Molecular mechanisms of doxorubicin-induced cardiotoxicity: novel roles of sirtuin 1-mediated signaling pathways

Abstract

Doxorubicin (DOX) is an anthracycline chemotherapy drug used in the treatment of various types of cancer. However, short-term and long-term cardiotoxicity limits the clinical application of DOX. Currently, dexrazoxane is the only approved treatment by the United States Food and Drug Administration to prevent DOX-induced cardiotoxicity. However, a recent study found that pre-treatment with dexrazoxane could not fully improve myocardial toxicity of DOX. Therefore, further targeted cardioprotective prophylaxis and treatment strategies are an urgent requirement for cancer patients receiving DOX treatment to reduce the occurrence of cardiotoxicity. Accumulating evidence manifested that Sirtuin 1 (SIRT1) could play a crucially protective role in heart diseases. Recently, numerous studies have concentrated on the role of SIRT1 in DOX-induced cardiotoxicity, which might be related to the activity and deacetylation of SIRT1 downstream targets. Therefore, the aim of this review was to summarize the recent advances related to the protective effects, mechanisms, and deficiencies in clinical application of SIRT1 in DOX-induced cardiotoxicity. Also, the pharmaceutical preparations that activate SIRT1 and affect DOX-induced cardiotoxicity have been listed in this review.

Introduction

Doxorubicin (DOX) is an anthracycline chemotherapeutic drug that is widely used to treat various neoplastic diseases, including lymphomas, breast cancer, prostatic cancer, osteosarcoma, lung cancer, and neuroblastoma [1]. DOX exerts an antitumor effect mainly by intercalation into DNA and inhibition of topoisomerase II. Regrettably, DOX also causes apoptosis or necrosis of healthy tissues, and hence, proving to be toxic to the brain, heart, liver, and kidney [2]. Clinical data have shown that the toxicities of this drug, especially cardiotoxicity, severely limit its application [3, 4]. The myocardial toxicity of DOX is closely associated with its dosage, which might ultimately result in progressive heart failure and irreversible cardiac dysfunction. The cumulative DOX doses of 400, 500, and 550 mg/m² were estimated to correlate with the incidence of heart failure equal to 5%, 16%, and 26%, respectively [5]. The mechanisms of DOX-induced cardiotoxicity have been studied extensively, including oxidative stress, inflammatory response, mitochondrial injury, endoplasmic reticulum (ER) stress, calcium (Ca²⁺) dyshomeostasis, apoptosis, fibrosis, and dysregulation of autophagy [6–11].

However, to date, there is no effective targeted therapeutic strategy, neither prophylactic nor curative, to protect against DOX-induced cardiotoxicity. Dexrazoxane is the only drug currently approved for reducing DOX-induced cardiotoxicity. Recently, Li et al. investigated the effects of continuous DOX infusion or dexrazoxane pre-treatment on DOX-induced cardiotoxicity. The results exhibited that changes in global longitudinal strain and left ventricular ejection fraction were scarcely observed. However, the increased levels of high-sensitivity troponin T were observed in > 59% of the patients who received continuous DOX infusion or dexrazoxane pre-treatment, suggesting that pre-treatment with dexrazoxane could not entirely improve subclinical DOX-induced cardiotoxicity [12]. Therefore, it is highly vital to investigate the prevention and treatment strategies as well as potential molecular mechanism for DOX-induced cardiotoxicity.

Sirtuin 1 (SIRT1), as a nicotinamide adenosine dinucleotide (NAD⁺)-dependent deacetylase, has been verified to play a critical role in DOX-induced cardiotoxicity. A previous study summarized some mechanisms of SIRT1 regarding DOX-induced cardiotoxicity, while the involved regulatory pathways are not comprehensive, only including the regulation of transforming growth factor-beta (TGF-β), tumor protein p53 (p53), and forkhead box O (FOXO) proteins [13]. In addition, we identified several shortcomings with respect to the application of SIRT1 in DOX-induced cardiotoxicity. Hence, this review summarizes the recent advances related to protective effects and mechanisms, as well as deficiencies in clinical application of SIRT1 in DOX-induced cardiotoxicity. First, we outline the mechanisms of DOX-induced cardiotoxicity and then explain the significant protective role of SIRT1 in the heart. Subsequently, we highlight and summarize SIRT1-mediated signaling pathways in DOX-induced cardiotoxicity. Finally, we indicate several SIRT1 agonists and their protective roles in DOX-induced cardiotoxicity, as well as deficiencies of SIRT1 in clinical application.

Mechanisms of doxorubicin-induced cardiotoxicity

Oxidative stress and inflammatory response

Oxidative stress refers to an imbalance between reactive oxygen species (ROS) production and antioxidant defense system, which is one of the main mechanisms of DOX-induced cardiotoxicity. Typically, mitochondria are the primary sites of ROS generation [14]. The enzymes that produce ROS in mitochondria can convert quinone moiety in ring C of DOX into semiquinone form through single-electron reduction. The semiquinone form of DOX is easy to react with oxygen molecules (O₂) to form superoxide anion (${\text{O}}_2^{·-}$), which is neutralized by superoxide dismutase (SOD) to become relatively stable and low-toxic hydrogen peroxide (H₂O₂). In the presence of iron (Fe²⁺), H₂O₂ may also generate highly reactive and toxic hydroxyl radical (OH∙). ROS reacts with DNA, proteins, and lipids, eventually leading to DNA damage and lipid peroxidation [15, 16] (Fig. 1). A previous study demonstrated that DOX could increase cardiac ROS generation and malondialdehyde (MDA, an end product of lipid hydroperoxide) level to result in oxidative stress. Nuclear factor erythroid 2-related factor 2 (NRF2) regulates numerous antioxidant proteins, including heme oxygenase-1 (HO-1), NAD(P)H quinone dehydrogenase 1 (NQO1), SOD, and catalase (CAT). Cardamonin increases the expression and reduces the degradation of NRF2 to improve DOX-induced oxidative stress through inhibiting ROS production and MDA level and upregulating the expression of antioxidant enzymes [17]. Reportedly, NAD phosphate (NADPH) oxidase 2 (NOX2) and NOX4 are involved in signal transduction during oxidative stress, which can be stimulated by DOX to increase ROS production and subsequent oxidative stress in cardiomyocytes [18]. Recently, Chen et al. found that metformin could inhibit the expression of mitogen-activated protein kinase (MAPK) and activate the expression of AMP-activated protein kinase (AMPK) to exert antioxidative and anti-apoptotic effects on cardiomyocytes during DOX treatment [19].

Fig. 1.

DOX-related redox cycling. The mitochondrial NADH-dependent enzymes can convert quinone moiety in ring C of DOX into semiquinone form through single-electron reduction. The semiquinone form of DOX reacts with O₂ to form ${\text{O}}_2^{·-}$, which is neutralized by SOD to H₂O₂. In the presence of Fe²⁺, H₂O₂ might also generate highly reactive and toxic OH∙, which reacts with DNA, proteins and lipids, leading to DNA damage and lipid peroxidation. DOX doxorubicin, NADH nicotinamide adenine dinucleotide hydrate, O₂ oxygen molecule, ${\text{O}}_2^{·-}$ superoxide anion, SOD superoxide dismutase, H₂O₂ hydrogen peroxide, Fe²⁺ iron, OH∙, hydroxyl radical, CAT catalase, GPx glutathione-dependent peroxidase, H₂O water

Inflammation is positively correlated with the oxidative stress, and thus, accelerates myocardial injury. Hwang et al. elucidated that oxidative stress could lead to inflammatory response through activating nuclear factor kappa B (NF-κB), a redox-sensitive transcription factor [20]. DOX has shown to upregulate the levels of several inflammatory factors, including interleukin-1β (IL-1β), IL-6, IL-17, and tumor necrosis factor-alpha (TNF-α) in the heart [21, 6]. In myeloid differentiation protein 1 (MD-1)-knockout mice, Toll-like receptor (TLR4)/MAPKs/NF-κB pathway was over-activated by DOX, accelerating myocardial inflammatory response and suggesting that MD-1 plays a pivotal role in DOX-induced cardiac inflammation [22]. NOD-like receptor family pyrin domain-containing protein 3 (NLRP3) inflammasome, as a critical regulator of the innate immune system, could be activated by DOX to result in myocardial inflammation [23]. Moreover, the transient receptor potential ankyrin 1 (TRPA1) channel is stimulated by DOX to cause cardiotoxicity through promoting oxidative stress and inflammation [6]. Based on the above-mentioned findings, oxidative stress and inflammation are regarded as significant risk factors for DOX-induced cardiotoxicity, while the specific correlation and mechanisms need to be further investigated.

