Fig 2: Blockage of ceramide generation reverses decreased thermogenesis and increased obesity phenotypes in Ormdl3−/− mice. (A–C) The Ormdl3−/− and their littermate WT mice were administered the β3 adrenergic agonist CL316243 plus myriocin or vehicle (n = 5 mice per group), and the body temperature change in mice was determined by cold challenge at 4 °C (A). Representative H&E (B) and immunohistochemical (C) staining of ingWAT, eWAT and iBAT. (D–F) Eight-week-old male WT and Ormdl3−/− mice were fed HFD for additional 16 weeks. From the 9th week, myriocin or saline was administered to mice twice per week. (D) Body weight of WT and Ormdl3−/− mice during 16 weeks (n = 6). (E) Left: Glucose tolerance tests. Right: Insulin tolerance tests (n = 5). (F) Western blot analysis of UCP1 and PGC1α protein levels. The data are presented as mean ± SEM. P values were determined by two-way ANOVA with Šidák's multiple comparisons test (A, WT + CL316243 versus Ormdl3−/− + CL316243; D and E, WT versus Ormdl3−/−). NS means no significance, ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001. Scale bar, 100 μm.

Fig 3: The thermogenic capacity of BAT is impaired in Ormdl3 deficient mice. (A) The relative mRNA levels of Ormdl3 in WAT and iBAT in Ormdl3−/− and WT mice (n = 4). (B–F) Eight-week-old Ormdl3−/− and their littermate WT mice (n = 6 mice per group) were housed at thermoneutral (30 °C) or exposed to cold environment (4 °C) for additional 2 weeks, and the following analysis was then performed: Body temperature in mice before and after cold challenge (B). The mice, which had been trained in thermoneutral or cold environment, were kept at room temperature for 24 h and then subjected to core body temperature analyses in acute cold exposure condition (C). Quantitative PCR analysis of thermogenic gene expression in iBAT (D). Western blot analysis of UCP1 and PGC1α protein levels in iBAT (E). Representative H&E and immunohistochemical staining of iBAT (F). Scale bar, 100 μm. The data are presented as mean ± SEM. P values were determined by unpaired nonparametric test (A), one-way ANOVA (B and D) or two-way ANOVA with Šidák's multiple comparisons test (C, WT + 4 °C versus Ormdl3−/− + 4 °C). NS, no significance; ∗p < 0.05, ∗∗p < 0.01.

Fig 4: Brown adipose tissue (BAT) transplantation improved the reproductive phenotype of polycystic ovary syndrome (PCOS) model rats. (A) Disordered estrous cycles were observed in the DHEA+Sham group, while BAT transplantation rescued the abnormal estrous cycles. (B) Representative results of ovarian H&E staining of the control (Ctrl) group, DHEA+Sham group, and DHEA+BAT group. Ovarian histology revealed that cystic follicles (arrow) and a few corpora lutea (asterisk) appeared in the DHEA+Sham group compared with the Ctrl group, while BAT transplantation reversed the phenotype caused by dehydroepiandrosterone (DHEA). H&E, hematoxylin–eosin; D, diestrus; E, estrus; M, metestrus; P, proestrus; UCP1, uncoupling protein 1.

Fig 5: Effects of PPH on activating the mitochondrion-related signaling pathway in scapular adipose tissue. Western blotting analyses of PGC 1α, UCP 1, and mitochondria complex protein expression (A); Quantification of the PGC 1α/β-Tubulin ratio, UCP 1/β-Tubulin ratio, and mitochondria complex/β-Tubulin ratio (B). CI: complex I; CII: complex II; CIII: complex III; CIV: complex IV; CV: complex V. *p < 0.05, **p < 0.01, and ***p < 0.001.