Cardio-protective effect of tetrahydrocurcumin, the primary hydrogenated metabolite of curcumin in vivo and in vitro: Induction of apoptosis and autophagy via PI3K/AKT/mTOR pathways

https://doi.org/10.1016/j.ejphar.2021.174495Get rights and content

Highlights

  • Tetrahydrocurcumin effectively prevents myocardial ischemia-reperfusion injury in vivo.

  • Tetrahydrocurcumin protects against hypoxia and reoxygenation-induced H9c2 injury in vitro.

  • Tetrahydrocurcumin attenuates oxidative stress, apoptosis and autophagy.

  • Tetrahydrocurcumin exerts cardio-protection partially via activating the PI3K/AKT/mTOR pathways.

Abstract

Tetrahydrocurcumin (THC) is an essential metabolite of curcumin, a major active component of the Curcuma species, which have been used traditionally for the treatment of cardiovascular diseases. The PI3K/AKT/mTOR signaling pathways serve a vital role during myocardial ischemia-reperfusion (MI/R) injury. The aim of the present study was to investigate the cardioprotective potential and mechanism of THC. In the in vivo study, an animal model of MI/R was induced by coronary occlusion. Results indicated that THC (50 mg/kg/day) protected the rat hearts from MI/R-induced heart failure by increasing ejection fraction (EF) and fractional shortening (FS) and decreasing left ventricular end systolic diameter (LVESD) and left ventricular end systolic volume (LVESV). THC also reduced myocardial infarct size and apoptosis. Furthermore, H9c2 cells were incubated with THC (20 μM) to explore its potential effect following exposure to hypoxia and reoxygenation (H/R). THC post-treatment significantly augmented cell viability and prevented lactate dehydrogenase (LDH) release after H/R exposure. THC effectively improved antioxidant activity by increasing SOD and CAT activities and decreasing MDA level. THC also enhanced mitochondrial membrane potential, inhibited apoptotic cell death, diminished the Bax/Bcl-2 ratio and cleaved caspase-3 level relative to the H/R model. In addition, THC effectively decreased Beclin1 expression and LC3 II/LC3 I ratio, but increased p62 expression, compared with the H/R model group, and decreased the formation of H/R-induced autophagosomes and autolysosomes. Furthermore, THC promoted the phosphorylation of PI3K/AKT/mTOR and induced the expression of hypoxia-inducible factor 1α (HIF-1α) after H/R. However, these effects on H9c2 cells were notably abolished by the PI3K inhibitor LY294002 and mTOR inhibitor rapamycin. In conclusion, THC effectively inhibited H/R-induced autophagy and apoptosis via, at least partially, activating the PI3K/AKT/mTOR pathways. THC might have the potential to be further developed into a potential candidate for the treatment of MI/R injury.

Introduction

Acute myocardial infarction (AMI) is a significant threat to health, and notable cause of death worldwide (Damiani et al., 2015; Zhang and Ren, 2014). Currently, effective treatments for the management of AMI mainly include reperfusion of the affected region by thrombolytic therapy or angioplasty. Nevertheless, ischemia/hypoxia and the resulting reperfusion lead to myocardial ischemia-reperfusion (MI/R) damage, consisting of arrhythmia and the persistent ventricular systolic malfunction (Ai et al., 2015; Hausenloy and Yellon, 2013). Furthermore, MI/R is the major cause of a poor prognosis following an AMI (Hausenloy and Yellon, 2015). Therefore, it is necessary to develop a novel therapeutic option to counteract the deterioration of MI/R following an AMI.

At present, several mechanisms, such as oxidative stress, apoptosis and autophagy, have been proposed to contribute to MI/R (Morrison and Li, 2011). Oxidative burst represents an imbalance of intracellular redox, and the increased production of reactive oxygen species (ROS) overwhelms the cell endogenous antioxidant system. Additionally, excessive ROS damage the myocardia by inducing apoptosis and dysfunctions of the mitochondria (Meng et al., 2014). Apoptosis represents a series of essential events that result in the programmed death of a cell in multicellular organisms, which is initiated following a myocardial infarction, and potently induces the reperfusion process (Gu et al., 2014). Previous studies have indicated that apoptosis serves a vital role in both AMI and MI/R (Sun et al., 2013). In addition, autophagy is one of the critical biological processes through which cellular equilibrium is maintained (Levine and Klionsky, 2004).

During mild ischemia, the upregulation of autophagy is associated with the preservation of the mitochondrial membrane potential (ΔΨm), and at the same time, it also results in delay in the initiation of apoptosis and necrosis (Loos et al., 2011). Nevertheless, increased autophagy often results in decreased levels of vital proteins and organelles, thus leading to cellular dysfunction (Levine and Yuan, 2005). Matsui et al. have shown that activation of autophagy may damage the heart during the reperfusion process (Matsui et al., 2007). Therefore, a strategy to protect the heart against MI/R-induced injury by identifying novel treatments with antioxidant, anti-apoptotic and anti-autophagic effects is required. Recently, increasing attention has been paid to traditional medicines as promising therapeutic alternatives for alleviating MI/R-induced injury, due to their broad range of pharmacological effects.

