High fat feeding induces only a minor adipose tissue macrophage infiltration in IL-1βKO mice.

<p>(<b>A</b>) Quantitative real-time PCR analysis of adipose tissue (epididymal fat pad) of <i>IL-1b, il6</i> and <i>tnf</i> (normalized to <i>tbp</i>, <i>18S</i> and <i>36b4</i>). n≥3 per group. (<b>B–D</b>) Representative X20 light microscopy images of adipose tissue stained with H&E or with anti-Mac2 antibody. The mean±SEM number of crown like structures (CLS) per X10 microscopic field was counted as described in Materials and Methods. mRNA levels of <i>f4/80</i>, a macrophage marker, was assessed by quantitative real-time PCR. (<b>E–G</b>) Similar analysis as described above (B–D), but for IL-1βKO-NC and IL-1βKO-HFF mice. n = 3–6 animals per group were included in the analysis. *p<0.05 compared to IL-1βKO-NC; ***p<0.001 compared to WT-NC.</p

Role of adipose IL-1β in hepatocyte insulin resistance as revealed by co-culture approach.

<p>(<b>A</b>) Schematic representation of the fat explants – primary hepatocyte co-culture experimental design. (<b>B</b>) Insulin-stimulated Akt and GSK3 phosphorylation in primary hepatocytes from IL-1RIKO liver co-cultured or not with fat explants from WT-NC or WT-HFF and densitometry analysis of 2–5 mice per group. *p = 0.05 compared to incubation with fat explants from WT-HFF mice. (<b>C</b>) Insulin-stimulated Akt phosphorylation in primary hepatocytes from WT mice co-cultured with fat explants from WT-HFF, IL-1βKO-HFF, or WT-HFF in the presence of IL-1 receptor antagonist (WT-HFF+RA). The right graph depicts densitometry analysis of 7–9 mice per group. *p<0.05 compared to the signal obtained from primary hepatocytes incubated with fat explants from WT-HFF mice.</p

Role of IL-1β in adipose tissue macrophage recruitment, ATM lipid content, and adipose inflammatory profile in dietary obesity.

<p>(<b>A</b>) FACS plots and gating of the stromal-vascular cells (SVCs) to detect adipose tissue macrophages (ATMs). Leucocytes (<b>B</b>), ATMs (<b>C</b>) in adipose tissue of WT-NC (n = 4), WT-HFF (n = 11), IL-1βKO-NC (n = 3) and IL-1βKO-HFF (n = 7). (<b>D</b>) Histogram of lipid content (determined with Bodipy) in representative mice of the 4 mouse groups (<b>E</b>)<b>.</b> Quantitative real-time PCR analysis of M1 or M2- genes in epididymal adipose tissue of (<b>F</b>) WT-NC, WT-HFF (n = 4, 11, respectively), and (<b>G</b>) IL-1βKO-NC and IL-1βKO-HFF (n = 3 and 7, respectively). The expression of each transcript was normalized to <i>tbp</i>, <i>18S</i> and <i>36b4</i> mRNA/rRNA, and a value of 1 was assigned to the normal chow group (NC) of each strain. *p<0.05, compared to NC; **p<0.01 compared to NC ***p<0.001.</p

IL-1β impact on liver and adipose tissue mass and adipose tissue expandability.

<p>(<b>A</b>) Representative Computed Tomography (CT) scans (mid-coronal sections) of WT-NC and WT-HFF mice, and excised epididymal white adipose tissue (eWAT) and livers, and the mean±SEM of their weights. (<b>B</b>) Similar to A, but for IL-1βKO mice. ***p<0.001 compared to NC. (<b>C</b>) Spearman correlation between epididymal fat pads' weight and liver weight in HFF mice. (<b>D</b>) Adipocyte size distribution in WT and IL-1βKO mice, quantified as described in Methods. n = 3–6 mice per group. (<b>E</b>) Quantitative real-time PCR analysis of the indicated genes in epididymal adipose tissue in WT and IL-1βKO mice, respectively. n = 3–6 per group. *p<0.05 compared to IL-1βKO-NC ***p<0.0001 compared to WT-NC.</p

Role of adipose IL-1β in adipose-liver cross-talk as revealed by portally-drained mesenteric adipose tissue transplantation.

<p>(<b>A</b>) Serum IL-1β levels were measured in peripheral (systemic) or portal vein blood in WT mice fed normal chow (WT-NC) or high fat diet (WT-HFF). Connecting lines indicate the paired systemic-portal samples from a single mouse, n = 17–19. Red symbols represent≥20% higher IL-1β level in the portal compared to the systemic blood; (<b>B</b>) Schematic representation of the mesenteric adipose tissue transplantation experimental flow. (<b>C</b>)Portal blood levels of IL-1β were measured in sham-operated (n = 9) and in mice receiving mesenteric adipose tissue transplantation from a littermate WT mouse (Trans-WT, n = 13)*p<0.05. (<b>D, E</b>) Intra-peritoneal pyruvate tolerance test (PTT, 2 gr/Kg body weight) was performed in Sham (n = 9), Trans-WT (n = 13), and in mice receiving transplants from IL-1βKO mice (Trans-IL-1βKO, n = 7) four weeks post-transplantation. Area under the glucose levels curve (AUC) was calculated; *p<0.05 compared to Sham-operated controls.</p