Gene expression of intracortical bone demonstrates loading-induced increases in Wnt1 and Ngf and inhibition of bone remodeling processes

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Harris, Taylor L.^1,2; Silva, Matthew J.¹

Gene expression of intracortical bone demonstrates loading-induced increases in Wnt1 and Ngf and inhibition of bone remodeling processes

Bone. September 2021;150:116019

©2021 Elsevier Inc. All rights reserved.

Affiliations

¹Department of Orthopaedic Surgery and Musculoskeletal Research Center, Washington University School of Medicine, Saint Louis, MO, United States

²Department of Biomedical Engineering, Washington University, Saint Louis, MO, United States
Abstract and keywords

Osteocytes are the primary mechanosensitive cells in bone. However, their location in mineralized matrix has limited the in vivo study of osteocytic genes induced by mechanical loading. Laser Capture Microdissection (LCM) allows isolation of intracortical bone (Intra-CB), enriched for osteocytes, from bone tissue for gene expression analysis. We used microarray to analyze gene expression from mouse tibial Intra-CB dissected using LCM 4 h after a single loading bout or after 5 days of loading. Osteocyte enrichment was supported by greater expression of Sost, Dmp1, Dkk1, and Mepe in Intra-CB regions vs. Mixed regions containing periosteum and muscle (fold-change (FC) = 3.4, 2.2, 5.1, 3.0, respectively). Over 150 differentially expressed genes (DEGs) due to loading (loaded vs. contralateral control) in Intra-CB were found on Day 1 and Day 5, but only 10 genes were differentially expressed on both days, including Ngf (Day 1 FC = 13.5, Day 5 FC = 11.1) and Wnt1 (Day 1 FC = 1.5, Day 5 FC = 5.1). The expression of Ngf and Wnt1 within Intra-CB was confirmed by in situ hybridization, and a significant increase in number of Wnt1 mRNA molecules occurred on day 1. We also found changes in extracellular matrix remodeling with Timp1 (FC = 3.1) increased on day 1 and MMP13 (FC = 0.3) decreased on day 5. Supporting this result, IHC for osteocytic MMP13 demonstrated a marginal decrease due to loading on day 5. Gene Ontology (GO) biological processes for loading DEGs indicated regulation of vasculature, neuronal and immune processes while cell-type specific gene lists suggested regulation of osteoclast, osteoblast, and endothelial related genes. In summary, microarray analysis of microdissected Intra-CB revealed differential regulation of Ngf, Wnt1, and MMP13 due to loading in osteocytes.

Keywords: Osteocytes; Laser Capture Microdissection; Mechanical loading; Bone formation; Gene expression
Links

DOI: 10.1016/j.bone.2021.116019

PubMed: 33971314

WoS: 000670402300011
History

Received: 2021-01-24

Revised: 2021-04-27

Accepted: 2021-05-18

Online: 2021-05-21

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Cited By (3)

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2023	Xu X, Yang H, Bullock WA, Gallant MA, Ohlsson C, Bellido TM, Main RP. Osteocyte estrogen receptor β (Ot-ERβ) regulates bone turnover and skeletal adaptive response to mechanical loading differently in male and female growing and adult mice. J Bone Miner Res. January 2023;38(1):186-197.
2023	Wee NKY, Novak S, Ghosh D, Root SH, Dickerson IM, Kalajzic I. Inhibition of CGRP signaling impairs fracture healing in mice. J Orthop Res. June 2023;41(6):1228-1239.
2021	Harris TL. Mechanisms for Osteoblast and Osteocyte Initiation and Sustainment of Bone Formation in Young-Adult and Aged Mice [PhD thesis]. St. Louis, MO: Washington University in St. Louis; August 2021.