A novel miR-466l-3p/FGF23 axis promotes osteogenic differentiation of human bone marrow mesenchymal stem cells

Xiang Zhang; Jin Xu

doi:10.1016/j.bone.2024.117123

A novel miR-466l-3p/FGF23 axis promotes osteogenic differentiation of human bone marrow mesenchymal stem cells

Bone. 2024 May 11:185:117123. doi: 10.1016/j.bone.2024.117123. Online ahead of print.

Authors

Xiang Zhang¹, Jin Xu²

Affiliations

¹ Key Laboratory of Endocrine Glucose & Lipids Metabolism and Brain Aging, Ministry of Education, Department of Endocrinology, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan, Shandong 250021, China; Shandong Key Laboratory of Endocrinology and Lipid Metabolism, Jinan, Shandong 250021, China; Shandong Institute of Endocrine and Metabolic Diseases, Jinan, Shandong 250021, China.
² Key Laboratory of Endocrine Glucose & Lipids Metabolism and Brain Aging, Ministry of Education, Department of Endocrinology, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan, Shandong 250021, China; Shandong Key Laboratory of Endocrinology and Lipid Metabolism, Jinan, Shandong 250021, China; Shandong Institute of Endocrine and Metabolic Diseases, Jinan, Shandong 250021, China; "Chuangxin China" Innovation Base of stem cell and Gene Therapy for endocrine Metabolic diseases, Jinan, Shandong 250021, China; Shandong Engineering Laboratory of Prevention and Control for Endocrine and Metabolic Diseases, Jinan, Shandong 250021, China; Shandong Engineering Research Center of Stem Cell and Gene Therapy for Endocrine and Metabolic Diseases, Jinan, Shandong 250021, China. Electronic address: xujin@sdfmu.edu.cn.

PMID: 38735373
DOI: 10.1016/j.bone.2024.117123

Abstract

Background: MicroRNAs (miRNAs) regulate osteogenic differentiation processes and influence the development of osteoporosis (OP). This study aimed to investigate the potential role of miR-466 l-3p in OP.

Methods: The expression levels of miR-466 l-3p and fibroblast growth factor 23 (FGF23) were quantified in the trabeculae of the femoral neck of 40 individuals with or without OP using quantitative reverse transcription-polymerase chain reaction (qRT-PCR). The impact of miR-466 l-3p or FGF23 expression on cell proliferation and the expression levels of runt-related transcription factor 2 (RUNX2), type I collagen (Col1), osteocalcin (OCN), osterix (OSX) and dentin matrix protein 1 (DMP1) was quantified in human bone marrow mesenchymal stem cells (hBMSCs) overexpressing miR-466 l-3p. Furthermore, alkaline phosphatase (ALP) staining and alizarin red staining were performed to measure ALP activity and the levels of calcium deposition, respectively. In addition, bioinformatics analysis, luciferase reporter assays, and RNA pull-down assays were conducted to explore the molecular mechanisms underlying the effects of miR-466 l-3p and FGF23 in osteogenic differentiation of hBMSCs.

Results: The expression levels of miR-466 l-3p were significantly lower in femoral neck trabeculae of patients with OP than in the control cohort, whereas FGF23 levels exhibited the opposite trend. Furthermore, miR-466 l-3p levels were upregulated and FGF23 levels were downregulated in hBMSCs during osteogenic differentiation. Moreover, the high miR-466 l-3p expression enhanced the mRNA expression of RUNX2, Col1, OCN, OSX and DMP1, as well as cell proliferation, ALP activity, and calcium deposition in hBMSCs. FGF23 was found to be a direct target of miR-466 l-3p. FGF23 overexpression downregulated the expression of osteoblast markers and inhibited the osteogenic differentiation induced by miR-466 l-3p overexpression. qRT-PCR and Western blot assays showed that miR-466 l-3p overexpression decreased the expression levels of mRNAs and proteins associated with the phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) signaling pathway, whereas FGF23 upregulation exhibited the opposite trend.

Conclusion: In conclusion, these findings suggest that miR-466 l-3p enhances the osteogenic differentiation of hBMSCs by suppressing FGF23 expression, ultimately preventing OP.

Keywords: Fibroblast growth factor 23; Human bone marrow mesenchymal stem cells; Osteoporosis; miRNAs.