Mitochondrial injury

As sites of cellular respiration and energy production, mitochondria play a central role in regulating pathophysiological activities, such as cell growth, proliferation, differentiation, damage, repair, and death [24]. Recent studies demonstrated that mitochondria are key targets of DOX-induced cardiotoxicity. The heart is more remarkably sensitive to toxicity of DOX than other organs owing to cardiolipin, which is localized in the mitochondrial inner membrane, has a high affinity to DOX and leads to the accumulation of DOX-cardiolipin complex in the heart [25]. When the accumulated dose of DOX is > 50–100 μM in the mitochondria, the electron transport chain is disrupted by inhibiting complexes I, and simultaneously, ROS production is increased [26]. Some studies have shown that the primary mechanism of DOX-induced mitochondrial dysfunction might be related to mitochondrial permeability transition (MPT). MPT is the phenomenon whereby the inner membrane suddenly allows free passage of solutes up to 1.5 kDa in size. The MPT pore (mPTP) could be opened by DOX-induced mitochondrial oxidative damage and Ca²⁺ overload, which might lead to the collapse of the inner membrane potential, halting of mitochondrial adenosine triphosphate (ATP) synthesis, and uncoupling of respiratory chain, promoting mitochondrial osmotic swelling and rupture, and cell death [27, 28]. While the biophysical properties of the mPTP are well-established, the exact molecular nature is yet an enigma. Cyclophilin D (CypD) is a genetically verified and undisputed regulator of mPTP function. Dhingra et al. reported that DOX could impair NF-κB signaling accompanied by mitochondrial injury and CypD-mediated mPTP opening through the activation of mitochondrial death protein Bcl-2/19 kDa interacting protein 3. However, the precise mechanism by which CypD controls mPTP opening has not been explored [29]. In addition, some studies suggested that adenine nucleotide translocator is not a requisite component of the mPTP but regulates mPTP activity in heart mitochondria from DOX-treated rats [27]. Recently, Catanzaro et al. pointed out that DOX could also enhance mitochondrial fragmentation and accelerate mitochondrial degradation by the lysosome, eventually leading to cardiotoxicity [30]. As an inhibitor of mitophagy, liensinine could decrease the phosphorylation of dynamin-related protein 1 (Drp1) at the Ser⁶¹⁶ site to inhibit mitochondrial fragmentation, mitophagy, and cytochrome c (Cyt C) leakage, thereby ameliorating mitochondrial dysfunction and cardiac injury following DOX treatment [7]. Although the effects of DOX on mitochondrial injury in the heart have been widely studied, the specific mechanisms and pathways by which DOX induces mitochondrial mPTP opening and mitochondrial fragmentation need to be further explored.

Endoplasmic/sarcoplasmic reticulum stress and calcium dyshomeostasis in cardiac cells

The changes in the function of ER lead to the accumulation of unfolded or misfolded proteins, resulting in a cellular condition called ER stress [31]. ER stress is mediated by three ER-resident transmembrane sensors, including activating transcription factor 6 (ATF6), protein RNA-like ER kinase(PERK), and inositol-requiring enzyme 1 (IRE1) [32]. When ER stress is sustained or aggravated, the c-JUN NH2-terminal kinase (JNK), caspase-12, and C/EBP homologous protein (CHOP) signaling pathways-mediated cell apoptosis occurs [33]. DOX could increase the expression of ER stress-related proteins, such as p-PERK, ATF6, ATF4, 78-kDa glucose-regulated protein precursor (GRP78), and CHOP to result in apoptosis. However, CACNA1H-specific inhibitor ABT-639 could reverse the DOX-mediated increased levels of ER stress-related proteins and pro-apoptotic proteins in H9c2 cells, suggesting that CACNA1H is involved in DOX-induced cardiotoxicity through effecting ER stress [34]. Another study found that DOX could increase transcriptional intermediary factor 1 (TIF1) expression but inhibit GRP78 expression to simulate the expression of ATF6 and IRE1, resulting in ER stress [35]. Recently, Wang et al. found that the DOX-activated TRPA1 channel in cardiomyocytes also could cause cardiotoxicity by promoting ER stress [6].

Sarcoplasmic reticulum (SR) has been considered as a specialized form of the ER in cardiac cells, which is primarily responsible for the regulation of Ca²⁺ fluxes and consequently, the control of excitation–contraction coupling [36]. Ca²⁺ homeostasis disequilibrium is involved in the development of DOX-induced cardiotoxicity because of its pivotal role in cardiac electrical activity and excitation–contraction coupling. Llach et al. showed that low-dose of DOX could disrupt the cardiac function with a reduction in ejection fraction and slight dilation at 15 weeks after the end of the treatment, which could be associated with a decline in the intracellular free Ca²⁺ concentration [8]. Early DOX treatment could induce SR Ca²⁺ leak by the interaction between DOX and ryanodine receptor and the sarco/ER Ca²⁺-ATPase; subsequently, Ca²⁺/calmodulin-dependent protein kinase II (CaMKII) was activated to result in DOX-induced cardiotoxicity. CaMKII is associated with caspase-dependent apoptosis pathways. SR Ca²⁺ homeostasis and ER stress are closely interlinked. GRP78 is not only a central mediator of the unfolded protein response during ER stress but also a regulator of Ca²⁺ homeostasis in the ER. Low levels of GRP78 overexpression could protect cardiomyocytes from DOX-induced cell apoptosis by reducing CaMKII activation and p53 accumulation. Mechanism study further demonstrated above CaMKII activation and p53 accumulation was occurred in a Ca²⁺-dependent manner, thereby explaining the beneficial action of GRP78 was to normalize Ca²⁺ handling [37]. Taken together, ER stress and Ca²⁺ dyshomeostasis are crucial in DOX-induced cardiotoxicity. However, the functional correlation between the ER and SR in cardiomyocytes is yet confusing; therefore, the interaction between ER stress and Ca²⁺ dyshomeostasis in DOX-induced cardiotoxicity need further investigation.

Apoptosis

Apoptosis is a form of programmed cell death in multicellular organisms. It plays a substantial role in the evolution of organisms, stability of internal environment, and normal cell renewal [38]. A study reported that DOX-induced cardiotoxicity is related to nuclear translocation of p53 and p53-dependent apoptosis. Extracellular signal-regulated kinases (ERKs) could be activated by DOX to phosphorylate p53 at Ser15. The phosphorylation of p53 could lead to cardiomyocyte apoptosis via downregulation of anti-apoptotic B-cell lymphoma 2 (Bcl-2), upregulation of pro-apoptotic Bcl-2-associated X protein (Bax), and activation of caspase-3, caspase-9, and poly-ADP-ribose polymerase [9, 39]. Moreover, DOX-induced oxidative stress opens the voltage-dependent anion channel (VDAC) to result in MPT and subsequently release Cyt C from the mitochondria [40]. In the presence of ATP, Cyt C modulates the allosteric activation and hepta-oligomerization of the adaptor molecule apoptosis-protease activating factor 1 to produce apoptosome, which recruits the dimers of caspase-9 to facilitate the activation of caspase-3, eventually initiating the apoptotic degradation phase [41]. Mitofusin 2 (Mfn2) is a mitochondrial GTPase fusion protein that plays a crucial role in mitochondrial fusion and fission. DOX treatment decreased the expression of Mfn2 accompanied by an increase in mitochondrial fragmentation and ROS generation, further causing cardiomyocyte apoptosis [42]. However, detailed mechanisms underlying the Mfn2 inhibition by DOX and ROS production regulation by Mfn2 are not yet clarified. Furthermore, Leboucher et al. found that Mfn2 is a substrate for JNK, which phosphorylates Mfn2 to result in its ubiquitin-dependent proteasomal degradation, leading to mitochondrial fragmentation and apoptosis in DOX-treated sarcoma U2OS cells [43]. Hitherto, the exact mechanism by which inhibition of Mfn2 leads to cardiomyocyte apoptosis in DOX-induced cardiotoxicity is unknown and needs to be further investigated. Intriguingly, death receptor-mediated apoptosis in cardiomyocytes could be a major mechanism of DOX-induced cardiotoxicity [44]. Recently, a study explored the role of nuclear factor of activated T cells (NFAT)/Fas/Fas ligand (FasL) axis in cardiomyocyte apoptosis during DOX treatment in rats and found that DOX could increase the levels of Fas, FasL and NFAT to induce caspase-8-mediated intrinsic cell death, resulting in elevated expression of Bax or caspase-3. Further mechanistic investigation unveiled that the activation of NFAT/Fas/FasL axis was mediated by upregulated expression of calcineurin and p38 MAPK and downregulated expression of mammalian target of rapamycin (mTOR) [45]. The activation of E2F transcription factor 1 (E2F1)/AMPKα2 signaling pathway was also found to be involved in DOX-induced cardiomyocyte apoptosis [46]. Although the effects and mechanisms of DOX on cardiomyocyte apoptosis have been studied extensively, the specific signaling pathways involved in the DOX-induced apoptosis are deemed complicated.

