Targeting both ferroptosis and pyroptosis may represent potential therapies for acute liver failure

Abstract

In this editorial, we comment on the article published in the recent issue of the World Journal of Gastroenterology. Acute liver failure (ALF) is a fatal disease that causes uncontrolled massive hepatocyte death and rapid loss of liver function. Ferroptosis and pyroptosis, cell death forms that can be initiated or blocked concurrently, can play significant roles in developing inflammation and various malignancies. However, their roles in ALF remain unclear. The article discovered the positive feedback between ferroptosis and pyroptosis in the progression of ALF, and revealed that the silent information regulator sirtuin 1 (SIRT1) inhibits both pathways through p53, dramatically reducing inflammation and protecting hepatocytes. This suggests the potential use of SIRT1 and its downstream molecules as therapeutics for ALF. Thus, we will discuss the role of ferroptosis and pyroptosis in ALF and the crosstalk between these cell death mechanisms. Additionally, we address potential treatments that could alleviate ALF by simultaneously inhibiting both cell death pathways, as well as examples of SIRT1 activators being used as disease treatment strategies, providing new insights into the therapy of ALF.

Key Words: Acute liver failure; Ferroptosis; Pyroptosis; Crosstalk; Silent information regulator sirtuin 1; P53; Glutathione peroxidase 4; Gasdermin D; Treatment

Core Tip: Acute liver failure (ALF) is a life-threatening disease characterized by uncontrolled death of hepatocytes. Ferroptosis and pyroptosis are two recently discovered types of cell death that can occur simultaneously. However, their roles in ALF remain unclear. The findings show that these two cell death pathways work together to advance ALF and suggest that silent information regulator sirtuin 1 (SIRT1) and its downstream molecules could be potential therapeutics for ALF. Therefore, we will discuss the roles and crosstalk of ferroptosis and pyroptosis in ALF. Activation of SIRT1 and suppression of both cell death pathways may offer new insights into therapeutic targets for ALF.

INTRODUCTION

Acute liver failure (ALF) is an acute clinical syndrome that occurs in patients without a history of liver disease and is life-threatening[1]. In developed countries, there are approximately 1–6 cases per million individuals. Although the incidence is rare, ALF causes a mortality rate as high as 30%[2,3]. The most prevalent causes of ALF are hepatic toxicity induced by medications or poisons, acute viral hepatitis, autoimmune and metabolic disorders, and unexplained cryptogenic liver failure[1,3]. ALF can lead to severe complications, including coagulopathy, elevated transaminases, hepatic encephalopathy, and multi-organ failure. This is often attributed to a series of severe proinflammatory states triggered by extensive hepatocyte damage, resulting in DNA damage, oxidative stress, and an accompanying inflammatory factor storm[4-6].

The main event in ALF is the excessive and uncontrolled death of hepatocytes by apoptosis, necroptosis, and necrosis[5,7,8]. Further study has found that various types of cell death are related to liver diseases such as ALF, including ferroptosis and pyroptosis[9]. These cell death pathways can coexist in a pathological environment, and several shared overlapping mechanisms can be used as “backup” death strategies to maintain biological balance within the organism when the death induction threshold is reached, mediating various immune effects and inflammatory responses[10].

FERROPTOSIS IN ALF

Dixon et al[11] originally described ferroptosis, an iron-dependent regulated cell death, in 2012. It is characterized by iron metabolism disorder and an accumulation of excessive intracellular lipid peroxides, which lead to redox imbalance and ultimately cell death[12].

When iron metabolism is disrupted, excess iron can be released from iron storage proteins into the cytoplasm or other organelles. Free iron can react with intracellular hydrogen peroxide or other oxidants to cause lipid peroxidation of polyunsaturated fatty acids in the cell membrane via the Fenton reaction, which produces a variety of reactive oxygen species (ROS) and lipid peroxidation radicals[13,14]. System X_c^- is a key regulatory component of ferroptosis, consisting of solute carrier family 7 member 11 (SLC7A11) and solute carrier family 3 member 2[15], whose antioxidant system typically protects cells from oxidative stress[16]. However, during ferroptosis, the antioxidant system may fail to efficiently neutralize the produced ROS, resulting in the cell’s incapacity to withstand lipid peroxidation[17].

