Zheng Zhang1, Weiwei Jiang2, Miao Hu1, Rui Gao3, Xuhui Zhou4. 1. Department of Orthopedics, Changzheng Hospital, Second Military Medical University, 415 Fengyang Road, Shanghai, 200003, PR China; College of Basic Medicine, Second Military Medical University, 800 Xiangyin Road, Shanghai, 200433, PR China. 2. Department of Critical Care Medicine, Changzheng Hospital, Second Military Medical University, 415 Fengyang Road, Shanghai, 200003, PR China. 3. Department of Orthopedics, Changzheng Hospital, Second Military Medical University, 415 Fengyang Road, Shanghai, 200003, PR China. 4. Department of Orthopedics, Changzheng Hospital, Second Military Medical University, 415 Fengyang Road, Shanghai, 200003, PR China. Electronic address: zhouxuhui@smmu.edu.cn.
Abstract
BACKGROUND: Dysfunction in the osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) leads to bone loss/osteoporosis. The catenin beta interacting protein 1 (CTNNBIP1) is an inhibitor of Wnt/β-catenin signaling, whose role in osteogenesis remains elusive. This study aimed to reveal the effects of miR-486-3p/CTNNBIP1 in osteogenesis. METHODS: Bone marrow samples from healthy individuals and osteoporosis patients and mice with sham or ovariectomy (OVX) surgeries were collected. Levels of CTNNBIP1 and miR-486-3p were assessed. A dual-luciferase reporter assay was used to confirm the interactions between CTNNBIP1 and miR-486-3p. MiR-486-3p mimics/inhibitor or CTNNBIP1 overexpression lentiviruses were transfected to human BMSCs (hBMSCs) and an osteogenic assay was performed. Alizarin red S (ARS) and Alkaline phosphatase (ALP) intensity and expression of osteogenic genes Runx2, Alp, Cola1 and Bglap were measured. Key proteins in the Wnt/β-catenin pathway including active β-catenin, Bcl-2, and Cyclin D1 were assessed. RESULTS: CTNNBIP1 was upregulated while miR-486-3p was downregulated in osteoporosis patients and OVX mice. CTNNBIP1 was confirmed as a target of miR-486-3p. MiR-486-3p overexpression promoted, while miR-486-3p knockdown suppressed, osteogenic differentiation and Wnt/β-catenin signaling. Rescue experiments confirmed the negative effects of CTNNBIP1 overexpression on osteoblastic differentiation and that miR-486-3p mimics could reverse canonical Wnt signaling. CONCLUSION: This study demonstrated that miR-486-3p targets CTNNBIP1, thus activating the Wnt/β-catenin signaling pathway to promote osteogenesis of BMSCs.
BACKGROUND: Dysfunction in the osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) leads to bone loss/osteoporosis. The catenin beta interacting protein 1 (CTNNBIP1) is an inhibitor of Wnt/β-catenin signaling, whose role in osteogenesis remains elusive. This study aimed to reveal the effects of miR-486-3p/CTNNBIP1 in osteogenesis. METHODS: Bone marrow samples from healthy individuals and osteoporosispatients and mice with sham or ovariectomy (OVX) surgeries were collected. Levels of CTNNBIP1 and miR-486-3p were assessed. A dual-luciferase reporter assay was used to confirm the interactions between CTNNBIP1 and miR-486-3p. MiR-486-3p mimics/inhibitor or CTNNBIP1 overexpression lentiviruses were transfected to human BMSCs (hBMSCs) and an osteogenic assay was performed. Alizarin red S (ARS) and Alkaline phosphatase (ALP) intensity and expression of osteogenic genes Runx2, Alp, Cola1 and Bglap were measured. Key proteins in the Wnt/β-catenin pathway including active β-catenin, Bcl-2, and Cyclin D1 were assessed. RESULTS:CTNNBIP1 was upregulated while miR-486-3p was downregulated in osteoporosispatients and OVX mice. CTNNBIP1 was confirmed as a target of miR-486-3p. MiR-486-3p overexpression promoted, while miR-486-3p knockdown suppressed, osteogenic differentiation and Wnt/β-catenin signaling. Rescue experiments confirmed the negative effects of CTNNBIP1 overexpression on osteoblastic differentiation and that miR-486-3p mimics could reverse canonical Wnt signaling. CONCLUSION: This study demonstrated that miR-486-3p targets CTNNBIP1, thus activating the Wnt/β-catenin signaling pathway to promote osteogenesis of BMSCs.