Research Paper Volume 15, Issue 20 pp 11471—11488
The Wnt/β-catenin signaling pathway inhibits osteoporosis by regulating the expression of TERT: an in vivo and in vitro study
- 1 Department of Orthopaedics, The First Affiliated Hospital, Dalian Medical University, Xigang, Dalian 116011, China
- 2 Department of Clinical Pharmacology, College of Pharmacy, Dalian Medical University, Lvshunkou, Dalian 116044, China
Received: August 3, 2023 Accepted: October 2, 2023 Published: October 19, 2023
https://doi.org/10.18632/aging.205136How to Cite
Copyright: © 2023 Cai et al. This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Abstract
Our study was performed to investigate whether the Wingless and int-1 (Wnt) signaling pathway promotes osteogenic differentiation and inhibits apoptosis in bone marrow mesenchymal stem cells (BMSCs) by regulating telomerase reverse transcriptase (TERT) expression. An in vivo model of osteoporosis (OP) in C57BL/6J mice by bilateral ovariectomy (OVX) and an in vitro model of H2O2-induced BMSCs were established separately. Western blotting was used to detect the expression of the pathway-related proteins TERT, β-catenin, and phosphorylated-glycogen synthase kinase-3beta (p-GSK3β)/GSK3β, the osteogenic-related markers osteopontin (OPN), bone morphogenetic protein 2 (BMP2), and runt-related transcription factor 2 (Runx2), and the apoptosis-related indicators B-cell lymphoma-2 (Bcl-2) and BAX. Osteoblastic phenotypes were also evaluated by alkaline phosphatase (ALP) staining and serum ALP activity assays. Osteogenic differentiation phenotypes in mice were verified by H&E staining, micro-CT, and parameter analysis of the femur. Western blotting results showed that the expression of the pathway-related proteins TERT, β-catenin, p-GSK3β/GSK3β was reduced in OVX mice and H2O2-induced BMSCs, accompanied by downregulated protein expression of osteogenic-related markers and antiapoptotic indicators and upregulated protein expression of apoptotic proteins compared to those in the control group. Mechanistic studies showed that the activation of Wnt signaling pathway in BMSCs promoted β-catenin translocation to the nucleus, as verified by immunofluorescence and facilitated colocalization between β-catenin and TERT, as verified by double-labeling immunofluorescence, thereby promoting osteogenic differentiation and reducing apoptosis. In summary, our experiments confirmed that the GSK3β/β-catenin/TERT pathway could regulate the osteogenic differentiation and apoptosis of BMSCs and that TERT might be a promising target for the future treatment of osteoporosis.
Abbreviations
BMSCs: bone marrow mesenchymal stem cells; TERT: telomerase reverse transcriptase; OP: osteoporosis; OVX: bilateral ovariectomy; LiCl: Lithium chloride; p-GSK3β: phosphorylated-glycogen synthase kinase-3beta; OPN: osteopontin; BMP2: Bone morphogenetic protein 2; Runx2: runt-related transcription factor 2; Bcl-2: B-cell lymphoma-2; ALP: alkaline phosphatase; Wnt: Wingless and int-1; TGF-β: Transforming growth factor beta; TCF-4: T-cell factor 4; LEF-1: lymphoid enhancer-binding factor 1; BRG1: Brahma-related gene 1; LiCl: Lithium chloride; H&E: hematoxylin and eosin; Co-IP: immunoprecipitation; FASL: FAS ligand; EMT: epithelial-mesenchymal transition.