Autophagy

Autophagy is a catabolic process involving the elimination of cellular misfolded proteins and damaged or old organelles to maintain cellular homeostasis by the regulation of autophagy-related (Atg) genes and lysosomal proteolysis [47, 48]. Some studies have demonstrated the role of autophagy in DOX-induced cardiotoxicity (Table 1). On the one hand, a number of studies pointed out that DOX could stimulate cardiac autophagy and conduce to the pathogenic mechanism of DOX-induced cardiotoxicity [35, 49–53]. Xu et al. demonstrated that DOX could stimulate autophagy through increased ratio of microtubule-associated proteins 1A/1B light chain 3-II (LC3-II)/LC3-I and upregulated expression of p62, Beclin-1, and Atg5 by stimulating the expression of JNK1 and p70S6 kinase (p70S6K) [35]. Moreover, mTOR has been proved to be one of the major negative regulators of autophagy. The inhibition of mTOR by DOX promotes autophagy by increased expression of LC3-II, Atg5, Atg6, Atg8 and Atg12 via AMPK activation and p38 MAPK inhibition [49]. Rutin is a polyphenolic flavonoid that suppresses DOX-induced autophagy by increased phosphorylation of protein kinase B (Akt) [50]. However, the effect of DOX on autophagy regulation in cardiomyocytes is controversial. Gu et al. found that DOX could cause cardiotoxicity via inhibition of autophagy rather than stimulation of autophagic flux [10, 46, 54, 55–57]. DOX inhibits autophagy via a sequence of pathways, such as AMPK/Unc-51 like autophagy activating kinase 1 (ULK1) [56] and E2F1/mTOR complex 1 (mTORC1) [46]. Recently, Pan et al. reported that autophagy in human AC16 cells was upregulated by the protective mechanism of cytotoxicity, and autophagy-related proteins (for example, the ratio of LC3-II/LC3-I and Beclin-1) showed a short-term increase in the early stage of DOX intervention. As the concentration and time of DOX intervention increased, the autophagy flux would be decompensated and reduced to lose the ability of self-cleaning and gradually move towards apoptosis [58]. The above-mentioned controversial findings about the effects of DOX on cardiac autophagy might be associated with the specific species, dosage of DOX, experimental conditions, and observation of different and intertwined pathways.

Table 1.

The effect of autophagy in DOX-induced cardiotoxicity

Animal or cellular model	DOX dose	Treatment period	Autophagy markers	Effect on autophagy	Mechanism	Reference
C57BL/6 mice	24 mg/kg (cumulative dose)	12 days	LC3; p62; Beclin-1; Atg5	Induction	DOX stimulates autophagy through increased ratio of LC3-II/LC3-I, and increased expression of p62, Beclin-1, and Atg5 by upregulated expression of JNK1 and p70S6K	[35]
C57BL/6 mice and NRCs	21 mg/kg (cumulative dose) and 1 μM	2 weeks and 24 h	LC3; p62; Atg5	Induction	DOX stimulates autophagy through increased expression of LC3-II, p62, and Atg5 by reduced phosphorylation of Akt	[50]
C57BL/6 mice and H9c2 cells	32 mg/kg (cumulative dose) and 10 μM	4 weeks and 24 h	LC3; Atg5; Atg6; Atg8; Atg12	Induction	DOX stimulates autophagy through increased expression of LC3-II, Atg5, Atg6, Atg8, and Atg12 by AMPK activation and p38 MAPK inhibition and subsequent mTOR inhibition	[49]
H9c2 cells	5 μg/ml	24 h	LC3; p62; Beclin-1	Induction	DOX stimulates autophagy through increased expression of LC3-II and Beclin-1 and decreased expression of p62	[51]
NRCs	1 μM	18 h	LC3; p62; Atg5; Atg12-Atg5 complex	Induction	DOX stimulates autophagy through increased expression of Atg5 and Atg12-Atg5 complex and decreased expression of p62 by upregulation of p70S6K expression	[52]
NRCs	1 μM	18 h	LC3; p62; Beclin-1; Atg5; Atg7; Atg12	Induction	DOX stimulates autophagy through increased expression of LC3-II, Beclin-1, Atg5, Atg7, and Atg12 and decreased expression of p62 by depleting GATA4 protein levels	[53]
C57BL/6 mice and H9c2 cells	20 mg/kg (cumulative dose) and 1 μM	4 weeks and 24 h	LC3; p62; Beclin-1; LAMP1	Inhibition	DOX inhibits autophagy through suppression of Beclin-1/LAMP1 pathway	[10]
C57BL/6 mice	15 mg/kg (single dose)	3 days	Beclin-1	Inhibition	DOX inhibits autophagy through decreased Beclin-1 by inhibition of AMPK and activation of mTOR	[54]
C57BL/6 mice and H9c2 cells	20 mg/kg (cumulative dose) and 1 μM	4 weeks and 24 h	LC3	Inhibition	DOX inhibits autophagy through decreased ratio of LC3-II/LC3-I by activating E2F1/mTORC1 pathway	[46]
GFP-LC3 transgenic mice and H9c2 cells	20 mg/kg (cumulative dose) and 3 μM	15 days and 24 h	LC3; p62	Inhibition	DOX inhibits autophagy as shown in the accumulation of LC3-I and p62	[55]
GFP-LC3 transgenic mice and neonatal mice cardiomyocytes	20 mg/kg (cumulative dose) and 0.1 μM	5 days and 6 h	LC3; p62	Inhibition	DOX inhibits autophagy through decreased expression of AMPK and ULK1	[56]

DOX doxorubicin, H9c2 cells rat cardiomyocyte cell line, NRCs neonatal rat ventricular cardiomyocytes, LC3 microtubule-associated proteins 1A/1B light chain 3, Atg autophagy-related, LAMP1 lysosomal-associated membrane proteins 1, JNK1 c-JUN NH2-terminal kinase 1, p70S6K p70S6 kinase, Akt protein kinase B, AMPK AMP-activated protein kinase, MAPK mitogen-activated protein kinase, mTORC1 mammalian target of rapamycin complex 1, E2F1 E2F transcription factor 1, ULK1 Unc-51 like autophagy activating kinase 1

Fibrosis

Cardiac fibrosis is a vital mechanism of DOX-induced cardiotoxicity [59, 60]. It has been proved that matrix metalloproteinases (MMPs) regulate cardiac fibrosis via degradation of extracellular matrix (ECM). Narikawa et al. demonstrated that DOX could increase the expression of MMP1, TGF-β, and collagen in human cardiac fibroblasts and speculated that DOX-induced upregulation of MMP1 could impair the balance of ECM via activating phosphoinositide 3-kinase (PI3K)/Akt signaling pathway to result in cardiac dysfunction and fibrosis [61]. Moreover, MMP2 activation is involved in DOX-induced cardiotoxicity, causing intracellular matrix and ECM remodeling through myofilament lysis by proteolyzing cardiac titin [11]. Another study found that DOX treatment could induce cardiac fibrosis via the neurokinin 1 receptor (NK-1R) and collagen production by cardiac fibroblasts [62]. Currently, only limited information is available about the effect of cardiac fibrosis on DOX-induced cardiotoxicity, and the specific mechanisms and pathways underlying DOX-induced cardiac fibrosis need further exploration. A schematic diagram of mechanisms involved in DOX-induced cardiotoxicity is illustrated in Fig. 2.

Fig. 2.

