Taxifolin ameliorates DEHP-induced cardiomyocyte hypertrophy via attenuating mitochondrial dysfunction and glycometabolism disorder in chicken

Highlights

• DEHP-induced cardiac hypertrophy is ameliorated after TAX treatment in vitro.

• DEHP induces cardiac hypertrophy via the IL-6/JAK/STAT3 pathway in vitro.

• TAX alleviates DEHP-induced glycometabolism disorder via the IGF1/PI3K pathway.

• TAX mitigates DEHP-induced mitochondrial dysfunction via the PPARs/PGC-1α pathway.

Abstract

Di-(2-ethylhexyl) phthalate (DEHP) is a prevalent environmental contaminant that severely impacts the health of human and animals. Taxifolin (TAX), a plant flavonoid isolated from yew, exerts protective effects on cardiac diseases. Nevertheless, whether DEHP could induce cardiomyocyte hypertrophy and its mechanism remains unclear. This study aimed to highlight the specific molecular mechanisms of DEHP-induced cardiomyocyte hypertrophy and the protective potential of TAX against it. Chicken primary cardiomyocytes were treated with DEHP (500 μM) and/or TAX (0.5 μM) for 24 h. The levels of glucose and adenosine triphosphate (ATP) were detected, and cardiac hypertrophy-related genes were validated by real-time quantitative PCR (qRT-PCR) and Western blot (WB) in vitro. The results showed that DEHP-induced cardiac hypertrophy was ameliorated by TAX, as indicated by the increased cardiomyocyte area and expression of atrial natriuretic peptide (ANP), natriuretic peptides A-like (BNP) and β-myosin heavy cardiac muscle (β-MHC). Furthermore, DEHP induced cardiac hypertrophy via the interleukin 6 (IL-6)/Janus kinase (JAK)/signal transducer and activator of transcription 3 (STAT3) pathway in vitro. In addition, DEHP disrupted mitochondrial function and glycometabolism by activating the insulin-like growth factor 1 (IGF1)/phosphatidylinositol 3-kinase (PI3K) pathway and the peroxisome proliferator activated receptors (PPARs)/PPARG coactivator 1 alpha (PGC-1α) pathway to induce cardiac hypertrophy in vitro. Intriguingly, those DEHP-induced changes were obviously alleviated by TAX treatment. Taken together, cardiac hypertrophy was induced by DEHP via activating the IL-6/JAK/STAT3 signaling pathway, triggering glycometabolism disorder and mitochondrial dysfunction in vitro, can be ameliorated by TAX. Our findings may provide a feasible molecular mechanism for the treatment of cardiomyocyte hypertrophy induced by DEHP.

Introduction

Di-(2-ethylhexyl) phthalate (DEHP) is omnipresent and widely used in a variety of polyethylene plastic products including industrial plastic, medical devices, pharmaceuticals and food containers (Kelley et al., 2012; Koch et al., 2006). Monitoring results indicate that DEHP is widely found in air, water, food, soil, plants and sediment (Arukwe et al., 2017). Due to its wide usage it is constantly released into the environment, especially since DEHP is easily separated from plastics and seriously pollutes food. Some studies have confirmed that plasticizer residues can also be detected in poultry. As chicken muscles are one of the main protein sources of humans, DEHP could accumulate in a human body and possibly cause chronic poisoning (Talsness et al., 2009). As an early warning animal specimen for toxin residues in the ecosystem, chickens can accurately and quickly respond to fluctuations in the concentration of toxic substances in the environment, which is manifested as damage to the body of the chicken. As one of the most sensitive poultry in the environment, the toxic effects of exogenous toxicants on chicken have been gradually studied (Wood and Bitman, 1980). Previous studies have shown that DEHP exposure has adverse effects on the central nervous system (Lin et al., 2011), kidneys (Rothenbacher et al., 1998), liver (Lutz, 1986) and lungs (Rosicarelli and Stefanini, 2009). Phthalates may induce heart rate variability and cardiovascular reactivity to disrupt cardiovascular health (Schulz et al., 1975). Clinical studies have also shown that plasticizers can induce the sudden death of birds because they affect the cardiovascular system, and myocardial regulation of the heat shock response is related to cardiac toxicity of DEHP on quail (Wang et al., 2019a,b). In recent years, despite a plethora of in vitro evidence showing the potential toxicity of DEHP on the heart, its cardiac toxicology on the chicken has yet to be fully defined. Thus, considering the lack of preventive strategies, knowledge of the mechanisms of cardiac disease induced by DEHP requires urgent attention.

