Genetic depletion of adipose-derived Isthmin-1 causes hepatic steatosis

Figures

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  Figure 1. Adipose-specific Ism1 ablation results in lower respiratory exchange ratio. A. Genotyping of WT (Ism1 floxed allele fl/fl) and AdipoQ-Ism1-KO (Ism1 floxed allele fl/fl crossed to Adiponectin-cre). B. Ism1 gene expression in lung, skin, gut, brain, liver, muscle, inguinal white adipose (iWAT), and brown fat (BAT) tissues from WT and AdipoQ-Ism1-KO (N = 3–8 biological replicates). C. Body weights in WT and AdipoQ-Ism1-KO mice (N = 5–8 mice per group). D, G. Oxygen consumption (VO2) in WT and AdipoQ-Ism1-KO mice (N = 4 mice per group). E, H. Carbon dioxide production (VCO2) in WT and AdipoQ-Ism1-KO mice (N = 4 mice per group). F, I. Respiratory exchange ratio (RER) in WT and AdipoQ-Ism1-KO mice (N = 4 mice per group). J. Accumulated food intake in WT and AdipoQ-Ism1-KO mice (N = 4 mice per group). K. Locomotor activity in WT and AdipoQ-Ism1-KO mice (N = 4 mice per group). L. Total energy balance expressed as Kcal in WT and AdipoQ-Ism1-KO mice (N = 4 mice per group). Data are presented as mean ± SEM. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001 by t-test (B, C) or two-way ANOVA (D–L).

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  Figure 2. Loss of adipose-derived Ism1 exacerbates MASLD. A. Strategy to induce MASLD in WT and AdipoQ-Ism1-KO mice (see Materials and Methods). B. Glycerol levels in pooled serum samples from WT (N = 5) and AdipoQ-Ism1-KO (N = 3) mice, measured in technical duplicates. C. NEFA levels in pooled serum samples from WT (N = 5) and AdipoQ-Ism1-KO (N = 3) mice, measured in technical duplicates. D. Representative liver photographs from WT or AdipoQ-Ism1-KO mice fed a MASLD diet for 6 weeks. E. Liver weights of WT (N = 5) or AdipoQ-Ism1-KO (N = 8) mice fed a MASLD diet for 6 weeks. F. ALT activity of WT (N = 5) or AdipoQ-Ism1-KO mice (N = 8) fed a MASLD diet for 6 weeks. G. Representative H&E and Oil Red O (lipid) staining in livers from WT or AdipoQ-Ism1-KO mice fed a MASLD diet for 6 weeks. H. Representative LipidTOX (lipid) staining in livers from WT (N = 5) or AdipoQ-Ism1-KO (N = 8) mice fed a MASLD diet for 6 weeks. I. Quantification of the LipidTOX staining (2 images per 5 or 8 biological replicates) in livers from WT or AdipoQ-Ism1-KO mice fed a MASLD diet for 6 weeks. Data are presented as mean ± SEM. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001 by t-test (B, C, E, F, I).

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  Figure 3. Adipose-specific Ism1 deletion elevates deleterious lipid species. A. Schematic illustration showing lipid extraction from mouse livers from MASLD mice and identification of lipids by untargeted LC-MS lipidomic profiling. B. Cloud plot of significantly enriched (green) or de-enriched (red) lipid species in livers from AdipoQ-Ism1-KO mice (N = 8) compared with WT mice (N = 5). C. LC-MS lipidomic profiling of hepatic triacylglycerol (22:5/22:6/22:6) in AdipoQ-Ism1-KO (N = 8) mice compared with WT mice (N = 5). D. LC-MS lipidomic profiling of hepatic diacylglycerol (17:1/18:1/0:0) in AdipoQ-Ism1-KO (N = 8) mice compared with WT mice (N = 5). E. LC-M lipidomic profiling of hepatic diacylglycerol (18:1/14:1/0:0) in AdipoQ-Ism1-KO (N = 8) mice compared with WT mice (N = 5). F. LC-MS lipidomic profiling of hepatic ceramides (d18:1/16:0) in AdipoQ-Ism1-KO (N = 8) mice compared with WT mice (N = 5). G. Plasma diacylglycerol levels in WT (N = 6) or AdipoQ-Ism1-KO (N = 7) mice fed a MASLD diet for 6 weeks. H. Plasma ceramide levels in WT (N = 6) or AdipoQ-Ism1-KO (N = 7) mice fed a MASLD diet for 6 weeks.Data are presented as mean ± S.E.M. ∗P < 0.05, ∗∗P < 0.01, ∗∗∗P < 0.001 by two-way…

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  Figure 4. Ism1 administration reduces several hepatic lipid species. A. Overview of therapeutic administration of vehicle, 5 mg/kg Ism1, or 30 mg/kg GW4064 for 2 weeks in MASLD-induced mice (N = 5 mice/group). B. Representative fluorescent images of LipidTOX in livers from chow mice, or mice with MASLD treated with vehicle, 5 mg/kg Ism1, or 30 mg/kg GW4064 for 14 days (N = 5 mice per group). C. Quantification of LipidTOX (lipid) staining (3 images per 5 biological replicates in total, N = 5 mice per group). P values ∗∗∗∗ represent <0.0001 by one-way Anova Kruskal–Wallis test for multiple comparisons. D. Cloud plot of significantly enriched (green) or de-enriched (red) lipid species detected by LC-MS lipidomics in livers from mice with MASLD treated with 5 mg/kg Ism1 compared with vehicle (N = 5 pe group). E. LC-MS profiling of hepatic palmitic acid in Ism1-treated mice compared with vehicle-treated MASLD mice (N = 5 per group). F. LC-MS profiling of hepatic monoacylglycerol (MG) in Ism1-treated mice compared with vehicle-treated MASLD mice (N = 5 per group). G. LC-MS profiling of hepatic diacylglycerol (DG) in Ism1-treated mice compared with vehicle-treated MASLD mice (N = 5 per group). H.…