Black mulberry (Morus nigra L.) polysaccharide ameliorates palmitate-induced lipotoxicity in hepatocytes by activating Nrf2 signaling pathway

Abstract

Black mulberry (Morus nigra L.) has shown health benefits against metabolic disorders. Lipotoxicity is considered as a potentially cause of metabolic syndrome, and there is no effective treatment. However, the protective effect and its mechanism of black mulberry against lipotoxicity are unclear. In this study, three polysaccharide fractions (BP1, BP2, BP3) were isolated from black mulberry by stepwise precipitation with 30%, 60%, and 90% of ethanol and analyzed by GPC, HPLC and FT-IR methods. BP1 exhibited a better protective effect than BP2 and BP3 on palmitic acid (PA)-induced lipotoxicity in HepG2 cells. BP1 effectively reduced PA-induced lipotoxicity by eliminating accumulation of ROS, improving mitochondrial function, reversing glutathione depletion and enhancing antioxidant enzyme activities. Mechanistically, BP1 activated the Nrf2 signaling pathway, a master regulator of the antioxidant defense system, through increasing Nrf2 nuclear translocation and phosphorylation. Collectively, these results demonstrate that BP1 has the great potential for applications in lipid disorders.

Introduction

Palmitic acid (PA) is a kind of saturated fatty acid widely existing in animal cell and adipose tissue, and it is also the main component of refined palm oil [1,2]. Generally, it is obtained from diet or synthesized endogenously through de novo synthetic reaction in human body [3,4]. However, increasing evidence indicates that PA-induced lipotoxicity is closely related to the pathological processes of oxidative stress, ROS overproduction and hepatocytes dysfunction, which also resulted in metabolism disorder and many diseases including obesity, diabetes, cardiovascular diseases and cancer [[5], [6], [7]]. At present, although some clinical drugs could promote lipid oxidation, mitigate ectopic fat accumulation, prevent or delay the damage caused by lipotoxicity, their potential side effects could not be neglected. This limits the long-term use of clinical treatment. Therefore, it is urgent to develop a natural, safe and effective agent to prevent lipotoxicity-associated metabolic disorders.

Polysaccharide, a kind of high molecular weight polymer, is derived from natural products and consists of many kinds of monosaccharides through glycosidic bonds [8]. In recent years, polysaccharides isolated and purified from healthy food, such as Ginseng, Hirsutella sinensis, Zizyphus Jujuba, and Chinese wolfberry have received increasing attentions [[9], [10], [11], [12]]. Moreover, epidemiological data have shown that plant polysaccharides have beneficial biological effects to ameliorate inflammation and oxidative stress, especially the cytotoxicity [[13], [14], [15]]. Xiong C et al. reported that MIP, a crude polysaccharide extracted from Morchella importuna, prevented PC12 cells from H₂O₂-induced cytotoxicity by reducing oxidative stress [16]. Kim K et al. found that aloe polysaccharides (APs) have a protective effect on free fatty acid-induced lipotoxicity in pancreatic β-cells [17]. In addition, Rubus chingii Hu-derived polysaccharides ameliorated palmitic acid-induced lipotoxicity in HepG2 cells [18]. Morus nigra L. (black mulberry), as a functional food with high edible and medicinal value, is often used in immune regulation, anti-inflammation and other purposes [19]. However, it remains unclear whether the black mulberry polysaccharide has a protective effect against hepatic lipotoxicity.

In this study, three different polysaccharides fractions were extracted from black mulberry by graded precipitation with 30%, 60% and 90% ethanol and analyzed by HPLC, FT-IR, GPC methods. Subsequently, the protective effect of black mulberry polysaccharides on PA-induced lipotoxicity in HepG2 cells and its potential molecular mechanism were investigated. In accordance with the obtained research results, black mulberry polysaccharide plays an important role in maintaining the normal redox status of hepatocytes and alleviating the disorder of lipid metabolism. Therefore, black mulberry polysaccharide has a potential ability to be used as a safe and efficient natural detoxification agent for PA-induced lipotoxicity.