Effects of high fat diet on lipid accumulation, oxidative stress and autophagy in the liver of Chinese softshell turtle (Pelodiscus sinensis)

https://doi.org/10.1016/j.cbpb.2019.110331Get rights and content

Highlights

  • High fat diet feeding induced lipid deposition and increased ROS level in the liver of Chinese softshell turtle;

  • The decreasing of enzyme activities and gene expression of antioxidant enzymes lead to oxidative stress.

  • Signaling molecules (Nrf2, Keap1, mTOR, S6K1) mediated the down-regulation of expression of antioxidant enzyme gene.

  • High fat diet feeding promote autophagy flux through inducing the expression of autophagy genes.

Abstract

The present study was performed to determine the effect of high fat diet in lipid accumulation, oxidative stress and autophagy, and to explore the underlying molecular mechanism of high fat diet induced hepatic oxidative damage in Chinese softshell turtle. To this end, the control group were fed a normal fat diet (NFD, 6.38% lipid) and the experimental group were bred high fat diet (HFD, 13.89% lipid) for eight weeks. Lipid accumulation, oxidative stress and autophagy, as well as the mRNA expression of genes related to the antioxidant system were determined in the liver. Results showed that high fat diet not only exacerbated lipid accumulation in the liver and serum through increasing contents of triglyceride, total cholesterol and low-density lipoprotein and decreasing content of high-density lipoprotein, but also induced liver injury through increasing activities of alanine aminotransferase and aspartate aminotransferase in the serum. In addition, the experimental subject induced oxidative injury for the increase of reactive oxygen species, malondialdehyde and protein carbonyl contents and the reduction of glutathione contents, anti-superoxide anion capacity and catalase, total superoxide dismutase, glutathione peroxidase, glutathione-S transferase activities. Meanwhile, antioxidant-related signaling molecule expression were also decreased, which might attribute to regulate antioxidant-related signaling molecule. On top of that, it indicated promote the occurrence of liver autophagy via up-regulating expressions of AMP activated protein kinase, UNC-51-like kinase 1, Microtubule-associated proteins 1A/1B light chain 3 and down-regulating gene expression of mammalian target of rapamycin. In conclusion, high fat diet could enhance lipid accumulation in the liver and serum, lead to liver injury and oxidative damage, impair liver antioxidant capacity, regulate antioxidant-related signaling molecule expression and activate hepatic autophagy.

Introduction

In recent years, high fat diet has been widely used in aquaculture to provide more energy through lipids rather than proteins (Boujard et al., 2004). However, excessive intake of large amounts of lipids can lead to obesity (Itziar et al., 2010). In mammals, nonalcoholic fatty liver disease is partially associated with long-term high fat intake, which is often accompanied with obesity, hyperlipidemia, and insulin resistance (Marchesini et al., 1999; Marrero et al., 2002; Elizabeth, 2007). In aquatic animal, high fat diet also induced hepatic steatosis due to unnecessary fat deposition (Zhang et al., 2014; Lu et al., 2016). Excess lipid accumulation in tissues such as liver could lead to tissues injury.

The link between high fat diet and disease progression is caused by oxidative imbalance, which mediated by excessive production of reactive oxygen species (ROS) (Maithili et al., 2014). In order to prevent oxidative damage, animals have developed an antioxidant system (Martínez-Álvarez et al., 2005). Aquatic animal's antioxidant system is generally composed of nonenzymatic compounds (GSH) and antioxidant enzymes (involving glutathione peroxidase (GPX), glutathione-S transferase (GST), superoxide dismutase (SOD) and catalase (CAT)) (Martínez-Álvarez et al., 2005). The antioxidant enzymes play a critical role in eliminating ROS (Chen et al., 2013). The expression of antioxidant enzyme genes is regulated by a variety of transcription factors. NF-E2-related nuclear factor 2 (Nrf2) is a key transcription factor that binds to antioxidant response elements (ARE) and induce the expression of antioxidant enzyme genes, such as SOD, CAT and GPX (Kobayashi and Yamamoto, 2006; Giuliani et al., 2017). Kelch-like ECH-associated protein 1 (Keap1) is identified as Nrf2-binding protein that inhibits Nrf2 translocation to the nucleus (Ma, 2013). Feng et al. (2017) reported that mTOR and its downstream target ribosome protein S6 kinase 1(S6K1) were involved in the up-regulation of the expression of Nrf2 in the gill of fish.

