Caloric restriction improves glucose homeostasis, yet increases cardiometabolic risk in caveolin-1-deficient mice

Abstract

Background and purpose: The plasma membrane protein caveolin-1 (CAV-1) has been shown to be involved in modulating glucose homeostasis and the actions of the renin-angiotensin-aldosterone system (RAAS). Caloric restriction (CR) is widely accepted as an effective therapeutic approach to improve insulin sensitivity and reduce the severity of diabetes. Recent data indicate that polymorphisms of the CAV-1 gene are strongly associated with insulin resistance, hypertension and metabolic abnormalities in non-obese individuals. Therefore, we sought to determine whether CR improves the metabolic and cardiovascular (CV) risk factors in the lean CAV-1 KO mice.

Materials/methods: Twelve- to fourteen-week-old CAV-1 knockout (KO) and genetically matched wild-type (WT) male mice were randomized by genotype to one of two dietary regimens: ad libitum (ad lib) food intake or 40% CR for 4 weeks. Three weeks following the onset of dietary restriction, all groups were assessed for insulin sensitivity. At the end of the study, all groups were assessed for fasting glucose, insulin, HOMA-IR, lipids, corticosterone levels and blood pressure (BP). Aldosterone secretion was determined from acutely isolated Zona Glomerulosa cells.

Results: We confirmed that the CAV-1 KO mice on the ad lib diet display a phenotype consistent with the cardiometabolic syndrome, as shown by higher systolic BP (SBP), plasma glucose, HOMA-IR and aldosterone levels despite lower body weight compared with WT mice on the ad lib diet. CAV-1 KO mice maintained their body weight on the ad lib diet, but had substantially greater weight loss with CR, as compared to caloric restricted WT mice. CR-mediated changes in weight were associated with dramatic improvements in glucose and insulin tolerance in both genotypes. These responses to CR, however, were more robust in CAV-1KO vs. WT mice and were accompanied by reductions in plasma glucose, insulin and HOMA-IR in CAV-1KO but not WT mice. Surprisingly, in the CAV-1 KO, but not in WT mice, CR was associated with increased SBP and aldosterone levels, suggesting that in CAV-1 KO mice CR induced an increase in some CV risk factors.

Conclusions: CR improved the metabolic phenotype in CAV-1 KO mice by increasing insulin sensitivity; nevertheless, this intervention also increased CV risk by inappropriate adaptive responses in the RAAS and BP.

Keywords: Aldosterone; Caloric restriction; Cardiometabolic dysfunction; Caveolin; Insulin resistance; The metabolic syndrome.

Figure 1. Experimental design

On day −5 mice of both genotypes were acclimatized in an individual cage with free access to ad lib diet and water. On day 0, WT and CAV-1 KO mice were randomized into ad lib (control for each genotype) or CR groups for 4 wk. The timing for performance of the GTT, ITT, BP measurement, urine collection and sacrifice are indicated.

Figure 2. Body weights and food consumption in CAV-1 KO vs. WT mice

(A) Growth curves for WT and CAV-1 KO mice over 4 weeks of ad lib diet. (B) Growth curves for WT and CAV-1 KO mice over 4 weeks of CR diet. (C) Absolute body weight changes (g) after 4 weeks of ad lib or CR diet. (D) Food intake in WT and CAV-1 KO mice after 4 weeks of ad lib or CR diet. *p < 0.05 and ** p < 0.001 vs. WT on the same diet, ^# p < 0.001 vs. the same genotype on ad lib diet.

Figure 3. Effect of CR on insulin resistance in WT and CAV-1 KO mice

(A)–(B) glucose excursion during the ip-GTT in WT (A) and CAV-1 KO mice (B) (data expressed as % of the highest glucose levels in the ad lib diet group); (C) area under the curve (AUC) during the GTT; (D) glucose levels during the ip-ITT (data expressed as % of fasting glucose); (E) inverse area under the curve (AUC) during the ITT. Data were analyzed using an analysis of variance (ANOVA) of the GTT and ITT profiles assessing the interaction between exposure group (ad lib vs. CR) and time during the test. *p < 0.01 vs. WT on the same diet, ^# p < 0.01 vs. the same genotype on ad lib diet.

Figure 4. Effect of CR on the lipid profile in WT and CAV-1 KO

Triglycerides (A) and total cholesterol (B) levels were measured in all groups after 10 hour fasting. *p < 0.05 vs. WT on the same diet, ^$p<0.001 and ^# p < 0.05 vs. the same genotype on ad lib diet.

Figure 5. Effect of CR on hemodynamic parameters and Na⁺ excretion

SBP levels (A), PP (B), RPP (C) and 24 hr urinary Na⁺ levels (D) in WT and CAV-1 KO maintained on ad lib vs. CR diet. *p < 0.05, ** p < 0.005 vs. WT on the same diet; ^# p < 0.05 vs. the same genotype on ad lib diet.

Figure 6. Effect of CR on RAAS and adrenal function

Circulating aldosterone (A), PRA (B), aldosterone to PRA ratio (C), corticosterone (E), and aldosterone to corticosterone ratio (F) in WT and CAV-1 KO maintained on ad lib vs. CR diet. Aldosterone release in response to 10⁻⁸ M Ang II, 8.7 mM K⁺ and 10⁻¹⁰ M ACTH (D) was measured in Zona Glomerulosa cells acutely isolated from mice in the four experimental groups. *p < 0.05, ** p < 0.005 vs. WT on the same diet; ^# p < 0.05 vs. the same genotype on ad lib diet.

Figure 7. Effect of CR on renal SGK1 (A) and ENaC (B) expression in CAV-1 KO and WT mice

ENaC and SGK1 transcript levels were assessed in kidney tissues from WT ad lib, WT-CR, CAV-1 KO ad lib and CAV-1 KO CR mice. *p < 0.05, ** p < 0.005 vs. WT on the same diet; ^# p < 0.05 vs. the same genotype on ad lib diet.