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. 2022 Nov 15;3(11):100813. doi: 10.1016/j.xcrm.2022.100813

Figure 2.

Figure 2

Liraglutide induces IL-6 signaling and adipocyte browning in mice

(A) Mouse immortalized brown pre-adipocytes seeded at confluency were induced for brown adipogenesis in DMEM/10% fetal bovine serum (FBS) medium containing 50 nM insulin/0.5 mM IBMX, 1 μM dexamethasone, 1 nM 3,5,3′-Triiodothyronine (T3), and 5 μM rosiglitazone for 24 h, were washed and cultured in DMEM/10% FBS medium containing 500 pg/mL mouse IL-6, 100 nM liraglutide, or their combination, for 6 and 24 h. UCP1 expression was measured by western blotting. Arrow: UCP1; ns, nonspecific band. Graph: quantification of band intensity normalized to actin. Shown are mean values ± SEM. ∗p < 0.001, one-way ANOVA.

(B) HFD-fed overweight C57BL/6 females (n = 3) were injected with a single s.c. dose of liraglutide (0.4 mg/kg BW). Plasma was collected at indicated time points after administration and IL-6 levels were measured by ELISA. Shown are mean values ± SEM. ∗p < 0.001, one-way ANOVA.

(C) Left: Echo MRI analysis of HFD-fed overweight C57BL/6 males after 2 and 4 weeks of liraglutide, treatment (0.2 mg/kg) shows a reduction of fat mass in liraglutide-treated mice. Right: mass of isolated individual AT depots at week 4. n = 5. ∗p = 0.05, Student’s t test.

(D) Cold tolerance test in mice from experiment (C) at week 4. ∗p < 0.05, Student’s t test.

(E) Analysis of SAT, VAT, and BAT sections from mice in (C) at week 4. Increased UCP1 expression is indicated (arrows). Adipocytes are revealed by perilipin-1 (PLN1) IF. Nuclei are blue. Scale bar, 50 μm.

(F) Quantification of data from (E). ∗p < 0.05, Student’s t test.

(G) Analysis of VAT from mice in (C) at week 4 by RT-PCR showing relative UCP1 expression (normalized to 18S RNA) increased by liraglutide (Lira).

(H) Analysis of VAT from mice in (C) at week 4 by immunoblotting with the indicated antibodies in two mice per group (randomly selected) showing liraglutide-increased UCP1 and pAMPK signal.

(I) Nine-week-old C57BL/6 males were injected with a single s.c. dose of liraglutide (0.2 mg/kg BW). Analysis of SAT and VAT by RT-PCR after 30 min shows liraglutide-increased relative SOCS3 and ADAM10 expression (normalized to 18S RNA). ∗p < 0.05, ∗∗p < 0.01, Student’s t test.

(J) C57BL/6 males were injected with a single s.c. dose of liraglutide (0.2 mg/kg BW). Analysis of VAT by immunoblotting shows liraglutide-induced pAMPK and pSTAT3 peaking at 120 min and decreasing at 240 min post-injection.