Curcumin is not only a dietary polyphenolic component found in turmeric, but is also a spice that originates from the rhizomes of Curcuma longa Linn. Generally, turmeric is widely used in Asian cuisines, and has been applied as a medicine for centuries (Kumar et al., 2016). Evidence obtained from preclinical studies has shown that curcumin has several beneficial pharmacological effects, including anti-inflammatory, anti-oxidant, anti-tumor, lipid-lowering and anti-atherosclerotic effects (Qin et al., 2017; Shin et al., 2014; Susan and Douglas, 2017). Additionally, it has been shown that curcumin exhibits cardioprotective effect against various types of injuries, hence it can be used to treat cardiovascular diseases (Jiang et al., 2017; Srivastava and Mehta, 2009). In 2015, Huang et al. (2015) have demonstrated that curcumin inhibits autophagy and apoptosis in H/R-induced myocytes. Similarly, curcumin also attenuates H/R-induced cardiomyocyte injury by downregulating Notch signaling (Zhu et al., 2019). Furthermore, Wei et al. (2019) have suggested that curcumin exerts protective effect on cardiomyocytes via suppression of endoplasmic reticulum stress and the MAPK pathway.

Tetrahydrocurcumin (THC), the primary bioactive metabolite of curcumin, reportedly exhibits multiple biological effects superior to those of curcumin, including antioxidative, anti-inflammatory, antidiabetic and neuroprotective effects (Aggarwal et al., 2014; Osawa and Kato, 2006; Wei, 2017; Wu et al., 2013). It has been shown that oral administration with THC is more effective than curcumin in decreasing blood glucose levels, which also enhanced the superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase activities in the livers and kidneys of diabetic rats (Murugan and Pari, 2007). THC is also more potent than curcumin when used to decrease the deposition and cross-linking of collagen in type 2 diabetes rats (Pari and Murugan, 2008). In our previous study, THC has been shown to exert superior anti-inflammatory (Zhang et al., 2018), anti-tumor (Liu et al., 2017) and hepatic-protective (Luo et al., 2019) effects compared with curcumin. Nevertheless, the potential effects of THC on MI/R injury remain unknown.

The aim of the present study was to determine the potential of THC as a novel candidate for the treatment of MI/R. To our knowledge, it was the first endeavor to unravel the cardioprotective effect and underlying mechanism of THC in vivo and in vitro. This was the innovative study exploring the cardioprotective activity of hydrogenated metabolite of curcumin in a murine model of myocardial ischemia-reperfusion injury. The observations provided pioneering evidence that THC exerted cardiomyocyte protection, at least in part, via regulation of the PI3K/AKT/mTOR pathways.

The results gained novel insight into the cardioprotective effect of this primary hydrogenated metabolite of curcumin, which further corroborated the modern application of turmeric and contributed to its cardioprotective pharmacological validation. The promising cardioprotective activity of THC suggests that it holds potential to be developed into a promising therapeutic agent for the treatment of myocardial ischemia-reperfusion injury.

Section snippets

Materials and reagents

THC, LY294002 and rapamycin (all with a purity >98%) were obtained from Sigma-Aldrich; Merck KGaA. The chemical structure of THC is shown in Fig. 1A. Reagents for cell culture were obtained from Gibco; Thermo Fisher Scientific, Inc. All antibodies used in the present study were provided by Cell Signaling Technology, Inc., and chemicals were provided by Sigma-Aldrich; Merck KGaA unless otherwise specified.

Animals

Sprague-Dawley rats (180–200 g) were obtained from the Center for Laboratory Animal

THC posttreatment preserved cardiac function after I/R injury

Cardiac function was assessed by echocardiography in rats after MI/R injury (Fig. 1C). LVESD and LVESV were significantly increased (P < 0.01, Fig. 1F and G) and EF and FS were significantly decreased (P < 0.01, Fig. 1H and I) after I/R, indicating that cardiac dysfunction and structural alterations appeared. After THC treatment, the levels of EF and FS were elevated (P < 0.05) and LVEDD and LVESD levels were significantly reduced (P < 0.01). Then, we assessed the morphological changes of

Discussion

Apoptosis and autophagy have significant therapeutic implications for CHD. Several drugs involving RASS blocker, β receptor blocker, simvastatin, antimicrobial agents and metformin have been illustrated to exert regulatory function on apoptosis or autophagy (Dong et al., 2019). Furthermore, pharmacologic compounds that target the interplay of apoptosis and autophagy may possess more direct and applicable potential as therapeutic candidates.

Apoptosis is considered to be a key pathophysiological

Conclusion

In conclusion, the present study provides novel insights into the protective effect of THC on myocardial I/R damage. THC effectively protected cardiomyocytes against H/R-induced cellular injury, oxidative burst, and cardiomyocyte apoptosis and autophagy. These effects were at least partially mediated by PI3K/AKT/mTOR signaling. These results suggest that THC might be further developed as a potential therapeutic candidate for protection against myocardial I/R injury.

Funding

This work was supported by grants from Science and Technology Development Special Project of Guangdong Province (2017A050506044), Science and Technology Plan Project of Guangzhou (202102010305), Key Program for Subject Research of Guangzhou University of Chinese Medicine (XK2018016 & XK2019002), Characteristic Cultivation Program for Subject Research of Guangzhou University of Chinese Medicine (XKP2019007), National Science Foundation of China (No. 81673845), Guangdong Natural Science Foundation

Availability of data and materials

The datasets used or analyzed during the current study are available from the corresponding author on reasonable request.

Ethics approval and consent to participate

Not applicable.

Patient consent for publication

Not applicable.

CRediT authorship contribution statement

Xiaoying Chen: Writing – original draft, Formal analysis. Qingfeng Xie: performed the study. Ying Zhu: Formal analysis, Writing – original draft. Jiamin Xu: Writing – original draft. Guoshu Lin: Writing – original draft. Shujun Liu: Formal analysis. Ziren Su: Conceptualization, of and designed the study. Xiaoping Lai: Conceptualization, of and designed the study. Qian Li: performed the study. Jianhui Xie: performed the study, critically reviewed the manuscript.. Xiaobo Yang: performed the

Declaration of competing interest

The authors declare that they have no competing interests.

Acknowledgments

Not applicable.

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