Figure 4. The metabolic effects of auranofin occur independently of miR-30a.

(A) Guide RNAs (gRNA) flanked a 1 kb genomic region containing miR-30a. Non-homologous end-joining (NHEJ) generated miR-30a^−/− (KO). PCR genotyping of the targeted loci in genomic DNA from wild-type (+/+), miR-30a^+/−, miR-30a^−/− mice or ES cells (JM8). (B) qPCR of miR-30a and miR-30c (six month old miR-30a^−/− and littermate controls, n=3/group). (C) Body weight during high fat diet (HFD) for 12 weeks (n=21-22 mice/group). (D) Fasting glucose (n=13/group) and insulin levels (n=8-9/group). HFD-fed wild-type and miR-30a^−/− were i.p. injected with auranofin (AF, 1 mg/kg) for 4 weeks. (E) Body weight gain of HFD wild-type and miR-30a^−/− mice during auranofin treatment. (F) GTT and (G) changes (Δ) in fasting glucose after auranofin (n=4-5/group). (H) ITT and corresponding area under the curve measurements (n=5/group). Asterisks (gray-WT, blue-KO) indicate pre- and post- auranofin changes for each genotype. (I) Reverse phase protein array (RPPA) on eWAT of untreated obese wild-type and miR-30a^−/− mice shown as log₂ fold change KO/wild-type. (J) H&E stained eWAT (scale=50 μm) and immunofluorescent labeling of CD68 with quantification (five images/mouse; n=3/group; scale=100 μm). (K) Expression of inflammatory and fibrosis genes in eWAT after auranofin treatment.

All data are mean ± SEM. *p<0.05 by upaired t-test (B; D; G, vs ΔBG=0 ; I-K) and two-way ANOVA with Tukey’s multiple comparisons test (C; E; F; pre- and post-AF, H)