The p53 gene is essential in the cellular response to a variety of stressors, including DNA damage, hypoxia, nutrient starvation, and oncogene activation[18]. P53 can also regulate ferroptosis by directly acting on the SLC7A11 promoter to reduce its expression, thereby reducing extracellular cystine intake, decreasing glutathione synthesis, lowering glutathione peroxidase 4 (GPX4) activity, increasing lipid peroxide levels, and ultimately leading to ferroptosis[19]. P53 can also recruit the deubiquitinase ubiquitin-specific peptidase 7 to the histone H2B monoubiquitination modification (H2Bub1) in the promoter region of SLC7A11, reducing H2Bub1 on the SLC7A11 gene, resulting in lower SLC7A11 protein levels and ferroptosis[20]. However, in addition to promoting ferroptosis, p53 may also inhibit it by regulating the localization and activity of dipeptidyl peptidase 4 and other pathways, thereby promoting cell survival[21]. Therefore, p53 may exhibit a “dual role” in the regulation of ferroptosis.

Ferroptosis is thought to play a substantial role in hepatocyte death in ALF[22-24]. One study reported that the ferroptosis inhibitor ferrostatin-1 significantly prevented hepatotoxicity and lipid peroxidation in mice with acetaminophen (APAP) induced ALF, lowering the mortality rate[25]. Another study revealed similar outcomes[26]. The iron inhibitor UAMC3203 and the iron chelator deferoxamine both protected against APAP-induced liver damage by reducing ferroptosis[26-28]. Moreover, various medicines or biological extracts, such as sulforaphane[29], (+)-clausenamide[30], nitroflurbiprofen[31], avicularin[32], and glycyrrhizin[33], have been demonstrated to reduce liver damage by ferroptosis inhibition. These studies highlight the importance of ferroptosis in ALF, suggesting that blocking the ferroptosis pathway could be a viable strategy for ALF treatment. According to the study by Zhou et al[34], histone deacetylases silent information regulator sirtuin 1 (SIRT1) regulates ferroptosis through the p53 pathway, offering a prospective therapeutic option.

PYROPTOSIS IN ALF

Pyroptosis is a type of immunogenic cell death mediated by caspases during microbial infections[35]. It helps the body eliminate invading pathogens. The gasdermin D (GSDMD) protein is the central driver of pyroptosis. When a cell receives signals such as pathogen-associated molecular patterns, damage-associated molecular patterns, and lipopolysaccharide (LPS) via classical or non-classical pathways, caspases-1 and caspases-4/5/11 are recruited and activated. The activated caspases perform proteolytic activity, leading to the formation of the active N-terminal fragment of GSDMD (GSDMD-N)[35]. It also promotes the cleavage of interleukin-1beta precursor and interleukin-18 precursor to produce mature cytokines[36,37]. Subsequently, GSDMD-N binds to acidic phospholipids on the cell membrane, forming oligomerized death-inducing pores and increasing intracellular osmotic pressure, leading to cell swelling and rupture[38-40]. It leads to the release of interleukin-1beta (IL-1β), interleukin (IL)-18, tumor necrosis factor-alpha (TNFα), ATP, and other substances into the extracellular space, attracting immune cells to the injury site and mediating the inflammatory immune response[37,41,42].

