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. 2015 May;20(3):461-72.
doi: 10.1007/s12192-015-0571-6. Epub 2015 Jan 25.

Genetic Manipulation of Cardiac Hsp72 Levels Does Not Alter Substrate Metabolism but Reveals Insights Into High-Fat Feeding-Induced Cardiac Insulin Resistance

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Genetic Manipulation of Cardiac Hsp72 Levels Does Not Alter Substrate Metabolism but Reveals Insights Into High-Fat Feeding-Induced Cardiac Insulin Resistance

Darren C Henstridge et al. Cell Stress Chaperones. .
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Abstract

Heat shock protein 72 (Hsp72) protects cells against a variety of stressors, and multiple studies have suggested that Hsp72 plays a cardioprotective role. As skeletal muscle Hsp72 overexpression can protect against high-fat diet (HFD)-induced insulin resistance, alterations in substrate metabolism may be a mechanism by which Hsp72 is cardioprotective. We investigated the impact of transgenically overexpressing (Hsp72 Tg) or deleting Hsp72 (Hsp72 KO) on various aspects of cardiac metabolism. Mice were fed a normal chow (NC) or HFD for 12 weeks from 8 weeks of age to examine the impact of diet-induced obesity on metabolic parameters in the heart. The HFD resulted in an increase in cardiac fatty acid oxidation and a decrease in cardiac glucose oxidation and insulin-stimulated cardiac glucose clearance; however, there was no difference in Hsp72 Tg or Hsp72 KO mice in these rates compared with their respective wild-type control mice. Although HFD-induced cardiac insulin resistance was not rescued in the Hsp72 Tg mice, it was preserved in the skeletal muscle, suggesting tissue-specific effects of Hsp72 overexpression on substrate metabolism. Comparison of two different strains of mice (BALB/c vs. C57BL/6J) also identified strain-specific differences in regard to HFD-induced cardiac lipid accumulation and insulin resistance. These strain differences suggest that cardiac lipid accumulation can be dissociated from cardiac insulin resistance. Our study finds that genetic manipulation of Hsp72 does not lead to alterations in metabolic processes in cardiac tissue under resting conditions, but identifies mouse strain-specific differences in cardiac lipid accumulation and insulin-stimulated glucose clearance.