Schematic representation of mechanisms involved in DOX-induced cardiotoxicity. DOX increases ROS production that results in cardiac oxidative stress. Moreover, DOX accelerates myocardial inflammation by MD-1/TLR4/MAPKs/NF-κB and NLRP3 pathways. DOX increases TIF1 expression and inhibits GRP78 expression to simulate the expression of ATF6 and IRE1, resulting in ER stress. DOX-induced oxidative stress causes the release of Cyt C from mitochondria via the opening of VDAC to activate caspase-3, leading to the cardiomyocyte apoptosis. Furthermore, DOX inhibits Mfn2 expression to cause mitochondrial fragmentation, resulting in cardiomyocyte apoptosis, and induces cardiomyocyte apoptosis by ERKs/p53 and E2F1/AMPKα2 pathways. DOX stimulates cardiac autophagy via upregulating the expression of JNK1 and p70S6K, as well as activating AMPK/mTOR pathway and inhibiting Akt expression. Conversely, DOX inhibits cardiac autophagy through inhibiting AMPK/ULK1 pathway and activating E2F1/mTORC1 pathway. DOX-induced upregulation of MMP1 impairs the balance of ECM via activating PI3K/Akt pathway, resulting in cardiac fibrosis. DOX also induces cardiac fibrosis through NK-1R. DOX doxorubicin, ROS reactive oxygen species, MD-1 myeloid differentiation protein 1, TLR4 toll-like receptor, MAPK mitogen-activated protein kinase, NF-κB nuclear factor kappa B, NLRP3 NOD-like receptor family pyrin domain-containing protein 3, TIF1 transcriptional intermediary factor 1, GRP78 78-kDa glucose-regulated protein precursor, ATF6 activating transcription factor 6, IRE1 inositol-requiring enzyme 1, ER endoplasmic reticulum, Cyt C cytochrome c, VDAC voltage-dependent anion channel, Mfn2 mitofusin 2, ERKs extracellular signal-regulated kinases, p53 tumor protein p53, E2F1 E2F transcription factor 1, AMPK AMP-activated protein kinase, JNK1 c-JUN NH2-terminal kinase 1, p70S6K p70S6 kinase, mTORC1 mammalian target of rapamycin complex 1, Akt protein kinase B, ULK1 Unc-51 like autophagy activating kinase 1, MMP1 matrix metalloproteinase 1, ECM extracellular matrix, PI3K phosphoinositide 3-kinase, NK-1R neurokinin 1 receptor

The role of SIRT1 in the heart

In rodents, the terminal differentiation of cardiomyocytes occurs over the first 2 weeks of postnatal life, characterized by the exit from the cell cycle, binucleation and loss of ability to proliferate. The expression of SIRT1 was low in embryonic hearts and notably increased in rats’ hearts at postnatal day 7. In contrast to the expression trend of SIRT1, that of cardiac-specific micorRNA-133a (miR-133a) was remarkable in embryonic hearts while that decreased markedly in the hearts of rats at postnatal day 7. Notably, SIRT1 suppression could reduce binucleated cardiomyocytes after birth, suggesting that endogenous SIRT1 and its upstream miR-133a may play substantial roles in the terminal differentiation and maturation of cardiomyocytes during cardiac development [63]. Furthermore, SIRT1-deficient mice indicated cardiac developmental defects and rarely postnatal survival [64]. Therefore, SIRT1 is a favorable factor for cardiac development and may be a potential therapeutic strategy for heart injury or diseases. Accumulating evidence suggested a protective role of SIRT1 in myocardial ischemia/reperfusion (I/R) injury, diabetic cardiomyopathy (DCM), and cardiac aging (Fig. 3).

Fig. 3.

The role of SIRT1 in the heart. Diagram showing proposed downstream targets of SIRT1 and their related effects implicated in myocardial protection. SIRT1 can activate or inhibit some signaling pathways to exert a protective effect on the heart. SIRT1 Sirtuin 1, FOXO forkhead box O, PGC-1α peroxisome proliferator-activated receptor gamma coactivator-1 alpha, p53 tumor protein p53, LKB1 liver kinase B1, AMPK AMP-activated protein kinase, NF-κB nuclear factor kappa B, NRF2 nuclear factor erythroid 2-related factor 2, NLRP3 NOD-like receptor family pyrin domain-containing protein 3, eEF2K eukaryotic elongation factor-2 kinase, eIF2α eukaryotic initiation factor 2 alpha, TGF-β transforming growth factor-beta, ANP atrial natriuretic peptide, BNP brain natriuretic peptide, Bcl-2 B-cell lymphoma 2, Bax Bcl-2-associated X protein, HO-1 heme oxygenase-1, NQO1 NAD(P)H quinone dehydrogenase 1, SOD superoxide dismutase, CAT catalase, ROS reactive oxygen species, LC3-II microtubule-associated proteins 1A/1B light chain 3-II, TNF-α tumor necrosis factor-alpha, IL interleukin, ER endoplasmic reticulum

The role of SIRT1 in myocardial ischemia/reperfusion injury

I/R injury is a major cause of myocardial injury that results in cardiac remodeling and heart failure. Accumulating evidence suggested that changes in SIRT1 levels affect myocardial injury and cardiac function. Cardiomyocyte-specific knockout of Sirt1 gene sensitized myocardium to I/R injury via metabolic shift, oxidative stress, apoptosis, and impaired autophagic influx. On the contrary, SIRT1 overexpression could improve cardiac function and decrease the myocardial infarction size caused by I/R injury via liver kinase B1 (LKB1) deacetylation and subsequent AMPK activation [65]. Guan et al. demonstrated that oxidative stress and Ca²⁺ overload were major mechanisms in the processes of I/R injury in cardiomyocytes, which could be improved by CD38 deficiency via activation of SIRT1/FOXOs pathway [66]. Moreover, the suppression of apoptosis is one of the major mechanisms of SIRT1 to protect the heart from injury. A study reported that SIRT1 activation by quercetin could stimulate peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1α) to upregulate Bcl-2 expression and downregulate Bax expression, thereby playing a pivotal anti-apoptosis role in improving I/R-induced myocardial damage [67]. DJ-1, a member of the eponymous DJ-1 superfamily, can suppress oxidative stress, regulate transcription, and promote cell survival. The increase in DJ-1 expression by resveratrol could stimulate SIRT1 activity via directly binding to SIRT1, which inhibited p53 acetylation, thereby attenuating hypoxia/reoxygenation-induced cardiomyocyte apoptosis [68]. Furthermore, the role of SIRT1-mediated anti-ER stress is an essential mechanism in the treatment of myocardial I/R injury. Wang et al. demonstrated that crocin could stimulate SIRT1/NRF2 signaling pathway to increase the expression of anti-apoptotic Bcl-2 and decrease the expression of ER stress makers (GRP78 and CHOP) and pro-apoptotic Bax and caspase-3, thereby improving the cardiac damage caused by I/R injury [69]. In addition, SIRT1 could alleviate ER stressor tunicamycin-induced ER stress in cardiomyocytes via regulating eukaryotic initiation factor 2 alpha (eIF2α) [70] and eukaryotic elongation factor-2 kinase (eEF2K)/eEF2 pathways [71]. However, whether SIRT1/eIF2α or SIRT1/eEF2K/eEF2 pathway plays an anti-ER stress role in the treatment of myocardial I/R injury needs to be further explored.

