Mechanistic investigation of the crosstalk between hepatic ischemia-reperfusion injury and intestinal microbiota

Introduction

Liver surgery, including liver resection and transplantation, remains the only effective treatment for end-stage liver disease. Hepatic ischemia-reperfusion injury (HIRI) is a major contributor to liver damage in these procedures. It is a pathological process where tissue and organ damage from ischemic blood supply blockage worsens upon reperfusion (1,2). HIRI can trigger systemic inflammatory response syndrome (SIRS) and multiple organ dysfunction syndrome (MODS), significantly contributing to early liver graft dysfunction (3). Traditional research has predominantly concentrated on oxidative stress and inflammatory responses. Recent research suggests that gut microbiota imbalance significantly contributes to the development of HIRI. The gut-liver axis, linking the gut and liver through the portal vein, enables gut microbiota metabolites like short-chain fatty acids (SCFAs), bile acids (BAs), indole derivatives, and lipopolysaccharides (LPS) to regulate hepatic immunity, metabolism, and oxidative balance (4). Although initial studies have explored the therapeutic potential of gut microbiota and its metabolites for liver disease treatment, their effectiveness remains uncertain and debatable. Accordingly, this review seeks to systematically elucidate the bidirectional interactions between gut microbiota and HIRI, analyze the associated molecular regulatory mechanisms, and evaluate the translational potential of microbiota-targeted strategies for the prevention and management of HIRI. A comprehensive understanding of these interactions may provide valuable insights for the development of novel therapeutic targets aimed at mitigating HIRI.

The impact of HIRI on intestinal microbiota

Different types of HIRI, such as warm ischemia, cold storage, partial hepatectomy, and fatty liver transplantation, uniquely affect the gut microbiome and intestinal injury. They can cause intestinal damage by disrupting gut microbiota diversity and structure, leading to imbalanced microbial metabolism, damage to the intestinal mucosal barrier, and bacterial translocation (5) (Figure 1, Table 1). Warm ischemia-type HIRI is predominantly employed in procedures such as partial hepatectomy and liver trauma surgery, wherein the interruption of hepatic blood flow induces warm ischemia. The occlusion of the portal vein results in intestinal venous congestion and mucosal hypoxia, compromising the intestinal barrier and markedly enhancing bacterial and endotoxin translocation. The microbial alterations are characterized by a rapid dysbiosis, notably a reduction in lactobacilli and an increase in enterococci. Cold preservation injury, frequently observed in liver transplantation, predominantly manifests during the ex vivo preservation phase of the liver and following reperfusion post-transplantation. Notably, this process does not involve any disruption of the recipient’s intestinal blood flow. The initial intestinal injury is relatively mild and is primarily attributed to the systemic response elicited by damage-associated molecular patterns (DAMPs) and inflammatory mediators released by the liver after reperfusion, which indirectly compromise the intestinal barrier through the gut-liver axis. In cases where a fatty liver is present, regeneration is inhibited, and dysbiosis becomes more severe and persistent. Fatty livers exhibit heightened sensitivity to ischemia-reperfusion, suffering significant damage accompanied by pronounced sterile inflammation (14). Furthermore, fatty livers can independently alter BA metabolism, thereby affecting the intestinal microenvironment. The baseline microbiota in fatty livers is already altered, as evidenced by changes in the Bacteroidetes/Firmicutes ratio, leading to a distinct trajectory of microbiota changes under HIRI compared to non-fatty livers (15). Severe liver damage results in the release of an increased quantity of DAMPs, which in turn triggers SIRS. This exacerbates and accelerates damage to the gut barrier, leading to significant endotoxemia and bacterial translocation. Although the precise mechanisms underlying these processes remain unclear, the primary effects of HIRI on the gut microbiota can be summarized as follows.

Figure 1 Effect of HIRI on intestinal homeostasis: decreased microbial diversity, proliferation of opportunistic pathogens, intestinal epithelial injury, reduced tight junction proteins, and bacterial translocation. HIRI, hepatic ischemia-reperfusion injury.