Previous studies have indicated that pyroptosis plays a crucial role in liver diseases. ALF patients’ liver tissue exhibits elevated levels of molecules associated with pyroptosis, including GSDMD-N, caspases-1/4, IL-1β, IL-18, and TNFα. In vitro research has shown that reducing GSDMD can lower MCP1/CCR2 protein levels, thereby decreasing neutrophil-mediated immune injury in the liver. In mouse models, deletion of the GSDMD gene effectively reduces liver inflammatory injury and increases survival rates in mice with D-Galn/LPS-induced ALF[43]. Moreover, several studies have shown that the use of pyroptosis inhibitors such as VX-765[44], GSDMD inhibitor necrosulfonamide[45], GSK3β inhibitor TDZD-8[46], limonin[47], 3,4-dihydroxyphenylethanol glycoside (DAG)[48], and tyrosine-alanine (YA)[49], can alleviate liver cell pyroptosis, reduce oxidative stress and inflammation, and improve liver injury. However, the regulatory mechanisms remain unclear. Additionally, p53-induced pyroptosis has been reported in several studies[50-53], but it is rarely reported in the liver.

CROSSTALK BETWEEN FERROPTOSIS AND PYROPTOSIS IN ALF

Given that pyroptosis and ferroptosis are often simultaneously inhibited or encouraged in tissue injury or cancers[54-56], researchers have explored their relationship. Some studies have demonstrated a mutual regulatory link between ferroptosis and pyroptosis. For example, a deficiency of GPX4 in bone marrow cells might increase GSDMD production via caspase-1/11, resulting in pyroptosis[57]. In the diabetic retinopathy model, the ferroptosis inhibitor Ferr-1 can reduce GSDMD expression, thereby inhibiting pyroptosis and improving retinal tissue damage[58]. Chlorpyrifos promotes GSDMD cleavage and increases intracellular ROS levels, which in turn enhances p53-mediated ferroptosis[59]. The Stat3/p53/nuclear factor-E2-related factor 2 axis regulates both ferroptosis and pyroptosis in colorectal cancer cells[60]. These findings indicate that there may be crosstalk between ferroptosis and pyroptosis in diseases.

ALF is characterized by excessive death of hepatocytes. Therefore, crosstalk and co-activation of multiple death pathways are likely important mechanisms. However, there is currently little research on the combined role of ferroptosis and pyroptosis in ALF. A report revealed that treatment with YA and DAG can simultaneously reduce ferroptosis and pyroptosis in an ALF mouse model, thereby protecting the liver from damage and reducing mouse mortality[48,49]. Zhou et al[34] found that both ferroptosis and pyroptosis are triggered in the liver tissue of ALF patients. Inhibiting ferroptosis or pyroptosis protected mice from LPS/D-GalN-induced ALF. Furthermore, in the LPS/D-GalN-induced ALF mouse model, inhibiting GPX4 promoted ferroptosis and increased the expression of the pyroptosis marker GSDMD. Supplementation with GPX4 inhibited both ferroptosis and pyroptosis. It was found that the absence of GSDMD reduces not only pyroptosis but also ferroptosis in GSDMD knockout mice[34]. As a result, ferroptosis has a positive feedback regulatory effect on pyroptosis (Figure 1). Further identification of critical molecules or drugs that target the ferroptosis and pyroptosis processes may reveal potential strategies for ALF prevention and treatment.

Figure 1 The positive feedback loop between ferroptosis and pyroptosis in acute liver failure, as well as possible treatments that target ferroptosis and pyroptosis. Glutathione peroxidase 4 (GPX4) reduces the expression of gasdermin D (GSDMD), a pyroptosis marker. In turn, GSDMD can reduce the level of GPX4. Silent information regulator sirtuin 1 can inhibit GSDMD expression while elevating System X_c^-via p53, hence targeting ferroptosis and pyroptosis. Acute liver failure mice treated with tyrosine-alanine or 3,4-dihydroxyphenylethanol glycoside (DAG) show a similar role in suppressing both cell death pathways. GSDMD-N: N-terminal fragment of gasdermin D; GSSG: Oxidized glutathione; IL-1β: Interleukin-1beta; IL-18: Interleukin-18; LPCAT3: Lysophosphatidylcholine acyltransferase 3; LPS: Lipopolysaccharide; PUFAs: Polyunsaturated fatty acids; ROS: Reactive oxygen species; SIRT1: Silent information regulator sirtuin 1; SLC7A11: Solute carrier family 7 member 11; SLC3A2: Solute carrier family 3 member 2; TLR: Toll-like receptors; TNFα: Tumor necrosis factor-alpha; YA: Tyrosine-alanine.