The role of SIRT1 in diabetic cardiomyopathy

DCM is one of the major complications in people with diabetes, characterized by cardiac fibrosis, myocardial hypertrophy, and ventricular systolic and/or diastolic dysfunction. Numerous studies have demonstrated that increased expression of SIRT1 is a crucial therapeutic target for the prevention and treatment of DCM. SIRT1 activation could exercise antioxidant function by reducing ROS production and MDA level, as well as activating SOD. Concurrently, it effectuated the anti-fibrotic effect by inhibiting the expression of fibrotic markers (α-smooth muscle actin, type-I collagen, and type-III collagen) via suppressing TGF-β1/mothers against decapentaplegic homolog 3 (SMAD3) pathway in DCM [72]. NF-κB is translocated into the nucleus as a response to oxidative stress, where it induces the transcription of several proinflammatory cytokines, serving as a negatively regulated downstream target of SIRT1. Bagul et al. found that activation of SIRT1 by resveratrol could deacetylate p65 subunit of NF-κB at Lys310 and histone 3 at Lys9 to reduce the expression of NOX1, NOX4, TNF-α, and IL-6, thereby inhibiting DCM-mediated cardiac oxidative stress and inflammation [73]. In addition, SIRT1 also exerts a cardioprotective role through attenuating myocardial hypertrophy in DCM. Waldman et al. reported that caloric restriction could protect the heart from DCM by inhibiting the expression of TNF-α, TGF-β, atrial natriuretic peptide (ANP), and brain natriuretic peptide (BNP) in diabetic mice, and the molecular mechanism of protective effects might be associated with the increase in cardiac SIRT1 and PGC-1α levels [74]. Furthermore, emerging evidence indicated that mitochondrial dysfunction might be one of the vital factors underlying the pathology in DCM. A study on cardiac-specific Sirt1 knockout mice demonstrated that SIRT1 could promote the expression of mitochondrion-related genes, such as nuclear respiratory factor 1 (Nrf-1), Nrf-2, mitochondrial transcription factor A (Tfam), and estrogen-related receptor-α to improve DCM via PGC-1α deacetylation [75]. Additionally, SIRT1/PGC-1α pathway is also involved in the inhibition of mitochondrial fission and apoptosis in diabetic hearts [76]. As described above, SIRT1 plays a pivotal cardioprotective role in DCM through inhibiting cardiac oxidative stress, fibrosis, inflammation, apoptosis, and mitochondrial fission, as well as improving myocardial hypertrophy and mitochondrial dysfunction by targeting diverse cellular factors and signaling pathways.

The role of SIRT1 in cardiac aging

Cardiac aging is characterized by increased apoptosis and necrosis, myocyte nuclei proliferation, cardiac hypertrophy, interstitial fibrosis, and cardiac dysfunction. A previous study demonstrated that SIRT1 exerts a protective role in cardiovascular aging [77]. Ralph et al. reported that 2.5–7.5-fold cardiac-specific overexpression of SIRT1 could inhibit the cellular senescence through regulating the INK4/ARF family proteins and p53 protein. In addition, 7.5-fold cardiac-specific overexpression of SIRT1 could upregulate the levels of some molecule markers such as heat shock protein 40 (HSP40), HSP70, HSP90, telomere reverse transcriptase, telomere repeat-binding factor 2, and Klotho, and subsequently improve age-dependent cardiac hypertrophy, fibrosis, apoptosis, and cardiac dysfunction. Conversely, the excessive elevation of SIRT1 might not always be advantageous. Transgenic mice with 12.5-fold overexpression of SIRT1 showed significantly increased cardiac β-myosin heavy chain, atrial natriuretic factor, α-skeletal actin, and TUNEL-positive cell, which eventually leaded to cardiac dysfunction [78]. In addition, exercise training could improve aging-induced inflammation, which might be associated with the upregulation of SIRT1 and PGC-1α in the hearts of aging rats [79]. Another study found that exercise training could also attenuate aging-induced cardiomyocyte apoptosis via stimulating SIRT1 expression [80]. SIRT1 also plays a cardioprotective role by regulating autophagy and mitochondrial integrity in aging hearts. Ren et al. demonstrated that Akt2 ablation could improve cardiac aging through restoring FOXO1-related mitochondrial integrity, which might be associated with SIRT1-mediated autophagy regulation [81]. Another study focused on the role of SIRT1 in aging human hearts and found that the expression of SIRT1 could be associated with gender. Lower expression of antioxidative protein SOD2 and higher expression of macrophages and proinflammatory cytokines were more obviously observed in old than young female hearts, which could be related to the diverse expression of cardiac SIRT1 and SIRT3 [82]. Although several studies have confirmed the anti-aging role of SIRT1 in myocardium, the specific mechanisms need to be further assessed.

Pathway regulation of SIRT1 in doxorubicin-induce cardiotoxicity

SIRT1 is involved in the regulation of oxidative stress [83–85], mitochondrial function [83], apoptosis [85, 86], inflammatory response [86], ER stress [87], fibrosis [88], and other processes by mediating various signaling pathways to play a significant role in DOX-induced cardiotoxicity. Thus, we summarize and introduce the signaling pathways regulated by SIRT1 in DOX-induced cardiotoxicity, including SIRT1/PGC-1α, SIRT1/AMPK, SIRT1/p53, SIRT1/p66Shc, SIRT1/NF-κB, SIRT1/p38 MAPK, SIRT1/FOXOs, SIRT1/TGF-β, and SIRT1/NLRP3 (Table 2; Figs. 3, 4).

Table 2.

Downstream targets of SIRT1 that are regulated during DOX-induced cardiotoxicity

Substrate	Animal or cell model	DOX dose	Treatment period	Functions of SIRT1	Mechanism	Reference
PGC-1α	C57BL/6 mice and H9c2 cells	20 mg/kg (cumulative dose) and 1 μM	6 days and 24 h	Antioxidation and regulate mitochondrial function	Deacetylate PGC-1α	[97]
	Human cardiomyocyte AC16 cells	125 nM	24 h	Regulate mitochondrial function and biogenesis	Deacetylate PGC-1α	[95]
AMPK	129S1/SvlmJ mice and H9c2 cells	20 mg/kg (cumulative dose) and 5 μg/ml	4 weeks and 22 h	Anti-inflammation, antioxidation and anti-apoptosis	Deacetylate LKB1 and activate AMPK	[105]
p53	Balb/c mice	24 mg/kg (cumulative dose)	3 weeks	Anti-apoptosis	Deacetylate p53	[110]
	H9c2 cells	1 μM	24 h	Anti-apoptosis	Deacetylate p53	[111]
p66Shc	Sprague–Dawley rats	16 mg/kg or 32 mg/kg (cumulative dose)	4 or 8 weeks	Anti-apoptosis	Suppress p66Shc expression	[116]
	Sprague–Dawley rats	20 mg/kg (cumulative dose)	9 days	Regulate mitochondrial function, antioxidation and anti-apoptosis	Suppress p66Shc expression	[117]
NF-κB	C57BL/6 mice	15 mg/kg (single dose) or 15 mg/kg (cumulative dose)	8 days or 6 weeks	Anti-inflammation	Suppress nuclear translocation of  NF-κB	[86]
p38 MAPK	C57BL/6 mice	20 mg/kg (single dose)	5 days	Antioxidation and anti-apoptosis	Suppress p38 MAPK expression	[129]
FOXOs	H9c2 cells	5 μM	24 h	Anti-apoptosis	Suppress FOXO1 expression	[136]
TGF-β	Fischer 344 rats	15 mg/kg (cumulative dose)	2 weeks	Anti-fibrosis	Suppress TGF-β/SMAD3 pathway	[88]
NLRP3	Sprague–Dawley rats and H9c2 cells	15 mg/kg (cumulative dose) and 5 μM	6 weeks and 24 h	Anti-inflammation	Suppress the NLRP3 inflammasome	[23]
	Kunming mice and H9c2 cells	15 mg/kg (single dose) and 5 μM	7 days and 24 h	Anti-inflammation	Suppress the NLRP3 inflammasome	[151]

SIRT1 Sirtuin 1, DOX doxorubicin, PGC-1α peroxisome proliferator-activated receptor gamma coactivator-1 alpha, LKB1 liver kinase B1, AMPK AMP-activated protein kinase, p53 tumor protein p53, NF-κB nuclear factor kappa B, MAPK mitogen-activated protein kinase, FOXO forkhead box O, TGF-β transforming growth factor-beta, NLRP3 NOD-like receptor family pyrin domain-containing protein 3, H9c2 cells rat cardiomyocyte cell line, SMAD3 mothers against decapentaplegic homolog 3

Fig. 4.