Diversity and structural alterations of the microbiota

The gut microbiota, considered a silent organ, is vital for health and disease. Numerous studies have demonstrated a strong correlation between gut microbiota and human health, indicating that a balanced gut microbiota is essential for maintaining overall health (16). The gut microbiota experiences significant changes in diversity and composition during HIRI (6). These modifications directly influence intestinal immune function, the severity of liver damage, and the liver’s regenerative capacity. HIRI significantly decreases gut microbiota diversity by disrupting intestinal barrier integrity and inducing systemic inflammation. This is demonstrated by decreases in alpha diversity (e.g., Shannon and Chao1 indices) and beta diversity (7). A reduction in alpha diversity signifies a homogenization of gut microbiota composition, reflecting a disruption in the stability and complexity of the intestinal microecology. The reduction in beta diversity reflects notable changes in community structure, often marked by a reduction in beneficial gut bacteria and a rise in pathogenic microorganisms. The phenomenon is marked by a rise in pathogenic bacteria like Escherichia coli, Salmonella and Enterococcus, and a decline in beneficial bacteria such as Bifidobacteria and Lactobacilli. Clinical investigations involving liver transplant samples have demonstrated that patients exhibiting elevated levels of Lachnospiraceae-bacterium (Lachn.) tend to experience milder HIRI (8). This dysbiosis of the gut microbiota not only intensifies immune damage in the liver but may also exacerbate liver inflammation via the gut-liver axis. In HIRI, gut microbiota changes are characterized by reduced diversity and structural imbalance. These alterations are intricately linked to the progression and recovery of HIRI, indicating that the gut microbiota may play a critical role in both the pathogenesis and therapeutic management of HIRI.

Intestinal mucosal barrier injury and bacterial translocation

A pivotal factor contributing to gut microbiota imbalance during HIRI is the disruption of intestinal barrier function. HIRI reduces intestinal blood flow and impairs the intestinal barrier’s integrity by triggering a systemic inflammatory response. Intestinal barrier damage allows harmful substances, pathogens, and LPS to translocate from the gut into the bloodstream, reaching the liver through the portal vein and worsening hepatic injury. Research shows that intestinal barrier dysfunction is mainly marked by heightened permeability and reduced levels of tight junction proteins like zonula occludens-1 (ZO-1) and occluding (9,17). This results in the loosening or disintegration of the tight junctions between intestinal epithelial cells, thereby enhancing intestinal permeability. Consequently, the breakdown of intestinal barrier integrity and the onset of metabolic endotoxemia permit gut-derived endotoxins to traverse the compromised intestinal wall into the portal circulation. Endotoxins, bacterial DNA, and short peptides reach the liver through the portal vein, activating Kupffer cells and liver sinusoidal endothelial cells. The activation initiates pro-inflammatory responses via the toll-like receptor 4/myeloid differentiation primary response gene 88/nuclear factor kappa B (TLR4-MyD88-NF-κB) pathway, exacerbating hepatocyte damage and possibly leading to SIRS and MODS (18).

Disruption of the gut microbiota metabolic profile

Metabolomic analysis reveals that HIRI results in the suppression of SCFAs production pathways in the gut, attributed to a reduction in beneficial bacterial populations, including Bifidobacteria and Lactobacilli, with a pronounced impact on butyrate synthesis. Butyrate, a SCFA with strong anti-inflammatory effects, is essential for preserving gut barrier integrity. Reduced production can impair gut barrier function, leading to increased intestinal permeability and the translocation of pathogenic bacteria and LPS into the systemic circulation (10). Concurrently, the composition and proportion of BAs undergo substantial alterations. Under physiological conditions, the synthesis and transformation of BAs are orchestrated by the liver and gut microbiota; however, HIRI perturbs this regulatory axis, precipitating BAs metabolic dysregulation. Changes in gut microbiota affect BA transformation and reabsorption, leading to increased levels of secondary BAs like deoxycholic acid and lithocholic acid, while hindering the production of hepatoprotective primary BAs, thus worsening liver damage (11,19).

Furthermore, the metabolism of aromatic amino acids, including tryptophan and phenylalanine, as well as branched-chain amino acids by the gut microbiota, is also impacted. HIRI inhibits the metabolic pathway that allows gut microbiota to convert tryptophan into indole and its derivatives, including indolepropionic acid. This inhibition results in reduced levels of these metabolites and a diminished cellular protective function against HIRI (20). Conversely, the metabolic activity of pathogenic bacteria may be enhanced, leading to increased production of deleterious metabolites such as phenylacetate and p-cresol, which can accumulate in the intestine and bloodstream. Currently, no direct empirical evidence shows that HIRI increases harmful microbial metabolites. Alterations in lipid metabolism during HIRI also represent a critical aspect of gut microbiota metabolic disturbances. HIRI-induced dysbiosis and intestinal barrier disruption significantly elevate soluble LPS levels in the intestinal lumen and portal vein, worsening liver injury (12).

