A Mouse Model of Metabolic Syndrome: Insulin Resistance, Fatty Liver and Non-Alcoholic Fatty Pancreas Disease (NAFPD) in C57BL/6 Mice Fed a High Fat Diet

Diet-induced obesity in C57BL/6 mice triggers common features of human metabolic syndrome (MetS). The purpose is to assess the suitability of a diet-induced obesity model for investigating non-alcoholic fatty pancreatic disease (NAFPD), fatty liver and insulin resistance. Adult C57BL/6 mice were fed either high-fat chow (HFC, 60% fat) or standard chow (SC, 10% fat) during a 16-week period. We evaluated in both groups: hepatopancreatic injuries, pancreatic islets size, alpha and beta-cell immunodensities, intraperitoneal insulin tolerance test (IPITT) and oral glucose tolerance test (OGTT). The HFC mice displayed greater mass gain (p<0.0001) and total visceral fat pads (p<0.001). OGTT showed impairment of glucose clearance in HFC mice (p<0.0001). IPITT revealed insulin resistance in HFC mice (p<0.0001). The HFC mice showed larger pancreatic islet size and significantly greater alpha and beta-cell immunodensities than SC mice. Pancreas and liver from HFC were heavier and contained higher fat concentration. In conclusion, C57BL/6 mice fed a high-fat diet develop features of NAFPD. Insulin resistance and ectopic accumulation of hepatic fat are well known to occur in MetS. Additionally, the importance of fat accumulation in the pancreas has been recently highlighted. Therefore, this model could help to elucidate target organ alterations associated with metabolic syndrome

Weight cycling enhances adipose tissue inflammatory responses in male mice.

BACKGROUND: Obesity is associated with low-grade chronic inflammation attributed to dysregulated production, release of cytokines and adipokines and to dysregulated glucose-insulin homeostasis and dyslipidemia. Nutritional interventions such as dieting are often accompanied by repeated bouts of weight loss and regain, a phenomenon known as weight cycling (WC). METHODS: In this work we studied the effects of WC on the feed efficiency, blood lipids, carbohydrate metabolism, adiposity and inflammatory markers in C57BL/6 male mice that WC two or three consecutive times by alternation of a high-fat (HF) diet with standard chow (SC). RESULTS: The body mass (BM) grew up in each cycle of HF feeding, and decreased after each cycle of SC feeding. The alterations observed in the animals feeding HF diet in the oral glucose tolerance test, in blood lipids, and in serum and adipose tissue expression of adipokines were not recuperated after WC. Moreover, the longer the HF feeding was (two, four and six months), more severe the adiposity was. After three consecutive WC, less marked was the BM reduction during SC feeding, while more severe was the BM increase during HF feeding. CONCLUSION: In conclusion, the results of the present study showed that both the HF diet and WC are relevant to BM evolution and fat pad remodeling in mice, with repercussion in blood lipids, homeostasis of glucose-insulin and adipokine levels. The simple reduction of the BM during a WC is not able to recover the high levels of adipokines in the serum and adipose tissue as well as the pro-inflammatory cytokines enhanced during a cycle of HF diet. These findings are significant because a milieu with altered adipokines in association with WC potentially aggravates the chronic inflammation attributed to dysregulated production and release of adipokines in mice

Fat pad masses and serum adipokines in the cycles of the diet.

<p>Values are mean and standard error of the mean (n = 8 per group). In the signaled cases (one-way ANOVA and post-hoc test of Bonferroni), <i>P<</i> 0.05, when:</p>[a]<p>different SC counterpart;</p>[b]<p>different HF counterpart,</p>[c]<p>different from SC₂/HF₂; and,</p>[d]<p>different from SC₂/HF₂/SC₂. Q_A[adipocytes] is numerical density per area of adipocytes.</p

Trends of the body mass evolution determined using the allometric model <i>log y = a+b log x</i>.

<p>The regressions show how the standard chow (SC) or the high-fat diet (HF), or the switching between these two diets throughout the experiment changed the body mass of the animals. In the signaled cases (comparison of slopes), <i>P</i><0.05, when: [a] different from SC; [b] different from SC2/HF2; [c] different from SC2/HF2/SC2; [A] different from HF counterpart; § different from 2^nd cycle. The details about the calculated equations as well as the significance of the comparison of their slopes are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039837#pone-0039837-t002" target="_blank">Table 2</a>.</p

Linear regressions of the body mass against the cycles of different diets (in weeks) based on the allometric model with log-transformed data, log y = log a+b log x (where a is the intercept and b is the slope).

<p><i>R</i> is the coefficient of correlation of Pearson, <i>P</i> is the probability. The group abbreviations were defined in the text (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039837#s2" target="_blank">Material and Methods</a>). Ns, not significant.</p

Schematic design of the experiment detailing the formation of the groups and sample size.

<p>SC is standard chow; HF is high-fat diet; n is the number of mice in the sample. The subscript number after the acronym of the group indicates the number of months the diet was consumed by animals. The gray charts represent the final groups studied.</p

Sectional area of adipocytes and photomicrographs of corresponding groups (n = 8 per group).

<p>Values are means with their standard error shown by vertical bars. In the signaled cases (one-way ANOVA and post-hoc test of Bonferroni), <i>P<</i>0.05, when: [a] different from SC counterpart; [b] different from HF counterpart; [c] different from SC₂/HF₂; and [d] different from SC₂/HF₂/SC₂.</p

Adipose tissue expression of IL-6.

<p>At the top are representative western blots with bands that correspond to groups, in order (n = 5 per group). Values are means with their standard errors shown by vertical bars. In the signaled cases (one-way ANOVA and post-hoc test of Bonferroni), <i>P<</i>0.05, when: [a] different from SC counterpart; [b] different from HF counterpart; [c] different from SC₂/HF₂; and [d] different from SC₂/HF₂/SC₂.</p

Food intake, feed efficiency, blood total cholesterol, triglycerides, oral glucose tolerance test (OGTT, a.u.c is the area under de curve in arbitrary units), serum insulin of groups after the cycles of diets.

<p>SC is standard chow; HF is high-fat diet; n is the number of mice in the sample. The subscript number after the acronym of the group indicates the number of months the diet was consumed by animals. Values are means and standard error of the mean (n = 8 per group In the signaled cases (one-way ANOVA and post-hoc test of Bonferroni), <i>P<</i>0.05, when:</p>[a]<p>different from SC counterpart;</p>[b]<p>different from HF counterpart,</p>[c]<p>different from SC₂/HF₂;</p>[d]<p>different from SC₂/HF₂/SC₂.</p

Adipose tissue expression of adiponectin. In the top, representative western blots with bands corresponding to groups, in order (n = 5 per group).

<p>Values are means with their standard errors shown by vertical bars. In the signaled cases (one-way ANOVA and post-hoc test of Bonferroni), <i>P<</i>0.05, when: [a] different from SC counterpart; [b] different from HF counterpart; [c] different from SC₂/HF₂; and [d] different from SC₂/HF₂/SC₂.</p