Loss of Nlrp3 Does Not Protect Mice from Western Diet-Induced Adipose Tissue Inflammation and Glucose Intolerance.

We tested the hypothesis that loss of Nlrp3 would protect mice from Western diet-induced adipose tissue (AT) inflammation and associated glucose intolerance and cardiovascular complications. Five-week old C57BL6J wild-type (WT) and Nlrp3 knockout (Nlrp3-/-) mice were randomized to either a control diet (10% kcal from fat) or Western diet (45% kcal from fat and 1% cholesterol) for 24 weeks (n = 8/group). Contrary to our hypothesis that obesity-mediated white AT inflammation is Nlrp3-dependent, we found that Western diet-induced expression of AT inflammatory markers (i.e., Cd68, Cd11c, Emr1, Itgam, Lgals, Il18, Mcp1, Tnf, Ccr2, Ccl5 mRNAs, and Mac-2 protein) were not accompanied by increased caspase-1 cleavage, a hallmark feature of NLRP3 inflammasome activation. Furthermore, Nlrp3 null mice were not protected from Western diet-induced white or brown AT inflammation. Although Western diet promoted glucose intolerance in both WT and Nlrp3-/- mice, Nlrp3-/- mice were protected from Western diet-induced aortic stiffening. Additionally, Nlrp3-/- mice exhibited smaller cardiomyocytes and reduced cardiac fibrosis, independent of diet. Collectively, these findings suggest that presence of the Nlrp3 gene is not required for Western diet-induced AT inflammation and/or glucose intolerance; yet Nlrp3 appears to play a role in potentiating arterial stiffening, cardiac hypertrophy and fibrosis

Liver characterization in WT and <i>Nlrp3</i>^-/- mice fed a control diet versus Western diet.

<p>(A) Liver weight; (B) liver triglycerides (TG); representative histological 20X images stained for H&E. Data are expressed as means ± SE. WT, wild-type; KO, <i>Nlrp3</i> knockout, CD, control diet; WD, Western diet; D, main effect of diet; G, main effect of genotype; DxG, diet by genotype interaction. Significant p values (<0.05) are highlighted in bold.</p

Body weight and composition in WT and <i>Nlrp3</i>^-/- mice fed a control diet versus Western diet.

<p>(A) Weekly body weights; (B) final body weight; (C) final % body fat. Data are expressed as means ± SE. WT, wild-type; KO, <i>Nlrp3</i> knockout, CD, control diet; WD, Western diet; D, main effect of diet; G, main effect of genotype; DxG, diet by genotype interaction. Significant p values (<0.05) are highlighted in bold.</p

Visceral white AT characterization in WT and <i>Nlrp3</i>^-/- mice fed a control diet versus Western diet.

<p>(A) Representative immunohistochemical 10X images of retroperitoneal white AT stained for Mac-2; (B) gene expression in stromal vascular cells isolated from epididymal white AT; (C) average adipocyte size in retroperitoneal white AT; (D) Mac-2 positive immunostained area in retroperitoneal white AT; (E) protein content of Mac-2 via Western blotting in retroperitoneal white AT; (F) protein content of pro-caspase-1 via Western blotting in retroperitoneal white AT; (G) protein content of p10 caspase-1 (cleavage) via Western blotting in retroperitoneal white AT; (H) representative Western blot bans. Data are expressed as means ± SE. WT, wild-type; KO, <i>Nlrp3</i> knockout, CD, control diet; WD, Western diet; D, main effect of diet; G, main effect of genotype; DxG, diet by genotype interaction. Significant p values (<0.05) are highlighted in bold. *denotes p<0.05 in panel B.</p

AT gene expression in WT and <i>Nlrp3</i>^-/- mice fed a control diet versus Western diet.

<p>(A) Visceral white (i.e., retroperitoneal) AT; (B) brown (i.e., interscapular) AT. Data are expressed as means ± SE. WT, wild-type; KO, <i>Nlrp3</i> knockout, CD, control diet; WD, Western diet; D, main effect of diet (p<0.05); G, main effect of genotype (p<0.05). No significant interactions were found.</p

Animal characteristics.

<p>Animal characteristics.</p

Aortic stiffness and cardiac characterization in WT and <i>Nlrp3</i>^-/- mice fed a control diet versus Western diet.

<p>(A) Aortic pulse wave velocity; (B) representative histological cardiac 10X images stained for trichrome blue (arrows point to trichrome blue positive-stained regions); (C) cardiomyocyte diameter; (D) cardiac fibrosis (quantification of trichrome blue positive-stained area). Data are expressed as means ± SE. WT, wild-type; KO, <i>Nlrp3</i> knockout, CD, control diet; WD, Western diet; D, main effect of diet; G, main effect of genotype; DxG, diet by genotype interaction. Significant p values (<0.05) are highlighted in bold.</p

Glucose tolerance testing (GTT) in WT and <i>Nlrp3</i>^-/- mice fed a control diet versus Western diet at 15 and 25 weeks of age.

<p>(A) Glucose levels after glucose injection; (B) glucose area under the curve (AUC) during GTT. Data are expressed as means ± SE. WT, wild-type; KO, <i>Nlrp3</i> knockout, CD, control diet; WD, Western diet; D, main effect of diet; G, main effect of genotype; DxG, diet by genotype interaction. Significant p values (<0.05) are highlighted in bold.</p