Intestinal microbiota regulates HIRI

Gut microbiota-derived metabolites are key molecular mediators linking the gut microecology and HIRI. A set of core functional metabolites, such as SCFAs, BAs, γ-aminobutyric acid (GABA) and indole derivatives, together with LPS, participate in the regulation of liver immunity, metabolism, and oxidative balance (Figures 2,3, Table 2).

Figure 2 Regulation of inflammation in hepatic ischemia-reperfusion injury by intestinal microbiota metabolites (SCFAs, LPS, BAs). ASC, apoptosis-associated speck-like protein containing a CARD; BAs, bile acids; cAMP, cyclic adenosine monophosphate; FXR, farnesoid X receptor; GPR41, G protein-coupled receptor 41; GPR43, G protein-coupled receptor 43; IL, interleukin; LPS, lipopolysaccharides; MyD88, myeloid differentiation primary response gene 88; NF-κB, nuclear factor kappa B; NLRP3, NOD-like receptor protein 3; PKA, protein kinase A; SCFAs, short-chain fatty acids; SHP, small heterodimer partner; TGR5, Takeda G protein-coupled receptor 5; TLR4, toll-like receptor 4; TNF-α, tumor necrosis factor-alpha.

Figure 3 Regulation of oxidative stress in hepatic ischemia-reperfusion injury by intestinal microbiota metabolites (SCFAs, GABA, indole derivatives). ACSL4, acyl-CoA synthetase long-chain family member 4; AhR, aryl hydrocarbon receptor; AKT, protein kinase B; ARE, antioxidant response element; Foxp3, forkhead box P3; FTH1, ferritin heavy chain 1; GABA, γ-aminobutyric acid; HO-1, heme oxygenase-1; IL, interleukin; JAK, janus kinase; NQO1, NAD(P)H:quinone oxidoreductase 1; Nrf2, nuclear factor erythroid 2-related factor 2; PI3K, phosphatidylinositol 3-kinase; ROS, reactive oxygen species; SCFAs, short-chain fatty acids; SOD, superoxide dismutase; STAT3, signal transducer and activator of transcription 3; TFR1, transferrin receptor 1.

SCFAs

SCFAs, or volatile fatty acids, are primarily produced by anaerobic bacteria in the colon via the fermentation of undigested carbohydrates such as oligosaccharides, non-starch polysaccharides, and resistant starch. Acetate, propionate, and butyrate are the most common SCFAs. In HIRI, SCFAs mainly offer protection due to their anti-inflammatory and antioxidant characteristics.

SCFAs modulate inflammatory responses and exert anti-inflammatory effects by interacting with G protein-coupled receptors (GPCRs) and inhibiting histone deacetylase (HDAC) activity (21). Within the GPCRs pathway, SCFAs function as natural agonists for G protein-coupled receptor 41 (GPR41) and G protein-coupled receptor 43 (GPR43). Through the activation of pro-inflammatory signaling pathways, such as NF-κB, they inhibit the activation of the NOD-like receptor protein 3 (NLRP3) inflammasome. This inhibition consequently reduces the release of pro-inflammatory cytokines, including tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β), and interleukin-6 (IL-6), from hepatic Kupffer cells and infiltrating neutrophils (13). Consequently, this mechanism attenuates the ‘cascade-like’ amplification of the inflammatory response associated with HIRI. SCFAs, notably butyrate, enhance histone acetylation through HDAC inhibition, promoting anti-inflammatory gene expression like forkhead box P3 (Foxp3) and supporting regulatory T cell (Treg) expansion. These Tregs secrete interleukin-10 (IL-10), which suppresses effector T cell and macrophage hyperactivation, promotes an immune-tolerant microenvironment, and reduces HIRI-induced damage (22).

