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. 2023 Feb 8;10(12):2207152. doi: 10.1002/advs.202207152

Figure 3.

Figure 3

Rspo1 deficiency promotes adipose thermogenesis and resists HFD‐induced obesity. A–C) A representative global image (A), body weight curve (n = 15–22 per group) (B), and body composition (n = 7–8 per group) (C) of Rspo1 −/− and WT mice fed HFD for 13 weeks. D) Hourly (left) and total (right) energy expenditure over 24 h between Rspo1 −/− and WT mice fed with 1‐week HFD (n = 8 per group). The hourly measurements were assessed by two‐way ANOVA model to evaluate the interaction between genotype and time, and pairwised t‐test with Benjamini–Hochberg correction was used as post‐hoc test for evaluating the differences between genotypes in each hour. The total energy expenditure over 24 h was compared with unpaired Student's t‐test. E) Representative images of H&E staining and UCP1 immunohistochemical staining in BAT of WT and Rspo1 −/− mice fed HFD (n = 4 per group). Scale bar, 100 µm. F) The mtDNA content of BAT in Rspo1 −/− and WT mice fed HFD (n = 7–8 per group). G) Representative images of H&E staining and UCP1 immunohistochemical staining in eWAT of WT and Rspo1 −/− mice exposed to chronic cold stimulation (4 °C) for 10 days (n = 4 per group). Scale bar, 100 µm. H) The volcano plot of genes differentially expressed in eWAT of Rspo1 −/− versus WT mice under chronic cold stimulation (n = 3 per group). Biomarkers associated with mitochondrial functions were indicated. I) The top‐upregulated pathways (FDR < 0.05) revealed by GSEA analysis based on GO:BP database in eWAT of Rspo1 −/− versus WT mice under chronic cold stimulation. J,K) GSEA results of mitochondrial gene expression (J) and adaptive thermogenesis (K) in eWAT of Rspo1 −/− versus WT mice under chronic cold stimulation. L,N) Relative mtDNA content (L), quantitative PCR analysis (M), and Western blotting (N) analysis of mitochondrial respiratory complexes and thermogenic genes in eWAT of Rspo1 −/− and WT mice under chronic cold stimulation (n = 7–8 per group). Data are shown as the mean ± sem, and statistical differences between genotypes were assessed by unpaired Student's t‐test (B,C,F,L,M); FDR below 0.05 was considered as the criteria for evaluating differential expressed genes between genotypes (H); FDR below 0.1 was considered as statistical significance in GSEA (I–K). *p < 0.05; **p < 0.01; ***p < 0.001.