption, accompanied with significant increases in CTX-1 serum levels, and a decrease in cell apoptotic rate ex vivo. Additionally, miR-150 KO mice showed increased osteoblast differentiation and decreased osteoclastogenesis ex vivo. Luciferase assay showed increased Osteoactivin/GPNMB expression in miR-150 KO osteoblasts compared to WT cells. CONCLUSION: Our data suggests that miR-150 influences osteoblast and osteoclast functionality and differentiation; specifically, miR-150 serves as a negative regulator for osteoblasts and a positive regulator for osteoclasts by regulating, at least in part, Osteoactivin/GPNMB expression.