SCFAs, particularly butyrate, contribute to the liver’s defense against oxidative stress through various mechanisms. SCFAs initiate the nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway, leading to the transcription of antioxidant genes like heme oxygenase-1 (HO-1) and NAD(P)H:quinone oxidoreductase 1 (NQO1). These genes aid in eliminating reactive oxygen species (ROS) and reducing oxidative damage (23,37). Additionally, SCFAs enhance mitochondrial function by promoting mitochondrial biogenesis and maintaining functional integrity, thereby decreasing ROS production at its source and further bolstering the liver cells’ antioxidant capacity (24).

GABA

GABA, a four-carbon non-proteinogenic amino acid, is widely found in organisms and serves as the main inhibitory neurotransmitter in the mammalian central nervous system. Recent research has identified GABA as a significant microbial metabolite synthesized by gut microbiota via pathways such as glutamate decarboxylase (GAD). It plays various roles in immune regulation and cytoprotection within peripheral tissues. Wang et al. (25) were the first to demonstrate that metformin mitigates HIRI by modulating the gut microbiota to enhance GABA production, which subsequently inhibits ferroptosis. Exogenous GABA supplementation significantly improves HIRI by reducing hepatic iron accumulation and lipid peroxidation, while downregulating pro-ferroptotic proteins such as acyl-CoA synthetase long-chain family member 4 (ACSL4) and transferrin receptor 1 (TFR1), and upregulating anti-ferroptotic proteins like ferritin heavy chain 1 (FTH1) (26). This study introduces a novel ‘metformin-gut microbiota-GABA-ferroptosis inhibition’ axis in the context of HIRI alleviation, although the precise mechanisms underlying this pathway require further elucidation. This finding enhances comprehension of the gut-liver axis and microbial metabolite functions, while offering robust theoretical backing for HIRI prevention and treatment via microbiota modulation.

BAs

BAs represent a significant category of host-microbe co-metabolites. Primary BAs, including cholic acid and chenodeoxycholic acid, are initially synthesized in the liver through cholesterol oxidation and subsequently conjugate with taurine or glycine to form bile salts. Gut microbiota modify these bile salts, significantly contributing to cholesterol catabolism and systemic cholesterol homeostasis (38). BAs play a crucial role in the processes of nutrient and exogenous substance absorption, distribution, metabolism, and excretion. The mechanisms through which BAs perform their diverse functions are intricate, involving interactions between the host and gut microbiota (39). Notably, BAs play a vital protective role in HIRI. This protective function primarily operates via the activation of the farnesoid X receptor (FXR) and the Takeda G protein-coupled receptor 5 (TGR5) (27,40). Gut microbiota convert primary BAs into secondary BAs, which act as high-affinity ligands to activate FXR when they return to the liver. FXR activation is crucial for the liver’s anti-inflammatory defense. It enhances small heterodimer partner (SHP) expression, which directly disrupts and inhibits the transcriptional activity of the core pro-inflammatory factor NF-κB. This process notably inhibits the release of inflammatory mediators, including TNF-α and IL-1β, from liver immune cells like Kupffer cells, thus preventing ongoing inflammatory cycles (41). Simultaneously, FXR activation boosts hepatocyte survival factor expression and suppresses BA synthesis, preventing cytotoxic hydrophobic BA accumulation in damaged liver tissue and reducing secondary damage (28). Conversely, secondary BAs serve as potent agonists for the membrane receptor TGR5. Research by Sinha et al. (29) demonstrated that supplementation with secondary BAs can alleviate intestinal inflammation, with this anti-inflammatory effect partially attributed to the reduction of colitis levels in mice by lithocholic acid and deoxycholic acid via TGR5 activation. Activation of TGR5 in liver immune cells, like Kupffer cells, triggers the cyclic adenosine monophosphate-protein kinase A signaling pathway (30,42). This signaling pathway can directly inhibit the NLRP3 inflammasome’s assembly and activation, subsequently hindering the maturation and release of IL-1β and interleukin-18 (IL-18) downstream of NF-κB (30,31). In conclusion, the BA signaling pathway, mediated by the dual roles of the FXR within the nucleus and the TGR5 on the cell membrane, forms a comprehensive, three-dimensional defense network. This network integrates gene transcription and rapid intracellular signaling to collectively mitigate the primary injury mechanism of HIRI-sterile inflammation-thereby preserving the homeostasis of the hepatobiliary system.

