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Updated: Jun 13, 2026
A Novel in vivo Gene Transfer Technique and in vitro Cell Based Assays for the Study of Bone Loss in Musculoskeletal Disorders
Published on: June 8, 2014
MiR-494-3p knockdown promotes osteogenic differentiation of bone marrow mesenchymal stem cells in ovariectomised rats
Jianping Lei1, Song Guo1, Jiabing Kuang2
1Department of Orthopaedics, Wuhan Fourth Hospital, Wuhan, China.
Introduction:
Osteoporosis is a bone disease characterised by low bone mass, deterioration of bone tissue, and disruption of bone microarchitecture. MicroRNAs have been found to play an important role in osteoporosis. MicroRNA (miR)-494 is inhibited during bone angiogenesis, and its overexpression reduces osteogenic differentiation gene expression. Sirtuin 1 (Sirt1) is a histone deacetylase with multiple cellular activities including increasing bone mass and delaying the onset of osteoporosis. MiR-494-3p has been predicted in computer-assisted bioinformatics analysis to target the 3'UTR of Sirt1 mRNA. The aim of the present study was to assess the effect of miR-494-3p on ovariectomy (OVX)-induced osteoporosis in rats and the relevant mechanisms.
Material And Methods:
Osteoporosis in female rats was induced by OVX, and bone microarchitectural changes were evaluated by means of microCT. MiR-494-3p and Sirt1 expression in femurs were evaluated by RT-qPCR or Western blotting. Bone marrow mesenchymal stem cells (BMSCs) were isolated from rat femurs and identified by flow cytometry. Then, BMSCs were transfected with miR-494-3p inhibitor/mimic, si-Sirt1 and negative controls as well as pcDNA3.1-TLR4 and empty pcDNA3.1 vector. Osteogenic cell differentiation was assessed via Alizarin Red, alkaline phosphatase (ALP) and Oil Red O staining.
Results:
MiR-494-3p level was upregulated, and Sirt1 mRNA and protein levels were downregulated, in femurs of OVX rats. Functionally, miR-494-3p inhibited osteogenic differentiation of cultured rat BMSCs. Mechanistically, miR-494-3p regulated the Sirt1 3'UTR to activate TLR4/NF-κB signalling, and Sirt1 inhibition and TLR4 overexpression reversed the enhancing effect of miR-494-3p knockdown on osteogenic differentiation.
Conclusions:
MiR-494-3p represses osteogenic differentiation of BMSCs in OVX rats through Sirt1/TLR4/NF-κB signalling.