Mechanisms of SIRT1-mediated signaling pathways involved in DOX-induced cardiotoxicity. Diagram showing proposed downstream targets of SIRT1 and their related effects implicated in DOX-induced cardiotoxicity. SIRT1 Sirtuin 1, DOX doxorubicin, PGC-1α peroxisome proliferator-activated receptor gamma coactivator-1 alpha, NRF-1 nuclear respiratory factor 1, TFAM mitochondrial transcription factor A, COXIV cytochrome c oxidase IV, ∆Ψm mitochondrial membrane potential, ROS reactive oxygen species, VDAC voltage-dependent anion channel, Cyt C cytochrome c, LKB1 liver kinase B1, AMPK AMP-activated protein kinase, NF-κB nuclear factor kappa B, TNF-α tumor necrosis factor-alpha, IL interleukin, NRF2 nuclear factor erythroid 2-related factor 2, NQO1 NAD(P)H quinone dehydrogenase 1, CAT catalase, HO-1 heme oxygenase-1, p53 tumor protein p53, Bcl-2 B-cell lymphoma 2, Bax Bcl-2-associated X protein, MAPK mitogen-activated protein kinase, FOXO1 forkhead box O 1, TGF-β transforming growth factor-beta, SMAD3 mothers against decapentaplegic homolog 3, NLRP3 NOD-like receptor family pyrin domain-containing protein 3

SIRT1/PGC-1α pathway

PGC-1α is a transcriptional coactivator that regulates mitochondrial biogenesis and function, antioxidant defense systems, and cell metabolism by interacting with specific co-activated genes [89, 90]. Previous studies have reported that SIRT1 is a significant upstream regulator of PGC-1α [91, 92]. When the energy storage of the cell is reduced, the increasing levels of NAD⁺ can activate SIRT1 expression, and then SIRT1 is combined with PGC-1α at 200–400 amino acids to deacetylate PGC-1α, thereby facilitating mitochondrial ATP production by mitochondrial substrate oxidation [93]. Fang et al. pointed out that SIRT1 could strengthen cellular antioxidant capacity and alleviate mitochondrial dysfunction to improve DCM by PGC-1α deacetylation [94]. Moreover, the activation of SIRT1/PGC-1α pathway could also protect the heart from I/R injury via inhibition of apoptosis [67]. A study concentrated on the DOX-treated human cardiomyocyte AC16 cells and demonstrated that DOX could increase PGC-1α acetylation and suppress the downstream genes related to mitochondrial function and biogenesis, such as SOD2, NRF-1, Cyt C oxidase IV and TFAM to result in oxidative stress, loss of mitochondrial membrane potential (∆Ψm) and disruption of mitochondrial energy, which could be reversed through SIRT1 activation by erythropoietin pre-treatment [95]. Moreover, Govender et al. found that melatonin could improve mitochondrial morphology, increase mitochondrial fusion, repair ATP generation and decrease myocardial apoptosis to ameliorate DOX-induced cardiotoxicity. The study on the underlying mechanism further demonstrated that the protective role of melatonin was associated with upregulation of cardiac SIRT1 and PGC-1α [96]. Recently, another study indicated that pterostilbene treatment could also upregulate the expression and deacetylation of PGC-1α via AMPK and SIRT1 cascades to improve DOX-induced mitochondrial injury and oxidative stress in cardiomyocytes [97]. Therefore, the efficient regulation of SIRT1/PGC-1α pathway plays a crucial role in the prevention and treatment of DOX-induced cardiotoxicity by maintaining mitochondrial homeostasis, as well as inhibiting oxidative stress and apoptosis.

SIRT1/AMPK pathway

AMPK is a conserved energy sensor that regulates cellular metabolism. The activation of AMPK increases the ATP-producing catabolism, including glucose metabolism and fatty acid oxidation, and decreases ATP-consuming anabolism, such as protein and lipid synthesis, aiming to maintain cellular energy storage [98]. The activation of AMPK also results in advanced mitochondrial biogenesis by increasing PGC-1α expression [99]. Kobashigawa et al. reported that AMPK is a sensitive target of DOX-induced cardiotoxicity [100]. DOX could inhibit AMPK phosphorylation and activate downstream mTOR to cause pathological cardiac phenotype [101]. SIRT1 was considered to be implicated in the metabolic regulation of AMPK in the heart. A study on SIRT1 in the treatment of I/R injury discovered that SIRT1 could improve cardiac function, reduce myocardial infarction size, and attenuate the oleate oxidation to rescue the tolerance of aged hearts to I/R injury via LKB1 deacetylation and subsequent AMPK activation [65]. In addition, the activation of SIRT1/AMPK pathway could improve cardiac metabolic remodeling in ventricular cardiomyocytes induced by aldosterone stimulation [102]. Alternatively, other studies revealed that AMPK could also activate SIRT1 by modulating intracellular NAD⁺ metabolism [103, 104]. Recently, we found that fibroblast growth factor 21 (FGF21) could facilitate the interaction of SIRT1 with LKB1 to enhance LKB1 deacetylation and subsequently activate AMPK to prevent DOX-induced oxidative stress, inflammation, and apoptosis, as well as ameliorate cardiac dysfunction partly through elevating nuclear NRF2 expression and inhibiting NF-κB expression both in vivo and in vitro[105]. In addition to the regulation of oxidative stress, inflammation, and apoptosis, AMPK is also a crucial regulator of autophagy in DOX-induced cardiotoxicity [49]; thus, whether this protective effect is dependent on the expression of SIRT1 remains obscure and needs to be further investigated.

SIRT1/p53 pathway

p53 is a primary tumor suppressor that mainly acts as a transcription factor in response to numerous stressors to control cell proliferation, senescence, apoptosis, and DNA repair [106]. Under normal circumstances, murine double minute-2 (Mdm2), an E3 ubiquitin ligase, targets p53 for proteasomal degradation and maintains the expression of p53 at a low-level. In response to acute stress, Mdm2 is inactivated, following which, it elevates p53 level that prohibits cell division and induces apoptosis [107]. A study on angiotensin II-induced cardiomyocyte apoptosis found that SIRT1 could inhibit p53 acetylation and its binding to the mitochondrial fission protein Drp1 promoter to suppress mitochondrial fission and cardiomyocyte apoptosis [108]. In addition, SIRT1/p53 pathway was also reported to be involved in the treatment of myocardial hypoxia/reoxygenation injury [68]. Generally, SIRT1 could regulate cardiac apoptosis through both p53 transcription-dependent and p53 transcription-independent pathways. The former is effectuated on the expression of the apoptosis-related target genes, such as Bax, PUMA, and NOXA [109], whereas the latter is initiated through Cyt C release from the mitochondria intramembrane [41]. For mouse models with DOX treatment, DOX-induced cardiomyocyte apoptosis was amplified through promoting Cyt C release from mitochondria and increasing p53 acetylation and p53-dependent transcription of Bax. These effects were inhibited by resveratrol. Moreover, resveratrol could also upregulate SIRT1 expression, suggesting that the cardioprotective effect of resveratrol was associated with SIRT1-mediated deacetylation of p53 and subsequent inhibition of p53 transcription-dependent and transcription-independent apoptosis [110]. Furthermore, Zhang et al. demonstrated that HSP25 knockdown could increase p53 acetylation on K379 by attenuating the correlation between SIRT1 and p53 to aggravate DOX-induced H9c2 cell apoptosis [111]. Therefore, pharmacological interventions that regulate SIRT1/p53 pathway might provide an effective pathway for the treatment of DOX-induced cardiotoxicity.

SIRT1/p66Shc pathway

p66Shc is a member of the spontaneous human combustion (shc) family that regulates the redox balance in the cells via elevation of mitochondrial ROS production [112]. p66Shc deletion could improve oxidative stress and cardiac dysfunction in pressure overload-induced heart failure [113]. Consistent with this study, Liu et al. reported that miR-124 could decrease ROS generation and MDA level, as well as increase activity of SOD to alleviate DOX-induced oxidative stress in the heart through inhibiting p66Shc expression [114], suggesting that inhibition of p66Shc might be a potential therapeutic strategy for DOX-induced cardiotoxicity. SIRT1 has emerged as a critical upstream regulator of p66Shc, while deletion of cardiomyocyte-specific Sirt1 gene could increase oxidative stress of aged hearts in response to ischemic insults by enhancing p66Shc phosphorylation [65]. Moreover, overexpression of SIRT1 could decrease the level of p66Shc by binding to the p66Shc promoter to prevent hyperglycemia-induced endothelial dysfunction through reducing oxidative stress and the expression of endothelial dysfunction marker plasminogen activator inhibitor-1 [115]. On the other hand, p66Shc was reported to be positively correlated with pro-apoptotic Bax expression and caspase-3 activation, while that is negatively associated with the expression of anti-apoptotic Bcl-2. Zhu et al. demonstrated that miR-34a-5p could inhibit SIRT1 expression and increase p66shc level to promote cardiomyocyte apoptosis after DOX treatment, highlighting that miR-34a-5p/SIRT1/p66Shc is a significant axis that is conducive to DOX-induced cardiotoxicity [116]. Moreover, another study showed that berberine could upregulate SIRT1 expression and subsequently downregulate p66Shc expression to exert antioxidative and anti-apoptotic effects, and improve mitochondrial dysfunction, thereby ameliorating DOX-induced cardiotoxicity [117]. The above-mentioned findings indicate a potential therapeutic target of SIRT1/p66Shc pathway in DOX-induced cardiotoxicity.