Indole derivatives

Indole derivatives, metabolites generated by intestinal microorganisms through the catabolism of tryptophan (43), establish a complex signaling axis from the gut microbiota to the hepatic immune system, demonstrating distinct cytoprotective roles in mitigating HIRI. The signaling axis begins with metabolites like indole-3-carboxaldehyde (ICA) and 5-hydroxyindole-3-acetic acid (5HIAA), produced by particular microbial communities. These metabolites are absorbed through the intestinal epithelium into the portal circulation and then transported to the liver. The main process entails activating the aryl hydrocarbon receptor (AhR) on liver immune cells (33). As a ligand-activated transcription factor, AhR undergoes a conformational transformation upon binding to indole derivatives, facilitating its translocation into the nucleus. This process stimulates hepatocytes to synthesize endogenous antioxidant molecules, including regeneration gene proteins and metallothioneins, and induces the secretion of antimicrobial peptides from the Reg3 family. These peptides exhibit bactericidal effects against translocated bacteria and help maintain gut mucosal barrier integrity, significantly reducing the invasion of pathogen-associated molecular patterns (PAMPs) (33,44,45). The indole derivative-AhR-IL-22 axis effectively harnesses microbial metabolites as signaling molecules, facilitating communication to immune cells via the AhR and subsequently transmitting these signals to hepatic cells through interleukin-22 (IL-22). This mechanism facilitates an accurate and efficient signaling pathway from the gut to the liver, connecting microbial and host cell interactions, and offering a strong defense against the main cause of cellular damage, which is cell death.

LPS

LPS, a unique macromolecule found on the outer membrane of Gram-negative bacteria, comprises lipid A, which anchors in the membrane, and outward-extending polysaccharide chains. This structural composition not only serves a protective role for the bacterial cell but also facilitates the activation of the host’s immune response when LPS translocates from the gut, thereby playing significant roles in both physiological and pathophysiological processes (46). During the development of HIRI, prolonged obstruction of the portal vein results in intestinal congestion, which induces a state of low perfusion and ischemia-hypoxia in the intestinal mucosal epithelial tissue. Subsequent reperfusion injury exacerbates this condition by damaging the sensitive intestinal epithelial tissue and impairing its barrier function, thereby facilitating the translocation of commensal gut bacteria. This phenomenon, in conjunction with liver injury, compromises the liver’s sinusoidal filtration capacity, leading to bacteremia and endotoxemia, which further aggravate liver dysfunction and the disruption of the intestinal mucosal barrier. In the context of portal vein congestion, there is a marked elevation of gut-derived endotoxins, which results in the substantial binding of LPS with lipopolysaccharide-binding proteins (LBP) within the portal vein or systemic circulation. This interaction facilitates the formation of complexes that readily translocate to the liver. Subsequently, Kupffer cells recognize circulating endotoxin complexes via the pattern recognition receptor TLR4 located on their cell membrane surface, initiating multiple signaling cascades that contribute to the progression of HIRI. Initially, the binding of endotoxin to TLR4 on the Kupffer cell surface activates the MyD88 dependent signaling pathway, which subsequently leads to the downstream activation of the NF-κB pathway. This activation significantly enhances the synthesis and robust secretion of key pro-inflammatory cytokines, such as TNF-α, IL-1β, and IL-6, by Kupffer cells, while also inducing the release of chemokines like C-X-C motif chemokine ligand 1 (CXCL1) and C-X-C motif chemokine ligand 2 (CXCL2) (34). Consequently, this process preliminarily amplifies local hepatic inflammatory infiltration and tissue damage. Secondly, during hepatic ischemia-reperfusion, substantial quantities of DAMPs, such as adenosine triphosphate, mitochondrial DNA, high mobility group box 1 (HMGB1), and ROS, are produced. These DAMPs serve as secondary activation signals for the inflammatory cascade, potentially leading to the complete assembly and activation of the NLRP3 inflammasome (35). Once activated, the NLRP3 inflammasome specifically activates caspase-1, which subsequently cleaves the precursors of IL-1β and IL-18, synthesized under the influence of the early NF-κB of activated B cells pathway, into mature cytokines with significant pro-inflammatory properties (36). Concurrently, this process initiates pyroptosis, a form of inflammatory programmed cell death in Kupffer cells, thereby markedly exacerbating hepatic inflammatory injury. Furthermore, the release of HMGB1 can facilitate the sequential activation of apoptosis signal-regulating kinase 1 (ASK1) and the mitogen-activated protein kinase (MAPK) pathway, precisely modulating the expression imbalance of downstream antioxidant and stress-related proteins, such as HO-1 and SIRT3, thereby exacerbating hepatic oxidative stress disorders. Multiple signaling pathways interact in a layered and progressive manner, leading to sustained activation and functional remodeling of Kupffer cells. This process establishes a vicious cycle of inflammation, oxidative stress, and cell pyroptosis, ultimately mediating and aggravating the pathological progression of HIRI (47,48).

