Brown fat-specific mitoribosomal function is crucial for preventing cold exposure-induced bone loss

Abstract

This study examines the interplay between ambient temperature, brown adipose tissue (BAT) function, and bone metabolism, emphasizing the effects of cold exposure and BAT mitochondrial activity on bone health. Utilizing ovariectomized (OVX) mice to model primary osteoporosis and BAT-specific mitochondrial dysfunction (BKO) mice, we evaluated the impact of housing temperature on bone density, immune modulation in bone marrow, and the protective role of BAT against bone loss. Cold exposure was found to universally reduce bone mass, enhance osteoclastogenesis, and alter bone marrow T-cell populations, implicating the immune system in bone remodeling under cold stress. The thermogenic function of BAT, driven by mitochondrial oxidative phosphorylation, was crucial in protecting against bone loss. Impaired BAT function, through surgical removal or mitochondrial dysfunction, exacerbated bone loss in cold environments, highlighting BAT's metabolic role in maintaining bone health. Furthermore, cold-induced changes in BAT function led to systemic metabolic shifts, including elevated long-chain fatty acids, which influenced osteoclast differentiation and activity. These findings suggest a systemic mechanism connecting environmental temperature and BAT metabolism with bone physiology, providing new insights into the metabolic and environmental determinants of bone health. Future research could lead to novel bone disease therapies targeting these pathways.

Keywords: Bone loss; Brown adipose tissue; Cold exposure; Mitochondria.

Fig. 1

Aggravation of ovariectomy-induced bone loss by low temperature housing conditions. A Illustrative micro-CT images showing the trabecular areas in the lower section of the femur. B Micro-CT measurement of trabecular bone mineral density (Tb.BMD); trabecular bone volume (Tb.BV); trabecular bone volume/tissue volume (Tb.BV/TV); trabecular bone surface/tissue volume (Tb.BS/TV); trabecular number Tb.N; and trabecular separation Tb.Sp C, D TRAP staining for osteoclasts and statistical analysis of osteoclast number. E, Serum levels of P1NP. The results were expressed as the mean ± SD. Statistical significance was estimated using unpaired t-tests. *, P < 0.05

Fig. 2

Temperature-mediated changes in osteoclastogenic bone marrow T cells and brown adipose tissue (BAT). A Representative contour plots from flow cytometry for regulatory T-cells (Treg; CD4 + CD25 + FOXP3+); RANKL (CD254) producing Treg cells; and IFN-γ or IL-17 A-producing CD4 + cells from bone marrow. B Statistical analysis of phenotypes defined by flow cytometry. C, D Representative western blots and band density measurements of UCP-1 and OxPhos complex subunits in the BAT isolated from mice housed for 12 weeks at 22–30 °C. E Representative immunohistochemical images of UCP-1 and SDH staining of BAT from mice housed at different temperatures. Scale bar, 50 μm. Data were expressed as the mean ± SD. Statistical significance was estimated using unpaired t-tests. *, P < 0.05, **, P < 0.01 ***, P < 0.001

Fig. 3

Removal of interscapular BAT promotes bone loss. A Images of mice at 12 weeks after post-surgical removal of BAT. B Representative images of micro-CT of cortical and trabecular regions in the distal femur. C Measurement of Tb.BMD, Tb.BV, Tb.BV/TV, Tb.N, Tb.Sp, Ct.BMD, Ct.BV, and Ct.Th in the femur of SHAM and BAT-deficient mice housed at 22 °C–14 °C for 12 weeks. Statistical significance was estimated using unpaired t-tests. *, P < 0.05, **, P < 0.01

Fig. 4

BAT-specific mitochondrial dysfunction (BKO) mice show impairment of OxPhos. A BKO mice generated by knockout of Crif1 in brown adipose tissue using UCP-1-cre mice and the Cre-loxP system. B, C Immunoblotting of OxPhos complex subunits and UCP-1 in BAT isolated from control and BKO mice housed at 22 °C at 20 weeks old. D Representative BN-PAGE image of the assembled OxPhos complex in BAT tissue. E, F Representative immunohistochemical images of UCP-1 and SDH staining in BAT paraffin sections. Scale bar, 50 μm. The statistical relevance of the findings was determined using unpaired t-tests. *, P < 0.05, **, P < 0.01 ***, P < 0.001. The analysis was conducted in comparison with the specified reference group

Fig. 5

BAT-specific mitochondrial dysfunction (BKO) mice exposed to cold stress exhibit bone loss and an osteoclastogenic bone marrow T-cell phenotype. A Micro-CT scans showing the cortical and trabecular structures in the distal part of the femur. B Measurement of Tb.BMD, Tb.BV/TV, Tb.Th, Tb.N, Tb.Sp, Ct.BMD, Ct.BV and Ct.Th in the femur. C Representative contour plots from flow cytometry for regulatory T-cells (Treg; CD4 + CD25 + FOXP3+); RANKL (CD254) producing Treg cells; and IFN-γ producing CD4 + cells from bone marrow. D Serum levels of RANKL. Data are expressed as the mean ± SD. Statistical significance was determined using one-way ANOVA for comparative analysis. *, P < 0.05 and **, P < 0.01 ***, P < 0.001

Fig. 6

Modifications of lipid composition in thermogenic adipose tissue. A Representative images of H&E staining of BAT and iWAT sections obtained from 20-week-old mice housed at 22 °C. Scale bar, 50 μm. B Statistical analysis of lipid area in BAT and lipid diameter in iWAT. C, D Representative western blots and band density measurements for PKA, p-PKA, HSL, p-HSL and ATGL in BAT of the control and BAT-specific mitochondrial dysfunction (BKO) mice. E Real-time PCR analysis of fatty acid oxidation related genes in the BAT of 20-week old control and BKO mice. F Quantification of serum free fatty acids. G Comparative analysis of BAT and serum metabolite levels in 20-week-old control and BKO mice. Statistical significance was analyzed by one-way ANOVA. *, P < 0.05 and **, P < 0.01 ***, P < 0.001 compared with the indicated group

Fig. 7

Treatment with palmitic acid increases the number of osteoclasts and promotes production of RANKL and IFN-γ in bone marrow T cells. A TRAP staining of osteoclasts and the number of TRAP-positive osteoclasts. Scale bar: 200 μm. B RANKL producing cells in Foxp3 + Treg populations treated with or without palmitic acid. C IFN-γ producing cells within the CD4 + T-cell populations treated with or without palmitic acid. Statistical significance was analyzed by one-way ANOVA. *, P < 0.05 and **, P < 0.01 compared with the indicated group