SIRT1/NF-κB pathway

NF-κB, known as a critical regulator of inflammatory and immunological responses, plays a pivotal role in the development of DOX-induced cardiotoxicity. A previous study pointed out that DOX could upregulate NF-κB expression to cause cardiac inflammation [118]. SIRT1 has been exhibited to inhibit NF-κB expression through deacetylation of RelA/p65 subunit of NF-κB on Lys310 to regulate inflammatory phases and energy supply for metabolic processes. Concurrently, the deacetylation of RelA/p65 protein on Lys310 makes its own methylation on Lys314 and Lys315 to enhance its ubiquitination and degradation [119, 120]. Bagul et al. found that SIRT1 activation by resveratrol pre-treatment could inhibit NF-κB expression to improve cardiac oxidative stress and inflammation in diabetic rats [73]. SIRT1/NF-κB pathway also participates in the inhibition of myocardial apoptosis to protect H9c2 cells against hypoxia-induced injury [121]. A recent study by Yuan et al. indicated that overexpression of SIRT1 by C1q/tumor necrosis factor-related protein-3 (CTRP3) could exert anti-inflammatory effect to improve DOX-induced cardiotoxicity through reduced levels of TNF-α and nuclear translocation of NF-κB [86]. Based on these findings, the anti-inflammatory effect of SIRT1 on DOX-induced cardiotoxicity might be partially due to the inhibition of NF-κB, while further investigation is required to elucidate the regulatory mechanism of NF-κB by SIRT1 in DOX-induced cardiotoxicity.

SIRT1/p38 MAPK pathway

p38 MAPK, a stress-activated kinase, can be induced by inflammatory response and osmotic stress. The activity of p38 MAPK is mediated by the upstream kinases, known as MAPK kinases (MKK), such as MKK3 and MKK6 [122]. Previous studies pointed out that p38 MAPK activation was involved in several cardiac pathological changes, and p38 MAPK inhibition could improve cardiac fibrosis [123], pressure-loaded right ventricular hypertrophy [124], cardiac oxidative stress, and myocardial I/R injury [125]. Guo et al. found that the inhibition of p38 MAPK/NF-κB pathway could decrease the levels of inflammatory factors IL-1β, IL-6, and TNF-α, which in turn, protected H9c2 cells from DOX toxicity [126]. SIRT1 has been reported to be a negative regulator of p38 MAPK. Previously, it was shown that the overexpression of SIRT1 could attenuate mitochondrial dysfunction, oxidative stress, and apoptosis to improve I/R injury of cardiomyocytes. Further mechanistic investigation revealed that the cardioprotective role of SIRT1 is involved in reduction of p38 MAPK and JNK phosphorylation as well as enhancement of ERK phosphorylation [127]. In addition, SIRT1/MAPK pathway is known to participate in the treatment of DCM [128]. Another study focused on SIRT1 in the treatment of DOX-induced cardiotoxicity and demonstrated that SIRT1 could inhibit oxidative stress and cardiomyocyte apoptosis partially via the inhibition of p38 MAPK expression [129]. Currently, only a limited number of studies are available on the role of SIRT1/p38 MAPK pathway in DOX-induced cardiotoxicity. Therefore, further studies are required to clarify the exact functions and mechanisms of SIRT1/p38 MAPK pathway.

SIRT1/FOXOs pathway

FOXO transcription factors, including four members, FOXO1, FOXO3, FOXO4, and FOXO6 in mammals, are widely involved in stress response, cellular homeostasis, and longevity. To carry out these functions, FOXOs regulate various cellular processes, such as oxidative stress, metabolic homeostasis, protein homeostasis, cell apoptosis, and repair of DNA damage [130]. In normal physiological conditions, FOXO1 are primarily localized in the cytoplasm. Under various stress stimulations, SIRT1 could deacetylate FOXO1 at K242, K245, and K262 and enhance the nuclear localization of FOXO1 in the heart [131–133]. Moreover, SIRT1 could enhance the abilities of oxidative stress resistance and DNA damage repair in the heart through the deacetylation and activation of FOXOs [132, 134]. On the contrary, other studies reported that SIRT1 negatively regulates the expression of FOXOs; for instance, Wang et al. found that the ability of FOXO3a could be inhibited by SIRT1 to improve oxidative stress-induced endothelial progenitor cell apoptosis through ubiquitination and degradation of FOXO3a [135]. Consistent with this study, resveratrol could protect the heart from DOX-induced apoptosis accompanied by increased expression of SIRT1, and reduced expression of FOXO1 and its target cell death gene Bim. Therefore, the anti-apoptotic function of SIRT1 on DOX-induced cardiotoxicity might be associated with the negative regulation of FOXO1 expression [136]. The above controversial reports about the regulation of FOXOs by SIRT1 might be related to the different post-translational modifications of FOXOs. FOXOs deacetylation by SIRT1 might increase its transcription and activity, whereas deacetylation of FOXOs promotes its own poly-ubiquitination and degradation [137]. Therefore, the specific effects and mechanisms related to the SIRT1/FOXOs pathway in DOX-induced cardiotoxicity need further investigation.

SIRT1/TGF-β pathway

TGF-β family is a multipotent cytokine involved in numerous cellular functions, such as modulating cell growth, proliferation, differentiation, and apoptosis. It can also regulate the production of ECM, including collagen and fibronectin. TGF-β family has three structurally similar isoforms: TGF-β1, TGF-β2, and TGF-β3, encoded by three different genes [138, 139]. A previous study demonstrated that TGF-β could directly stimulate its downstream targets SMADs to induce the overexpression of pro-fibrotic genes [140]. SIRT1 plays a potential regulatory role in abolishing TGF-β-induced collagen synthesis and myofibroblast differentiation. Li et al. found that SIRT1 activation by tetrahydrocurcumin could inhibit TGF-β1/SMAD3 pathway to exercise anti-fibrotic function and ameliorate DCM [72]. Another study on isoproterenol-induced cardiac fibrosis demonstrated that SIRT1 exerts a protective role by endothelial-to-mesenchymal transition via downregulation of TGF-β/SMAD2/3 pathway [141]. In addition, TGF-β is known to be involved in the occurrence of DOX-induced cardiotoxicity[142]. In a rat model of DOX-induced cardiotoxicity, activation of SIRT1 by resveratrol could decrease the levels of TGF-β and pSMAD3/SMAD3 ratio, as well as the expression of fibronectin and type-I collagen, which subsequently inhibited cardiac fibrosis and fibroblast activation to improve diastolic dysfunction and myocardial remodeling [88]. Therefore, the anti-fibrotic function mediated by SIRT1/TGF-β pathway may play an indispensable function in the treatment of DOX-induced cardiotoxicity.

SIRT1/NLRP3 pathway

NLRP3, a critical regulator of the innate immune system, is activated in response to the stimulation of endogenous and exogenous factors [143]. The activation of NLRP3 inflammasome results in the activation of caspase-1 and transforms pro-IL-1β and pro-IL-18 into their bioactive forms (IL-1β and IL-18), leading to inflammatory response and tissue injury [144]. Moreover, NLRP3-induced innate immune response could lead to pyroptosis, an inflammatory form of programmed cell death [145]. Previous studies showed that NLRP3 inflammasome plays a critical role in the pathogenic mechanism of cardiovascular diseases, including myocardial I/R injury [146], atherosclerosis [147], and DOX-induced cardiotoxicity [148]. Furthermore, it was previously found that SIRT1 could downregulate the NLRP3 inflammasome to modulate cellular inflammation [149]. A recent study on the protective role of SIRT1 in myocardial I/R injury demonstrated that SIRT1 could protect cardiomyocytes from I/R injury through inhibiting NLRP3-dependent inflammation and pyroptosis via Akt-dependent metabolic regulation [150]. Recently, Zhai et al. pointed out that SIRT1 activation by calycosin could not only upregulate the cardiac activity of glutathione peroxidase, CAT, and SOD but also decrease the levels of ROS, MDA, IL-1β, and NLRP3 in DOX-treated mice, suggesting that the protective effects of calycosin on DOX-induced cardiac dysfunction are involved in SIRT1/NLRP3 pathway [151]. In addition, dihydromyricetin could also protect against DOX-induced inflammation by SIRT1/NLRP3 pathway [23]. Hence, it could be concluded that SIRT1/NLRP3 signaling pathway is a potential therapeutic target for DOX-induced cardiotoxicity; however, the specific mechanisms related to the inhibition of NLRP3 by SIRT1 in DOX-induced cardiotoxicity remain elusive, highlighting the necessity of conducting further research.