Research progress on targeted intervention of intestinal microbiota in HIRI

In recent years, strategies aimed at targeting gut microbiota have become a hot topic in HIRI prevention and treatment, gradually forming a multi-dimensional tech system that includes selective gut decontamination, probiotic/prebiotic interventions, fecal transplants, colonization with engineered synthetic biology bacteria, and targeted regulation of microbiota metabolism and signaling pathways (Table 3).

Antibiotics

In HIRI, compromised intestinal barriers allow gut microbiota-derived PAMPs like LPS to translocate to the liver, significantly contributing to inflammation. Consequently, antibiotics, which can swiftly and directly modulate the gut microbiota, offer promising strategies for intervening in HIRI. The primary approach involves selective digestive decontamination (SDD), which employs non-absorbable or poorly absorbable oral antibiotics, such as vancomycin, neomycin, and polymyxin B, to selectively target and eliminate specific pathogenic bacteria in the gut, particularly Gram-negative bacilli (79). This method could significantly lower LPS levels in the system, reducing the translocation of PAMPs to the liver during HIRI and intestinal barrier disruption. Consequently, it alleviates endoplasmic reticulum stress and enhances hepatic autophagy, ultimately diminishing liver transplant-related injury (49-51). Despite the promising prospects of this strategy, it necessitates careful implementation. Broad-spectrum antibiotics, although effective against pathogenic bacteria, also adversely affect beneficial symbiotic bacteria. Inappropriate utilization of these antibiotics can profoundly disrupt the composition of the gut microbiota, resulting in dysbiosis and compromising the integrity of the intestinal barrier. Moreover, the eradication of sensitive symbiotic bacteria generates ecological niches that facilitate the proliferation of resistant strains, thereby promoting the overcolonization of resistant bacteria and the widespread expression of resistance genes (52). This process, mediated through enterohepatic circulation, persistently influences the hepatic microenvironment, thereby elevating the risk of postoperative infections with multidrug-resistant bacteria and prolonging liver injury (53-55).

Probiotics and prebiotics

Probiotics, as beneficial microbial consortia, play a pivotal role in modulating gut microbiota equilibrium. Their application not only facilitates the colonization and functionality of beneficial bacteria but also effectively suppresses pathogenic bacterial proliferation, thereby preserving gut microecological balance. Recent studies have increasingly highlighted the critical role of probiotic therapy in modulating microbial dynamics and optimizing the intestinal milieu. For instance, Lactobacillus reuteri modifies the composition of the BA pool, with a notable impact on the metabolism of ursodeoxycholic acid. It also diminishes the prevalence of cholestasis-associated microbiota during acute LPS stimulation, thereby effectively mitigating liver inflammation and hepatocyte apoptosis (56). Clostridium butyricum mitigates HIRI by augmenting butyrate production, activating the NRF2 antioxidant pathway, and inhibiting NF-κB activation. Lactobacillus rhamnosus GG upregulates intestinal barrier proteins like ZO-1 and claudin-1, while decreasing serum LPS levels (57,58). Commercial probiotic strains may inherently possess or acquire antibiotic resistance genes via horizontal gene transfer. Upon entering the intestinal environment, these genes can be disseminated to commensal and opportunistic pathogenic bacteria through mechanisms such as conjugative plasmids and transposons. Following translocation through the gut-liver axis, this process may elevate the risk of antibiotic-resistant infections in the liver (59,60). Furthermore, probiotics are subject to adverse environmental conditions and colonization resistance, which can hinder the maintenance of a sufficient population of viable probiotics necessary to reach the target site and exert their intended effects (80).