SIRT1 SERVES AS A THERAPEUTIC TO ALLEVIATE ALF VIA INHIBITING BOTH FERROPTOSIS AND PYROPTOSIS

SIRT1 is a class of nicotinamide adenine dinucleotide (+)-dependent histone deacetylases that regulate the deacetylation of histones and other proteins. The targets include p53, forkhead box class O1/3/4, heat shock factor1, hypoxia-inducible factor 1alpha, nuclear factor kappa B[61,62]. SIRT1 regulates a variety of activities associated with anti-aging and oxidative stress, including apoptosis, autophagy, mitochondrial function, DNA damage repair, metabolism, and inflammation[62,63]. SIRT1 down-regulation correlates with cell aging and increased inflammatory factors, such as IL6, IL8, and IL1B/IL-1β[64]. A recent study found that activating SIRT1 could inhibit ferroptosis induced by excessive iron through autophagy in foam cells, providing a new therapeutic target for atherosclerosis[65].

SIRT1 also plays a role in ALF (Table 1)[66-73]. Several studies have shown that the SIRT1 signal in ALF reduces cellular oxidative stress and inhibits hepatocyte death[66-68,74,75]. Further studies revealed that the p53 signaling pathway is involved in the process[69,70,76]. SIRT1 can deacetylate p53, promote autophagy, inhibit oxidative stress, and reduce inflammatory responses[70].

Table 1 Current studies on the role of silent information regulator sirtuin 1 in acute liver failure.

Article title	Core tips	Ref.
Sirtuin 1 attenuates ALF by reducing reactive oxygen species via hypoxia-inducible factor 1α	Resveratrol, a SIRT1 activator, deacetylates hypoxia-inducible factor 1alpha and inhibits its activity, reducing ALF caused by hypoxia, reactive oxygen species, and apoptosis	Cao et al[66]
Short-term fasting attenuates lipopolysaccharide/D-galactosamine-induced ALF through SIRT1-autophagy signaling in mice	Short-term dietary restriction activates the SIRT1 signaling pathway, regulates autophagy, and reduces hepatocytes apoptosis in ALF	Long et al[67]
Fusobacterium nucleatum promotes the development of ALF by inhibiting the NAD+ salvage metabolic pathway	Fusobacterium nucleatum suppresses NAD+ and the SIRT1/adenosine monophosphate activated protein kinase signaling pathway, causing liver damage in ALF	Cao et al[68]
Hepato-protective effect of resveratrol against acetaminophen-induced liver injury is associated with inhibition of CYP-mediated bioactivation and regulation of SIRT1-p53 signaling pathways	Resveratrol helps to heal liver impairment caused by APAP by blocking CYP-mediated APAP bioactivation and regulating SIRT1, p53, cyclin D1, and proliferating cell nuclear antigen	Wang et al[69]
Apigenin prevents acetaminophen-induced liver injury by activating the SIRT1 pathway	Apigenin prevents acetaminophen-induced liver injury by regulating the SIRT1-p53 axis, which promotes autophagy and reduces the inflammatory response and oxidative stress caused by acetaminophen	Zhao et al[70]
Sirtuin-activating compounds alleviate D-galactosamine/lipopolysaccharide-induced hepatotoxicity in rats: Involvement of sirtuin 1 and heme oxygenase 1	Quercetin and SRT1720 can activate SIRT1 protein and inhibit HO-1 expression, reducing liver damage caused by D-galactosamine/lipopolysaccharide in rats	Kemelo et al[71]
The sirtuin 1 activator SRT1720 alleviated endotoxin-induced fulminant hepatitis in mice	SRT1720, a SIRT1 activator, reduces fulminant hepatitis caused by lipopolysaccharide/D-Gal, potentially through inhibiting tumor necrosis factor-alpha production and activating the apoptotic cascade	Zhou et al[72]
Evaluation of the reparative effect of SIN in an acetaminophen-induced liver injury model	SIN successfully treats acetaminophen-induced liver injury by restoring SIRT1 levels, lowering oxidative stress, and repairing cell damage	Kayalı et al[73]