Figures

Fig. 1
Fig. 1
Hsp72 Tg mice are protected from HFD-induced weight gain and have normal heart size. Mice were fed a normal chow (NC) or high-fat diet (HFD) for 12 weeks from 8 weeks of age. a Body weight, b fat mass, c lean mass, and d body fat percentage, n = 8–9. (Body composition characteristics of this cohort of mice were included as part of a larger population of mice in Henstridge et al. (2014). e Heart mass, n = 8–15; f tibia length, n = 7–10; and g heart weight to tibia length ratio, n = 7–10 per group. ***p < 0.001 for dietary effect; p < 0.05, †† p < 0.01 for genotype effect. All data are presented as mean ± SEM
Fig. 2
Fig. 2
On a HFD, Hsp72 Tg mice maintain whole body insulin responsiveness and skeletal muscle glucose clearance but not cardiac glucose clearance. Mice were fed a normal chow (NC) or high-fat diet (HFD) for 12 weeks from 8 weeks of age. a Blood glucose response following an intravenous injection of insulin and b average blood glucose concentration over the 35-min period post insulin/tracer injection. c Cardiac glucose clearance rate, d skeletal muscle glucose clearance (quadriceps), e skeletal muscle glucose clearance (gastrocnemius), and f white adipose tissue (epididymal fat pad) glucose clearance, n = 6–8 per group. *p < 0.05, **p < 0.01, ***p < 0.001 for dietary effect; p < 0.05, †† p < 0.01 for genotype effect; ## p < 0.01 for a direct comparison between WT HFD and Hsp72 Tg HFD groups by a t test. All data are presented as mean ± SEM
Fig. 3
Fig. 3
High-fat feeding causes a decrease in glucose oxidation and an increase in palmitate oxidation rate which is not altered by overexpression of Hsp72. Mice were fed a normal chow (NC) or high-fat diet (HFD) for 12 weeks from 8 weeks of age. a Glucose oxidation rate, n = 6–9 per group. b Palmitate oxidation rate, n = 6–10 per group. The palmitate oxidation rates were calculated in the CO2 component and the acid-soluble metabolite (ASM) fraction, and the CO2 and ASM fractions summed to calculate total oxidation rate. Asterisk indicates trend for a main effect of diet. All data are presented as mean ± SEM. *p < 0.05, ***p < 0.001 for dietary effect. All data are presented as mean ± SEM
Fig. 4
Fig. 4
Lipidomic analysis of cardiac triacylglycerol (TG), diacylglycerol (DG), and ceramide in Hsp72 Tg mice fed a NC or HFD. Mice were fed a NC or HFD for 12 weeks from 8 weeks of age. a Total TG levels and b individual cardiac TG molecular lipid species. c Total DG levels and d individual cardiac DG molecular lipid species. e Total ceramide levels and f individual cardiac ceramide molecular lipid species. n = 5–9 per group. *p < 0.05, **p < 0.01 for dietary effect; p < 0.05 for genotype effect. All data are presented as mean ± SEM
Fig. 5
Fig. 5
At 20 weeks of age, Hsp72 KO do not have a whole body weight phenotype and have normal heart size. Mice were fed a normal chow (NC) or high-fat diet (HFD) for 12 weeks from 8 weeks of age. a Body weight, b fat mass, c lean mass, and d body fat percentage, n = 6–9. e Heart mass, f tibia length, and g heart weight to tibia length ratio, n = 3–6 per group. ***p < 0.001 for dietary effect. All data are presented as mean ± SEM
Fig. 6
Fig. 6
On a HFD, Hsp72 KO mice have similar whole body insulin responsiveness and skeletal muscle and cardiac glucose clearance as wild-type mice. Mice were fed a normal chow (NC) or high-fat diet (HFD) for 12 weeks from 8 weeks of age. a Blood glucose response following an intravenous injection of insulin and b average blood glucose concentration over the 35-min period, n = 4–9. c Cardiac glucose clearance rate, d skeletal muscle glucose clearance (quadriceps), e skeletal muscle glucose clearance (gastrocnemius), and f white adipose tissue (epididymal fat pad) glucose clearance, n = 4–7 per group. *p < 0.05, **p < 0.05, ***p < 0.001 for dietary effect. p < 0.05, †† p < 0.05 for genotype effect. All data are presented as mean ± SEM
Fig. 7
Fig. 7
High-fat feeding causes a decrease in glucose oxidation and an increase in palmitate oxidation rate which is not altered by deletion of Hsp72. Mice were fed a normal chow (NC) or high-fat diet (HFD) for 12 weeks from 8 weeks of age. a Glucose oxidation rate, n = 7–9 per group. b Palmitate oxidation rate, n = 7–8 per group. The palmitate oxidation rates were calculated in the CO2 component and the acid-soluble metabolite (ASM) fraction, and the CO2 and ASM fractions summed to calculate total oxidation rate. c Oxygen consumption rates in isolated cardiac mitochondrial fractions, n = 4–7. ***p < 0.001 for dietary effect. All data are presented as mean ± SEM
Fig. 8
Fig. 8
Cardiac lipidomic analysis of triacylglycerol (TG), diacylglycerol (DG), and ceramide lipids in Hsp72 KO NC or HFD mice. Mice were fed a NC or high-fat diet for 12 weeks from 8 weeks of age. a Total TG levels and b individual cardiac TG molecular lipid species. c Total DG levels and d individual cardiac DG molecular lipid species. e Total ceramide levels and f individual cardiac ceramide molecular lipid species. n = 4–6 per group. *p < 0.05, **p < 0.01, ***p < 0.001 for dietary effect; p < 0.05, †† p < 0.01 for genotype effect. All data are presented as mean ± SEM

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