Osteoblast and Osteoclast Activity Affect Bone Remodeling Upon Regulation by Mechanical Loading-Induced Leukemia Inhibitory Factor Expression in Osteocytes.

Jingke Du,Zihao He,Mingming Xu,Yiqi Yang,Xinhua Qu,Zhifeng Yu,Mengning Yan,Hanjun Li,Jiancheng Yang,Ning Zhao,Junqi Cui

doi:10.3389/fmolb.2020.585056

Abstract

PurposeBone remodeling is affected by mechanical stimulation. Osteocytes are the primary mechanical load-sensing cells in the bone, and can regulate osteoblast and osteoclast activity, thus playing a key role in bone remodeling. Further, bone mass during exercise is also regulated by Leukemia inhibitory factor (LIF). This study aimed to investigate the role of LIF in the mechanical response of the bone, in vivo and in vitro, and to elucidate the mechanism by which osteocytes secrete LIF to regulate osteoblasts and osteoclasts.MethodsA tail-suspension (TS) mouse model was used in this study to mimic muscular disuse. ELISA and immunohistochemistry were performed to detect bone and serum LIF levels. Micro-computed tomography (CT) of the mouse femurs was performed to measure three-dimensional bone structure parameters. Fluid shear stress (FSS) and microgravity simulation experiments were performed to study mechanical stress-induced LIF secretion and its resultant effects. Bone marrow macrophages (BMMs) and bone mesenchymal stem cells (BMSCs) were cultured to induce in vitro osteoclastogenesis and osteogenesis, respectively.ResultsMicro-CT results showed that TS mice exhibited deteriorated bone microstructure and lower serum LIF expression. LIF secretion by osteocytes was promoted by FSS and was repressed in a microgravity environment. Further experiments showed that LIF could elevate the tartrate-resistant acid phosphatase activity in BMM-derived osteoclasts through the STAT3 signaling pathway. LIF also enhanced alkaline phosphatase staining and osteogenesis-related gene expression during the osteogenic differentiation of BMSCs.ConclusionMechanical loading affected LIF expression levels in osteocytes, thereby altering the balance between osteoclastogenesis and osteogenesis.

Highlights

Mechanical loading plays an essential role in the maintenance of bone quality and quantity, and osteocytes are critical for sensing mechanical stimulation (Dallas et al, 2013; Bellido, 2014; Choy et al, 2020)
After performing Gene Ontology (GO) (Figure 2A) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway (Figure 2B) enrichment analyses, we found that differential genes were enriched in the cytokinerelated pathways
We validated a series of cytokine expression levels by Quantitative Reverse-Transcription Polymerase Chain Reaction (qRT-PCR) (Figure 2D) and confirmed that Leukemia inhibitory factor (LIF) expression increased on fluid shear stress (FSS) treatment

Summary

Introduction

Mechanical loading plays an essential role in the maintenance of bone quality and quantity, and osteocytes are critical for sensing mechanical stimulation (Dallas et al, 2013; Bellido, 2014; Choy et al, 2020). In vivo studies have shown that osteocytes always exist in a complex mechanical environment, which includes shear stress, tensile strain, and pressure (Iolascon et al, 2013). When stimulated by mechanical strain (Wang et al, 2019), osteocytes can regulate osteoblast proliferation and differentiation by releasing signaling molecules, such as nitric oxide, prostaglandin E2, and adenosine triphosphate (Bakker et al, 2001; Genetos et al, 2005). Studies have shown that bed-ridden patients and astronauts develop osteoporosis due to reduced mechanical bone stimulation, but the mechanism of this phenomenon remains unclear (Norvell et al, 2004; Sibonga, 2013; Yang et al, 2018). Previous studies have shown that muscle loading impact bone development, and LIF influences this process (Broholm et al, 2008). We hypothesized that mechanical loading-induced LIF in osteocytes might regulate bone remodeling

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