De-Yu Duan1, Jing Tang2, Hong-Tao Tian1, Yang-Yang Shi1, Jie Jia3. 1. Department of Orthopedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, Hubei Province, PR China. 2. Cancer Center, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, Hubei Province, PR China. 3. Department of Orthopedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, Hubei Province, PR China. Electronic address: jjwhunion@163.com.
Abstract
AIMS: Adipocyte-secreted microvesicles (MVs)-derived microRNAs (miRNAs) are relevant to adipogenic and osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) in osteonecrosis of the femoral head (ONFH). Our aims are to investigate the mechanism of adipocyte-derived MVs-miR-148a in ONFH. MATERIALS AND METHODS: Adipocyte-derived MVs were identified via transmission electron microscopy and specific markers expression. The adipogenic and osteogenic differentiation were investigated by Oil-Red O staining, alkaline phosphatase (ALP) activity, Alizarin Red S (ARS) staining and osteogenic or adipogenic factors levels. Genes and proteins expression were detected by using quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting. The relationship between miR-148a and Wnt5a was tested via dual-luciferase reporter analysis. The adipogenic differentiation and osteogenic differentiation in methylprednisolone (MPS)-induced ONFH rat model were assessed via hematoxylin-eosin (HE) staining, and immunohistochemical staining of collagen I (COL I). KEY FINDINGS: Adipocyte-derived MVs promoted adipogenic differentiation via increasing Oil-Red O staining positive cells, adiponectin (Adipoq), acid-binding protein 2 (aP2) and peroxisome proliferator-activated receptor γ (PPAR-γ) levels, and repressed osteogenic differentiation of BMSCs via decreasing ARS staining positive cells, ALP, Runt-related transcription factor 2 (RUNX2) and osteocalcin (OCN) levels. MiR-148a was present in adipocyte-derived MVs, and miR-148a knockdown inhibited adipogenic differentiation and promoted osteogenic differentiation. Furthermore, Wnt5a expression was regulated by miR-148a. MiR-148a overexpression facilitated adipogenic differentiation and suppressed osteogenic differentiation via regulating the Wnt5a/Ror2 pathway. Adipocyte-derived MVs promoted adipogenic differentiation and inhibited osteogenic differentiation in MPS-induced ONFH rat model. SIGNIFICANCE: Adipocyte-derived MVs-miR-148a promoted adipogenic differentiation and suppressed osteogenic differentiation via targeting the Wnt5a/Ror2 pathway.
AIMS: Adipocyte-secreted microvesicles (MVs)-derived microRNAs (miRNAs) are relevant to adipogenic and osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) in osteonecrosis of the femoral head (ONFH). Our aims are to investigate the mechanism of adipocyte-derived MVs-miR-148a in ONFH. MATERIALS AND METHODS: Adipocyte-derived MVs were identified via transmission electron microscopy and specific markers expression. The adipogenic and osteogenic differentiation were investigated by Oil-Red O staining, alkaline phosphatase (ALP) activity, Alizarin Red S (ARS) staining and osteogenic or adipogenic factors levels. Genes and proteins expression were detected by using quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting. The relationship between miR-148a and Wnt5a was tested via dual-luciferase reporter analysis. The adipogenic differentiation and osteogenic differentiation in methylprednisolone (MPS)-induced ONFH rat model were assessed via hematoxylin-eosin (HE) staining, and immunohistochemical staining of collagen I (COL I). KEY FINDINGS: Adipocyte-derived MVs promoted adipogenic differentiation via increasing Oil-Red O staining positive cells, adiponectin (Adipoq), acid-binding protein 2 (aP2) and peroxisome proliferator-activated receptor γ (PPAR-γ) levels, and repressed osteogenic differentiation of BMSCs via decreasing ARS staining positive cells, ALP, Runt-related transcription factor 2 (RUNX2) and osteocalcin (OCN) levels. MiR-148a was present in adipocyte-derived MVs, and miR-148a knockdown inhibited adipogenic differentiation and promoted osteogenic differentiation. Furthermore, Wnt5a expression was regulated by miR-148a. MiR-148a overexpression facilitated adipogenic differentiation and suppressed osteogenic differentiation via regulating the Wnt5a/Ror2 pathway. Adipocyte-derived MVs promoted adipogenic differentiation and inhibited osteogenic differentiation in MPS-induced ONFH rat model. SIGNIFICANCE: Adipocyte-derived MVs-miR-148a promoted adipogenic differentiation and suppressed osteogenic differentiation via targeting the Wnt5a/Ror2 pathway.