SIRT1 agonists and their roles in DOX-induced cardiotoxicity

As discussed earlier, the activation of SIRT1 is a major cardioprotective therapy for DOX-induced cardiotoxicity. A number of substances, such as resveratrol, pterostilbene, melatonin, erythropoietin, berberine, sesamin, CTRP3, FGF21, dihydromyricetin, and calycosin, stimulate SIRT1 to exert protective functions on DOX-induced cardiotoxicity (summarized in Table 3). Resveratrol is the earliest and most studied SIRT1 agonist that can stimulate SIRT1 to antagonize DOX-induced cardiotoxicity. It is a natural phytoalexin that belongs to polyphenols and is mainly found in the skin of grapes and red wines [152]. Numerous studies indicated that resveratrol increased SIRT1 expression to alleviate DOX-induced mitochondrial dysfunction, ER stress, cardiac fibrosis, and apoptosis [83, 87, 88, 110]. Pterostilbene is a natural resveratrol analogue and acknowledged as an antioxidant found in grapes and blueberries [153]. Relevant studies have shown that pterostilbene is an effective myocardial protective agent to prevent various heart diseases, such as I/R injury, myocardial infarction, and DCM through free radical elimination and anti-inflammatory roles [154–156]. It also upregulates SIRT1 and AMPK cascades and subsequently activates PGC-1α to resist DOX-induced mitochondrial injury and oxidative stress [97]. Notably, a clinical study demonstrated that pterostilbene is generally safe for the human body and can attenuate adult blood pressure [157].

Table 3.

The main roles of each SIRT1 agonist in DOX-induced cardiotoxicity

SIRT1 agonist	Roles in DOX-induced cardiotoxicity
Resveratrol	Regulate mitochondrial function, inhibit ER stress, anti-fibrosis and anti-apoptosis
Pterostilbene	Antioxidation and regulate mitochondrial function
Melatonin	Regulate mitochondrial function and anti-apoptosis
Erythropoietin	Regulate mitochondrial function and biogenesis
Berberine	Regulate mitochondrial function, antioxidation and anti-apoptosis
Sesamin	Antioxidation
CTRP3	Anti-inflammation and anti-apoptosis
FGF21	Antioxidation, anti-inflammation and anti-apoptosis
Dihydromyricetin	Anti-inflammation
Calycosin	Anti-inflammation

SIRT1 Sirtuin 1, DOX doxorubicin, ER endoplasmic reticulum, CTRP3 C1q/tumor necrosis factor-related protein-3, FGF21 fibroblast growth factor 21

Melatonin is another drug that enhances the expression of SIRT1 and PGC-1α and improves DOX-mediated mitochondrial dysfunction and cardiomyocyte apoptosis [96]. Reportedly, melatonin, as a mitochondria-targeted antioxidant, exerts an effect on mitochondrial homeostasis and functions [158]. Erythropoietin and its receptor on hearts have been manifested to play pivotal protective roles in myocardial I/R injury and cyanotic congenital heart disease by modulating mitochondrial morphology and biogenesis [159, 160]. The beneficial functions of erythropoietin mentioned above are also effective in antagonizing DOX-induced cardiotoxicity, which is implicated in the regulation of SIRT1/PGC-1α pathway [95]. In addition, berberine, a broad-spectrum antibiotic with antineoplastic and cardioprotective effects, also inhibits DOX-induced mitochondrial dysfunction by adjusting ∆Ψm, mitochondrial Ca²⁺ concentration, and mitochondrial biogenesis. Importantly, pre-treatment with berberine upregulates SIRT1 expression and downregulates p66Shc expression to exercise antioxidative and anti-apoptotic functions in the heart following DOX exposure [117, 161]. Sesamin, as another SIRT1 agonist with antioxidative and anti-hypertensive roles, ameliorates DOX-induced cardiotoxicity through the regulation of manganese SOD (MnSOD) expression [84]. Moreover, CTRP3, an endogenous lipopolysaccharide antagonist, activates SIRT1 and subsequently reduces DOX-induced inflammation and apoptosis [86]. As an effective adjuster of glucose and lipid metabolism in adipose and liver tissue, FGF21 is also verified to express in the myocardium. In a previous study, we found that it could restore the DOX-disrupted interaction of SIRT1 and LKB1 and then decrease LKB1 acetylation, thereby inducing AMPK activation to improve cardiac oxidative stress, inflammation, and apoptosis [105]. Recently, Sun et al. pointed out that dihydromyricetin, a flavonoid compound with cardioprotective effect extracted from the Japanese raisin tree (Hovenia dulcis), could inhibit DOX-induced activation of NLRP3 inflammasome to protect against cardiac inflammation through stimulating SIRT1 signaling [23]. Furthermore, as the primary active constituent in astragalus membranaceus with anti-tumor, anti-inflammatory, and cardioprotective roles, calycosin also activates SIRT1/NLRP3 pathway to ameliorate DOX-induced inflammation [151]. Although the effects of numerous SIRT1 agonists on DOX-induced cardiotoxicity have been reported, various animal or cellular models and different doses and timescales of DOX make it difficult to reach consensus on protective molecular mechanisms. Moreover, some SIRT1 agonists might not be specific, and further assessment is required to identify specific SIRT1 agonists for clinical application.

Conclusion

SIRT1 plays a protective role in DOX-induced cardiotoxicity by modulating various signaling pathways. Although previous studies have reported the protective effects of SIRT1 on DOX-induced cardiotoxicity in rodent models, no systematic study has yet evaluated the effects of clinical combination therapy and the optimum time to take precaution. In addition, several studies ignored many aspects of human cancer management in clinical practice; for example, various studies used a single dose or multiple high doses and intraperitoneal injection of DOX to treat animals, while human cancer management was typically implemented with continuous cycles of DOX and intravenous administration. Therefore, a recent editorial emphasized the importance of horizontally integrating research on DOX-induced cardiotoxicity, such as using intravenous administration and small repeated doses in animal models [162]. Notably, the experimental results in animals need to be observed and confirmed in humans. Another study on mouse model has verified that an appropriate amount of SIRT1 (2.5- to 7.5-fold) expression is beneficial for the heart, whereas higher levels of SIRT1 (12.5-fold or greater) can trigger cardiomyopathy [78]. Thus, appropriate doses of SIRT1 agonists should be carefully evaluated for the optimal use of the therapeutic potential of SIRT1 in clinical application.

Notably, although numerous studies have shown that resveratrol, as an effective agonist of SIRT1, plays a critical role in DOX-induced cardiotoxicity, its application in clinical practice is limited due to low solubility in water and poor systemic bioavailability [163]. In order to improve the bioavailability, newer methods of delivery need to be developed so that it can be adequately absorbed from the gut. Recently, Quagliariello et al. reported that nanoemulsions loaded with anti-inflammatory nutraceuticals and nano-encapsulation of coenzyme Q10 in nano-emulsions acted against DOX-induced cardiotoxicity, thereby providing an effective method of delivery [21, 164]. Resveratrol solid lipid nanoparticles and PLGA nanoparticles have been found to be submicron drug delivery systems for delivering resveratrol, enabling its gradual release and better distribution in the body [165, 166]. Furthermore, SIRT1, as a redox-sensitive deacetylase, can be downregulated under oxidative/inflammatory conditions through post-translational modification [167]. Thus, reversing the post-translational modification of SIRT1 might be an ideal approach to treat chronic diseases associated with oxidative stress and inflammation. In conclusion, further in-depth studies on the effects and mechanisms of SIRT1 would provide novel ideas for the prevention and treatment of DOX-induced cardiotoxicity.

Funding

This study was supported by Qilu Young Scholar’s Program of Shandong University (21330089963007), National Natural Science Foundation of China (81700329, 81770375) and Jilin Science and Technology Department (20200801061GH).

Compliance with ethical standards

Conflict of interest

There are no conflicts of interest.