Taxifolin (TAX), also known as dihydroquercetin, is a bioflavonoid essence extracted from the root of larch and yew, which has long been used in the clinical treatment of coronary heart disease and cardiovascular diseases. Increasing numbers of studies have proven that TAX exhibits a variety of biological effects, including anticancer, anti-inflammatory, anti-virus, antibacterial and antioxidative properties (Gupta et al., 1971; Kuang et al., 2017; Manigandan et al., 2015). TAX can inhibit cardiac apoptosis by using its antioxidant functions to ameliorate diabetic cardiomyopathy in streptozotocin-induced diabetic mice (Sun et al., 2014). Additionally, a previous study has shown that TAX can abolish hypoxia-induced cardiomyocyte damage via the hypoxia-inducible factor-1 (HIF1-a)/heme oxygenase-1 (HO-1)/autophagy pathway and thus improve cell viability (Lin et al., 2017). The production of reactive oxygen species (ROS) and nitric oxide (NO) during the process of cerebral ischemic reperfusion (CI/R)-injured brain in rats was significantly inhibited by TAX through activating the nuclear factor of erythroid 2-like 2 (Nrf2) antioxidative stress pathway (Yea-Hwey et al., 2006). Tang et al. found that TAX treatment attenuated cardiac I/R injury by regulating the mitochondrial apoptosis pathway and oxidative stress (Tang et al., 2019). TAX could be a potential candidate for curing cardiac hypertrophy and ventricular fibrosis. However, the effect of TAX on cardiac hypertrophy and its related molecular mechanism are still unclear.

Cardiac hypertrophy is an abnormal increase or thickening of the myocardium caused by an increase in the size of cardiomyocytes and other changes in cardiac components, which is a common cause of heart failure with high morbidity and mortality worldwide (Joerg and Molkentin, 2006). In the hypertrophic myocardium, the possible signaling pathways have been investigated in various cardiac diseases (Huss and Kelly, 2004). The interleukin 6 (IL-6)/Janus kinase (JAK)/signal transducer and activator of transcription 3 (STAT3) pathway is reported to be critically involved in the development of cardiomyocyte hypertrophy, especially JAK and STAT3, that have been considered to be essential regulators of a hypertrophic response (Wu and Liu, 2008; Zhang et al., 2007). Glycometabolism is an important metabolic mechanism in mammals and poultry. GLUT1,4-mediated cardiac glucose uptake is associated with cardiac hypertrophy and insulin-like growth factor 1 (IGF1)-dependent activation of phosphatidylinositol 3-kinase (PI3K) signaling is used to meet the energy demands of hypertrophic cardiomyocytes (Kaczmarczyk et al., 2003; Olianas et al., 2011). Related studies have shown that glycometabolism disorders could lead to an increase of glucose uptake and glycolysis in patients with pathological cardiac hypertrophy. Mitochondrial damage along with a decrease of ATP generation may accelerate cardiac hypertrophy (Zhou et al., 2013). Therefore, we intended to determine the exact mechanism of mitochondrial dysfunction and glycometabolism in cardiac hypertrophy.

We hypothesized that TAX may effectively ameliorate DEHP-induced cardiac hypertrophy via attenuating mitochondrial dysfunction and glycometabolism disorders in vitro. To validate our hypothesis, the current study was performed to explore the related genes of mitochondrial biosynthesis function, glucose metabolism and cardiac hypertrophy by detecting the glucose level and cell size in vitro. Ultimately, we found that TAX could prevent DEHP-induced cardiac hypertrophy through modulations of mitochondrial function and glycometabolism in chicken cardiomyocytes.