Galectin-3 and TRIM16 coregulate osteogenic differentiation of human bone marrow-derived mesenchymal stem cells at least partly via enhancing autophagy

Wen-Ting Chen; Fan Zhang; Xing-Qi Zhao; Bin Yu; Bo-Wei Wang

doi:10.1016/j.bone.2019.115059

Galectin-3 and TRIM16 coregulate osteogenic differentiation of human bone marrow-derived mesenchymal stem cells at least partly via enhancing autophagy

Bone. 2020 Feb:131:115059. doi: 10.1016/j.bone.2019.115059. Epub 2019 Sep 12.

Authors

Wen-Ting Chen¹, Fan Zhang², Xing-Qi Zhao², Bin Yu³, Bo-Wei Wang⁴

Affiliations

¹ Department of Anesthesiology, Nanfang Hospital, Southern Medical University, Guangzhou 510515, PR China.
² Department of Orthopaedics and Traumatology, Nanfang Hospital, Southern Medical University, Guangzhou 510515, PR China.
³ Guangdong Provincial Key Laboratory of Bone and Cartilage Regenerative Medicine, Nanfang Hospital, Southern Medical University, Guangzhou 510515, PR China. Electronic address: hhgbm653@163.com.
⁴ Department of Orthopaedics and Traumatology, Nanfang Hospital, Southern Medical University, Guangzhou 510515, PR China. Electronic address: wangbowei5563@163.com.

PMID: 31521826
DOI: 10.1016/j.bone.2019.115059

Abstract

Background: The osteogenic differentiation of human bone marrow-derived mesenchymal stem cells (hBMSCs) is critical for bone homeostasis. Here, we investigated the regulation of Galectin-3 and tripartite motif protein 16 (TRIM16) on osteogenic differentiation of hBMSCs through autophagy.

Methods: Quantitative PCR (qPCR) and western blot were performed to determine the expression of osteogenic markers, autophagic markers, Galectin-3 and TRIM16. Short-hairpin RNAs (shRNAs) and overexpression plasmids were used to manipulate the expression of Galectin-3, TRIM16 and Unc-51 like autophagy activating kinase 1 (ULK1). Alkaline phosphatase (ALP) activity was measured by ALP staining assay. Calcium deposition in differentiated hBMSCs was assessed by Alizarin Red S staining. LC3 puncta formation was monitored by immunofluorescence staining. The interaction between indicated proteins was confirmed by co-immunoprecipitation (Co-IP) assay.

Results: Either Galectin-3 or TRIM16 knockdown led to impaired ALP activity, reduced calcium deposition, down-regulation of pro-osteogenic markers as well as restrained autophagy in osteogenic-induced hBMSCs. However, overexpression of Galectin-3 or TRIM16 promoted osteogenic differentiation of hBMSCs, which was then compromised by autophagy inhibition. Co-IP experiment demonstrated that TRIM16 associated with Galectin-3 through ULK1. Meanwhile, osteogenic induction enhanced the association between TRIM16 and ULK1 or coiled-coil myosin-like BCL2-interacting protein (Beclin1), and TRIM16 increased the stability of ULK1 and Beclin1. Moreover, either TRIM16 or ULK1 knockdown dampened the pro-osteogenic effect of Galectin-3, which elucidated that Galectin-3 mediated osteogenic differentiation was at least partly dependent on TRIM16 and ULK1.

Conclusion: In summary, the present study revealed Galectin-3 and TRIM16 co-regulated osteogenic differentiation of hBMSCs at least partly via enhancing autophagy, which might provide a promising approach for osteoporosis treatment in future.

Keywords: Autophagy; Galectin-3; Osteogenic differentiation; TRIM16; ULK1; hBMSCs.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Autophagy
Bone Marrow
Cell Differentiation
Cells, Cultured
Galectin 3* / genetics
Humans
Mesenchymal Stem Cells* / cytology
Osteogenesis*
Tripartite Motif Proteins*
Ubiquitin-Protein Ligases*

Substances

Galectin 3
Tripartite Motif Proteins
TRIM16 protein, human
Ubiquitin-Protein Ligases