ALF: Acute liver failure; APAP: Acetaminophen; CYP: Cytochrome; NAD+: Nicotinamide adenine dinucleotide; SIN: Sinomenine; SIRT1: Silent information regulator sirtuin 1.

Zhou et al[34] discovered that SIRT1 expression was reduced in human ALF liver tissue. Using SIRT1 activators or overexpressing SIRT1 could inhibit both ferroptosis events (reduced iron deposition and ROS activity, decreased expression of Acyl-CoA synthetase long-chain family 4, and increased expression of SLC7A11 and GPX4) and the expression of the pyroptosis marker GSDMD, thereby alleviating acute liver injury. In LPS/D-GalN-induced in vitro and in vivo models, the deactivation of SIRT1 increased ferroptosis and pyroptosis, exacerbating liver injury. Further research revealed that the inhibition of ferroptosis and pyroptosis by SIRT1 may depend on p53 deacetylation[34]. These results suggest that SIRT1 may serve as a molecular target to suppress multiple death processes and as a potential treatment for ALF.

Classical SIRT1 activators have been discovered, including natural ones like resveratrol, and synthetic compounds derived from the core structure of imidazole (1,2-b) thiazole, such as SRT1720 and SRT2104[77]. There have been no known clinical investigations on ALF treatment using SIRT1 activators. However, SIRT1 activators have shown an excellent safety profile and positive therapeutic effects on several diseases. Clinical trial results indicate that the natural SIRT1 activator resveratrol is effective in treating Alzheimer’s disease[78], obesity and metabolic disorders[79], and polycystic ovary syndrome[80]. The synthetic SIRT1 activator SRT2104 has been demonstrated to ameliorate sepsis[81], psoriasis[82], and blood lipid profiles in older adults[83]. Although natural drugs have some cytotoxicity, synthetic SIRT1 activators address these concerns. However, current medication development remains challenging, and clinical investigations have shown that orally administered SRT2104 has an absolute bioavailability of only 14% (NCT00937872), which is inadequate. Therefore, it may be necessary to develop appropriate drug delivery techniques, modify pharmacochemical structures, or use combination therapy. Kemelo et al[71] reported that quercetin (a natural polyphenol) and SRT1720 showed the ability to improve disease in a rat ALF model, providing evidence that combination therapy may be effective in ALF treatment. In addition, further clinical research will advance the application of SIRT1 activators in ALF treatment.

CONCLUSION

Various forms of cell death, such as apoptosis, necroptosis, necrosis, ferroptosis, and pyroptosis, have been implicated in ALF. However, the crosstalk between them remains unclear. Zhou et al[34] proved that the positive feedback between ferroptosis and pyroptosis plays an important role in hepatocyte mortality in ALF. Mutual stimulation of these cell death pathways may significantly reduce the survival rates of ALF patients. Thus, identifying targets that control the activation of these cell death pathways, as well as medicines that directly suppress these cell death pathways, may provide novel therapeutic strategies to treat ALF. SIRT1 could act as a treatment for ALF, influencing both ferroptosis and pyroptosis. However, it is unclear if SIRT1 can influence other death pathways, such as apoptosis, necroptosis, and necrosis, all of which play essential roles in ALF. Molecular targets capable of effectively controlling multiple cell death pathways have not been reported either. In addition, the efficacy of SIRT1 activation in the clinical treatment of ALF has not been reported. Therefore, addressing these concerns may provide therapeutic targets for treating ALF.