Extended Data Fig. 2: Chronic irisin treatment has no effects on energy expenditure and glucose homeostasis in normal-chow-fed mice.

4-week-old male C57BL6/J mice were intravenously injected with AAV-GFP or AAV-Irisin. The mice were fed a normal chow continuously without switching diet. Mice were analyzed in CLAMS cages after another 12 weeks of continuous normal chow feeding, and gene expression analysis was performed after another 18 weeks of continuous normal chow feeding. (A) Body weight measurements in CLAMS cages over one week (n = 8, 8). The last 3.5 days of measurement were shown on the left and the averaged values in the light and dark periods were summarized on the right. Unpaired two-sided T-test was used for calculating p values. (B) Food consumption measurements in CLAMS cages over one week (n = 8, 7). The last 3.5 days of measurement were shown on the left and the averaged values in the light and dark periods were summarized on the right. Unpaired two-sided T-test was used for calculating p values. (C) Energy expenditure measured in CLAMS cages over one week (n = 8, 8). The last 3.5 days of measurement were shown on the left and the averaged values in the light and dark periods were summarized on the right. Two-way ANOVA (left) or unpaired two-sided T-test (right) was used for calculating p values. (D) Locomotive activity measured in CLAMS cages over one week (n = 8, 8). The last 3.5 days of measurement were shown on the left and the averaged values in the light and dark periods were summarized on the right. Two-way ANOVA (left) or unpaired two-sided T-test (right) was used for calculating p values. (E) Intraperitoneal glucose tolerance tests (GTTs) of mice fed a normal chow for 18 weeks (n = 15, 15). Two-way ANOVA was used for calculating p values. (F) Intraperitoneal insulin tolerance tests (ITTs) of mice fed a normal chow for 18 weeks (n = 15, 15). Two-way ANOVA was used for calculating p values. (G) Distance traveled measured in CLAMS cages over one week. n = 8, 8. The last 3.5 days of measurement were shown on the left and the averaged values in the light and dark periods were summarized on the right. Unpaired two-sided T-test was used for calculating p values. (H–J) RT-qPCR measuring mRNAs associated with thermogenesis and adipogenesis in BAT (H) (n = 15, 15), iWAT (I) (n = 15, 15), and eWAT (J) (n = 9, 12) of the mice received AAV-irisin or -GFP and fed a normal chow for 5 weeks followed by another 18 weeks of feeding without diet switch. Two-way ANOVA was used for calculating p values (BAT: * p = 0.0211, **** p < 0.0001, * p = 0.0276, * p = 0.0294, **** p < 0.0001, *** p = 0.0006, and ** p = 0.0046; iWAT: * p = 0.0420; eWAT: *** p = 0.0002). (K) RT-qPCR measuring mRNAs associated with lipolysis in iWAT of the indicated mice received AAV-irisin or -GFP, and fed a HFD for 18 weeks (n = 7, 8). Two-way ANOVA was used for calculating p values. Abbreviations as per Fig. 1. Mean ± SD.* p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, ns: not significant. Significant but irrelevant p values are not indicated. Unless specifically mentioned, experiments were repeated twice with similar results.

Source data