Exercise Promotes Bone Marrow Microenvironment by Inhibiting Adipsin in Diet-Induced Male Obese Mice

Figure 1. Exercise improved insulin sensitivity and bone density. (A) The eight-week-old male C57/B6 mice were fed with HFD for 6 weeks. The average body weight of the mice achieved 41.5 g after 6 weeks of HFD and before the mice started the 12-week treadmill running with HFD. Body composition was measured before and after exercise intervention. (B) Body composition was measured after exercise in the obesity and exercise groups using MRI. (C,D) measurement of plasma glucose during ITT and the AUC of the obesity (N = 9) and exercise (N = 7) groups after 8-week treadmill exercise. (E,F) femoral BS, BV in the trabecular regions of TV. (G) representative trabecular reconstruction image of the femur by μCT. The gray part of the picture shows reconstructed trabecular bones. (H) femoral bone density in the trabecular regions of TV. * p < 0.05, ** p < 0.01 2-sided Student t test. Data are presented the mean ± SD. AUC, areas under the curve; BW, body weight; HFD, high fat diet; ITT, insulin tolerance test; MRI, magnetic resonance imaging.

Figure 2. RNA-Seq of bone marrow indicated that exercise inhibited Spp1 and related ECM-receptor pathways. Representative bone marrow from tibia of both the obesity and exercise groups (N = 2,2) were isolated for RNA-Seq analysis. (A) volcano plot of bone marrow in the exercise group compared to the obesity group. Colored dots represent p value < 0.05 (−log10(p Value) > 1.3, gray line), log2Fold Change > 1 up-regulated genes (red), and log2Fold Change < −1 down-regulated genes (blue). Selected ECM-receptor pathway related genes are highlighted. (B) results of pathway enrichment analysis for highly variable genes identified by significant DEG in the bone marrow. (C) gene set enrichment analysis of ECM-receptor interaction. (D) protein interaction network diagram of Spp1. The original data come from the HINT (http://hint.yulab.org, accessed on 31 May 2022) database, a database of high-quality protein–protein interactomes. ECM, extracellular matrix.

Figure 3. Exercise inhibited bone resorption and protected bone by reducing adipsin and Spp1. Quantitative real-time PCR analyses and protein expression level of tibia BM. (A) Bone remodeling related gene expression quantitative analysis of bone marrow. The bone marrow was isolated from the male C57/B6 mice (N = 9) that were induced for 6 weeks of high-fat diet and intervened in twelve weeks of exercise. * p < 0.05, ** p < 0.01, *** p < 0.001 for obesity group (N = 9) vs exercise group (N = 7). (B,C) Western blotting and quantification (N = 6) in tibia bone marrow from obesity group and exercise group. * p < 0.05, for obesity group (N = 6) vs exercise group (N = 6). * p < 0.05, ** p < 0.01 2-sided Student t tests were used for statistical analyses. Data are presented as mean ± SD. BMAT, bone marrow adipose tissue; PCR, polymerase chain reaction.

Figure 4. Exercise inhibited adipogenic differentiation of BMSCs and decreased adipsin in vitro. Isolated tibia mesenchymal stem cells were divided into control group and stretch group during adipogenic differentiation (N = 3). (A) representative images of control and stretch groups under a 10× phase-contrast microscope at the end of adipogenic differentiation. (B) quantitative PCR analysis of adipogenic relative gene expression in the BMSCs, containing Pparg1 and Pparg2. (C,D) representative Western blotting images and protein expression quantification (N = 4) of adipsin from mesenchymal stem cells treated with adipogenic differentiation and stretch stimulation. * p < 0.05, ** p < 0.01 for control group vs stretch group. Two-sided Student t-tests were used for statistical analyses. Data are presented as the mean ± SD. BMSCs, bone mesenchymal stem cells.

Figure 5. Exercise inhibited adipsin-mediated complement pathway activation on osteoclasts. Adipsin plays an essential role in C3 convertase forming and activation of the subsequent alternative complement pathway. As the central component of the complement pathway, C3 and the complement pathway are important in osteoclast differentiation. Exercise inhibits the adipokine adipsin expression and affects osteoclasts through its mediated complement system. In addition, exercise can restrain BMSC adipogenic differentiation to protect bone health. IS, insulin sensitivity; BW, body weight.