Hypoxia training improves hepatic steatosis partly by downregulation of CB1 receptor in obese mice

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Highlights

  • Hypoxia training (HT) can reduce body weight and relieve hepatic steatosis.

  • HT down-regulates CB1 expression and endocannabinoid 2-AG contents.

  • HT improves hepatic steatosis partly by inhibition of CB1 mediated fat synthesis.

Abstract

Hypoxia training (HT) can reduce body weight and improve fatty liver. However, the mechanism is not clear. A previous study indicated that HT-induced weight loss might be associated with the endocannabinoid system (ECS), which has also been reported recently to be involved in the persistent lipid mediators after weight loss. The present study investigated the effects of HT, a new prospective weight-loss method, on nutritionally obese mice and demonstrated that HT significantly reduced body weight, fat mass, transcriptional expression of liver endocannabinoid receptor 1 (CB1), biosynthetic enzyme diacylglycerol lipase α (DAGLα) and improved the transcriptional expression of degrading enzyme monoacylglycerol lipase (MAGL). Liver endocannabinoids 2-arachidonoylglycerol (2-AG) but not anandamide (AEA) was evidently decreased in response to HT. Simultaneously, HT significantly reduced liver index, serum alanine aminotransferase (ALT) and liver fat contents. Western blot showed decreased expression of liver CB1, sterol regulatory element-binding protein-1 (SREBP-1), peroxisome proliferator-activated receptor γ (PPARγ) and increased expression of adipose triglyceride lipase (ATGL) and carnitine palmitoyltransferase-1 (CPT-1) levels after HT. However, intraperitoneal injection of CB1 receptor agonist WIN55212-2 offset the benefits by which HT reduced hepatic fat synthesis, with significant increased protein expression of SREBP-1 and PPARγ. Taken together, these findings reported the alleviation of obesity and hepatic steatosis through HT and provided a putative molecular mechanism by inhibiting the CB1-mediated fat synthesis.

Graphical abstract

Note: Blue font, effects of hypoxia training; red font, effects of the CB1 receptor agonist.

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Introduction

The obesity epidemic is closely associated with the rising prevalence and severity of nonalcoholic fatty liver disease (NAFLD) [1]. There is growing evidence that the endocannabinoid system (ECS) is related to hepatic lipid metabolic disorders [2]. As important mediators of metabolism regulation, the ECS is a complex signaling cascade consisting of cannabinoid receptors, ligands and metabolic enzymes and is expressed in various metabolic organs of the obese groups [3].

Traditional HT is used to increase exercise performance. Recently, various studies confirmed that normobaric HT causes more weight loss than normobaric normoxic training in obese groups [[4], [5], [6]]. However, few studies have focused on the effects and mechanism of HT on the lipid metabolism in the liver, especially in obese individuals. What has been established is that intermittent hypoxia (FIO2 cycles from 21% to 7% every 30s) exacerbates insulin intolerance and hepatic steatosis [7], while liver hypoxia-inducible factor-1α (HIF-1α) suppresses the excessive lipid accumulation in the mice of alcoholic fatty liver [8]. Furthermore, only a few studies have included the effects of hypoxia exposure combined with aerobic training on the lipid metabolic changes in the liver.

In this study, we explored that among the traditional methods, whether the new prospective weight loss method — hypoxia training (HT), would relieve the over-activation of liver ECS and improve lipid metabolism. Our previous study has demonstrated that serum endocannabinoids levels are associated with “Living High-Training Low”-induced weight loss [4]. Herein, we further explored the effects of HT (simulation of “Living High-Training Low”) on hepatic lipid metabolism and investigated the putative role of liver endocannabinoid receptor CB1 in nutritional obese mice. We hypothesized that HT might improve lipid metabolism through the downregulation of ECS components in the liver of obese mice. Therefore, this study will help to understand the mechanism underlying HT-regulated liver lipid metabolism.

Section snippets

Animals and experimental procedures

5-week-old C57Bl/6J male mice were fed HFD (D12492, Research Diets)) for 10 weeks, and the obese mice were selected and randomly divided into three groups (n = 9 for each group): obese control group (OB), HT, and HT plus CB1 receptor agonist WIN55212-2 (HT + WIN). All mice were fed HFD during the 4-week intervention period. Mice in all groups were fasted for 12 h followed by sacrifice after the final intervention session. Blood samples were allowed to stand for 2 h at 4 °C before centrifugation

Effects of HT on phenotypic changes in obese mice and the influence of CB1 receptor agonist

HT significantly reduced BW (from week 1–4, P < 0.01 at each time point; Fig. 1A) and FM (P < 0.05, Fig. 1B) as compared to the OB group. In addition, 4 weeks of HT did not affect the level of HDL-C (Fig. 1D), LDL-C (Fig. 1E), and TG (Fig. 1F) significantly. The HE-stained sections showed similar morphological changes among the three groups (Fig. 1G). Nonetheless, we found that irrespective of the CB1 receptor agonist, HT-induced phenotypic changes did not vary significantly. Previous studies

Discussion

In this study, we explored the promising weight-loss method, HT, an intervention that simulated the “Living High-Training Low” model on the liver ECS and lipid metabolism process. Under high-fat feeding conditions, HT partially improved the disorder of lipid metabolism and reversed the over-activation of liver ECS. Furthermore, we demonstrated that the improvement of hepatic steatosis by HT was associated with reduced fat synthesis and accelerated fat transport, hydrolysis, and fatty acid

Conclusion

In conclusion, we confirmed the benefits of HT on the weight loss and demonstrated that HT significantly inhibited the over-activation of liver ECS and rescued hepatic steatosis, putatively via the downregulation of CB1→SREBP-1/PPARγ signaling pathway in nutritionally obese mice. Thus, the current study provided a new perspective on the role of ECS in HT-mediated liver lipid metabolism. Further studies would focus on the hypoxia- and training factor-regulated ECS in obesity.

Declaration of competing interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgement

This work was supported by Fundamental Research Funds for the Central Universities (22120190127) and the National Natural Science Foundation of China (81472148). Thanks to Professor Wang Ru’s research team (Shanghai University of Sport) for their help and support.

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