Intestinal Epithelial-Derived Exosomes Under Cold Stimulation Promote Adipose Thermogenesis

Figures

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  Figure 1. IEC-Exo under cold stimulation promote adipose thermogenesis in mice. (A) Representative transmission electron microscopy images of IEC-Exo. (B) Nano sight-nanoparticle tracking analysis of IEC-Exo (n = 3). (C) Western blotting of exosomal markers CD9, CD63, TSG101, and IEC GPA33. (D) Fluorescence of DiR-labeled IEC-Exo detected by IVIS Spectrum in vivo imaging system in mice. (E–L) IEC-Exo was collected from 8-week-old mice after 7 days of cold exposure at 4 °C (Cold IEC-Exo), and IEC-Exo from mice raised at room temperature (RT IEC-Exo) was used as the control. Male C57BL/6J mice fed with NCD were administered RT IEC-Exo or Cold IEC-Exo at a concentration of 2 μg/g body weight through tail vein injection. Injections were conducted once a day for 7 consecutive days (n = 6). (E) Food intake. (F) Mice body weight before sacrifice. (G) Organ weight. (H) Mice body temperature after cold stimulation. (I,J) Representative pictures of body surface temperature of mice and the statistics of the highest body surface temperature. (K,L) Relative mRNA expression of thermogenesis and fatty acid oxidation-related genes in BAT and sWAT. Each point on the column represents a separate data point.…

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  Figure 2. IEC-Exo under cold stimulation promote the thermogenesis of primary adipocytes. Relative mRNA expression of thermogenesis (A) and fatty acid oxidation-related (B) genes (n = 6). (C,D) The OCR and proton leak (n = 6). The uptake of BODIPY (E) and 2NBDG (F) (n = 4). Each point on the column represents a separate data point. Data are presented as means ± SEM. * p < 0.05, ** p < 0.01 by T-TEST.

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  Figure 3. miR-674-3p in Cold IEC-Exo promotes thermogenesis in adipocytes. (A) Differentially expressed microRNAs in RT IEC-Exo and Cold IEC-Exo (n = 3). (B–D) The expression levels of miR-27a-5p, miR-139-3p, and miR-674-3p in primary brown adipocytes after transfection with miR-27a-5p, miR-139-3p, and miR-674-3p mimics, respectively (n = 6). (E) The mRNA expression of Ucp1 in primary brown adipocytes after transfection with miR-27a-5p, miR-139-3p, and miR-674-3p mimics, respectively (n = 6). (F) Relative mRNA expression of thermogenesis and fatty acid oxidation-related genes in primary brown adipocytes after transfection with miR-674-3p mimics (n = 6). (G,H) The OCR and proton leak (n = 6). (I) The uptake of BODIPY (n = 4). Each point on the column represents a separate data point. Data are presented as means ± SEM. * p < 0.05, ** p < 0.01 by T-TEST.

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  Figure 4. In situ overexpression of miR-674-3p in BAT and sWAT promotes thermogenesis in mice. (A–F) BAT and sWAT were injected in situ with AAV-miR-674-3p (n = 6). Mice were fed with NCD and experiments were conducted. The mice injected with Vehicle in situ were used as the control (n = 6). (A) Overexpression efficiency. (B) Mice body weight. (C) Organ weight. (D) Mice body temperature after cold stimulation. (E,F) Relative mRNA expression of thermogenesis and fatty acid oxidation-related genes in BAT and sWAT. (G–O) BAT and sWAT were injected in situ with AAV-miR-674-3p. Three weeks after recovery, the mice were fed with HFD for 12 weeks (n = 8). The mice injected with Vehicle in situ were used as the control (n = 8). (G) Overexpression efficiency. (H) Mice body weight. (I) Organ weight. (J,K) Glucose tolerance test and area under the curve (AUC). (L) Insulin tolerance test. (M) Mice body temperature after cold stimulation. (N,O) Relative mRNA expression of thermogenesis and fatty acid oxidation-related genes in BAT and sWAT. Each point on the column represents a separate data point. Data are presented as means ± SEM. * p < 0.05, ** p < 0.01 by T-TEST.