Fig. 1: Lin28A/B inhibition suppresses insulin-PI3K-mTOR signalling in liver and skeletal muscle. | Nature Communications

Fig. 1: Lin28A/B inhibition suppresses insulin-PI3K-mTOR signalling in liver and skeletal muscle.

From: Pharmacological inhibition of Lin28 promotes ketogenesis and restores lipid homeostasis in models of non-alcoholic fatty liver disease

Fig. 1

a C1632, C4019 and C1243 inhibit Lin28 and derepress let-7 biogenesis. b 7-week-old male C57BL/6J mice received intraperitoneal injections of C1632 (50 mg/kg) or vehicle for 5 consecutive days. Blood glucose was monitored daily (n = 8 mice/group; basal vs 1632 Day 2: *P  = 0.0457 and Day 4: ****P < 0.0001; vehicle vs 1632-Day 4: *P  = 0.0224). c Immunoblot analysis for Lin28A, Lin28B, p-AKT(S473), AKT in liver lysates in treated mice from (b) (Loading control: α-tubulin). d Immunoblot analysis in skeletal muscles from (b) (Loading control: α-tubulin). e Let-7 and miR-16 expression in livers from treated mice (n = 3 mice/group; let-7a: *P  = 0.0270; let-7g: **P = 0.0071). f Immunoblot analysis for Lin28B and INSR in HepG2 cells after C1632 treatment (Loading control: β-Actin). g Let-7 levels in HepG2 cells after C1632 (100 μM) treatment (n = 3 biologically independent samples/group; let-7c: ***P = 0.0004; let-7g: *P = 0.0465). h Glucose uptake was measured by flow-cytometry in HepG2 cells after C1632 (100 μM) or vehicle treatment, followed by NBDG (representative experiment from four biological replicates). i Lactate concentration in medium of HepG2 cells after C1632 (100 μM) or vehicle treatment (n = 4 biologically independent samples; **P = 0.0077). j Immunoblot analysis for Lin28A, Lin28B and INSR in C2C12 myotubes after C1632 treatment (loading control: α-tubulin). Immunoblot analysis for P-AKT (S473) and AKT in C2C12 myotubes after treatment with C1632 (loading control: α-tubulin). k Immunoblot analysis for Lin28A, Lin28B, INSR, p-AKT (S473), AKT, in C2C12 myotubes after transfection with empty vectors (EV), Lin28A/B-overexpression vectors (loading control: α-tubulin). l Immunoblot analysis for Lin28B and INSR in C2C12 myotubes after siRNA transfection against Lin28B (20 nM) (loading control: α-tubulin). Immunoblot analysis for Lin28B and INSR in HepG2 cells after transfection with siLin28B (20 nM) (loading control: β-actin). m Immunoblot analysis for Lin28B, INSR, p-AKT (S473), AKT in C2C12 myotubes after C4019 treatment. Immunoblot analysis for Lin28B, INSR in C2C12 myotubes after treatment with C1243. Densitometry of immunoblots shown in Fig. S1. Values are mean ± SEM. *P < 0.05; **P < 0.01; ***P  <  0.001; ****P  <  0.0001. Groups were compared using two-tailed unpaired Student’s t-test. Source data provided in Source Data file.

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