Autophagy is an evolutionarily conserved process involving organelles and catabolic cytoplasmic proteins (Xie and Klionsky, 2007). The process of autophagy is strictly controlled to maintain a balance between synthesis and reduction of cellular components, which is important to cell development, homeostasis and growth (Levine and Klionsky, 2004). Autophagy play a key role in the redistribution of energy from unnecessary processes to key processes of survival (Akiko et al., 2004). In addition, autophagy is essential to maintain the quality of proteins and organelles in the cell of liver (Masaaki et al., 2005), brain (Masaaki et al., 2006; Taichi et al., 2006) and heart (Atsuko et al., 2007). Autophagy is up-regulated when cells intend to destroy damaged cytoplasmic component, such as during accumulation of protein and infection (Beth and Guido, 2008).

Chinese softshell turtle Pelodiscus sinensis, with high nutrition and medicinal value, regarded as a water treasure in China (Jia et al., 2015). In addition, it is considered to be an important commercial farmed aquatic species, and the current annual output exceeds 200,000 tons in China (Li et al., 2008). High fat diet is used to improve Chinese softshell turtle growth performance and reduce feed costs in aquaculture. However, the using of high fat diet could lead to some detrimental effects to aquatic species. In this study, we provide molecular, enzyme activity and electron microscopy insights into how high fat diet regulates lipid accumulation, hepatic oxidative stress and autophagy in Chinese softshell turtle.

Section snippets

Animals and experimental design

Chinese softshell turtles were purchased from Tongxin turtle ecological breeding professional cooperative in Dingcheng District, Changde City. Feeding experiment at Changsha College (Changsha, Hunan, China). Chinese softshell turtles were acclimated for at least 2 weeks and fed with normal fat diet (6.38% lipid). Animal care conditions were similar to those described previously (Huang and Lin, 2002; Huang et al., 2015; Huang and Lin, 2015). At the beginning of the trial, 24 uniformly sized

Liver and serum biochemical parameters

Oil red O staining of livers in Chinese softshell turtle fed different diets were shown in Fig. 1. After 8 weeks of high fat diet feeding, the liver showed much more lipid droplets (Fig. 1). The contents of TG, TC and LDL were increased significantly and the content of HDL was decreased significantly in the serum and liver of HFD compared with NFD groups (P < .05) (Fig. 2A-B and Fig. 3A–B). In comparison with NFD group, the activities of ALT and AST in the serum were increased significantly in

Effects of high fat diet on liver and serum lipid accumulation and liver injury

The fundamental biological functions of lipids include signal transmission, energy storage and structural components of cell membranes (Fahy et al., 2009; Shankar et al., 2011). Excessive ingestion of lipids may lead to excessive lipid deposition in the liver, which is detrimental to growth of aquatic animals and decreases the anti-stress ability of aquatic animals (Reiter et al., 2000). Oil red O staining indicated that high fat diet can cause lipid accumulation in the liver of Chinese

Declaration of Competing Interest

We declare that we have no financial and personal relationships with other people or organizations that can inappropriately influence our work, there is no professional or other personal interest of any nature or kind in any product, service and company that could be construed as influencing the position presented in the manuscript entitled “Effects of high fat diet on lipid accumulation, oxidative stress and autophagy in the liver of Chinese softshell turtle (Pelodiscus sinensis)”.

Acknowledgements

This work was supported by the Key Research and Development Project of Hunan Province (2018NK2074) and Science and Technology Plan General Project of Changsha (k1705044).

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