Prebiotics, which are dietary components selectively utilized by gut microbiota (including oligosaccharides and cellulose), provide nutritional support for beneficial bacteria and enhance metabolic activity. Research has demonstrated that prebiotics, such as inulin, stimulate the growth of butyrate-producing bacteria and contribute to the restoration of the gut-liver metabolic balance (61). Prebiotic supplementation significantly boosts beneficial gut bacteria, enhances gut microbiota composition and function, and partially alleviates hepatic steatosis, potentially reducing ischemia-reperfusion injury (IRI) after liver transplantation.

Fecal microbiota transplantation (FMT) and engineered bacteria colonization

FMT transfers healthy donor fecal microbiota to a recipient to restore gut microecology. In recent years, clinical studies have explored the application of FMT to enhance the efficacy of treatments for diabetes, melanoma, fatty liver, and other diseases (62-65). However, these studies are often limited by small sample sizes. Research indicates that FMT exerts significant protective effects in animal models of HIRI; however, its clinical application in HIRI-related contexts, such as complications following liver transplantation, remains in the exploratory phase (66). The primary challenges include donor screening, standardization of preparation, safety concerns (such as the risk of pathogen transmission and unknown long-term effects), ethical and regulatory considerations, acceptance, and procedural complexity. Additionally, low-abundance antibiotic-resistant bacteria and mobile resistance genes present in the microbiota of healthy donors can be transferred to the recipient’s gut via FMT, where they may establish colonization and expansion (67). In the context of intestinal barrier damage induced by HIRI, bacterial translocation and the dissemination of resistance genes are more likely to occur, ultimately reaching the liver through the portal vein and triggering infections with resistant bacteria. Comprehensive quality control and extensive clinical trials are crucial to confirm the efficacy and safety of FMT for HIRI. Building on the seminal research by Funabashi and colleagues published in Nature (59), which elucidates the metabolic pathways of gut microbiota BAs, engineered bacterial colonization technology is emerging as a cutting-edge approach to modulate the gut-liver axis and address HIRI. This innovative technology facilitates the introduction of the complete BA 7α-dehydroxylation gene cluster into the non-pathogenic commensal bacterium Clostridium sporogenes through a multi-plasmid system. This approach ensures stable colonization and the execution of predefined metabolic functions within the complex intestinal milieu, representing a transition from traditional probiotic transplantation to the precision of synthetic biology. But, genetically engineered bacteria present risks associated with plasmid escape and abnormal gene integration. Once colonized, these bacteria may undergo genetic recombination with indigenous gut strains, potentially leading to the emergence of new antibiotic-resistant strains. Their prolonged retention in the enterohepatic circulation may perpetuate selective pressure, thereby exacerbating the dissemination and evolution of resistance along the gut-liver axis.

Targeting key molecular pathways

Recent insights into the gut-liver axis underscore the influence of gut microbiota and its metabolites as dynamic, modifiable units affecting HIRI progression by targeting key liver molecular pathways. Within HIRI, the TLR4/NLRP3 pathway triggers the NLRP3 inflammasome, leading to Caspase-1 activation. This activation leads to the maturation and release of pro-inflammatory factors, such as IL-1β, which directly induce hepatocyte pyroptosis (68). Additionally, the TNF-α signaling pathway facilitates the recruitment of neutrophils to the liver, activating mitochondrial apoptosis pathways and interacting with the pyroptosis pathway, thereby collectively exacerbating liver injury. The MAPK signaling pathway, triggered by cytokines, enhances pro-inflammatory mediator expression, thereby intensifying inflammation in HIRI (69). Wang et al. (70) utilized network pharmacology analysis to demonstrate that metabolites from the gut microbiota mitigate HIRI by inhibiting cell apoptosis through multiple signaling pathways. In addition, Pan et al. (71) Animal experiments demonstrated that intraperitoneal administration of N-acetyl-L-tryptophan (L-NAT) in mice mitigates HIRI. The exact molecular mechanisms responsible for L-NAT’s hepatoprotective effects have yet to be completely understood. Chen et al. (72) have engineered a nanozyme, Cu5.4O@CNDs, exhibiting cascade antioxidant activity. This innovative material emulates the functions of superoxide dismutase and catalase, efficiently neutralizes ROS, and demonstrates significant therapeutic efficacy in a murine HIRI model by modulating hepatic retinol metabolism and inflammatory pathways. In a separate study, Ma et al. (73) developed Kupffer cell membrane-mimicking nanoparticles, K-EVs@ZIF-8@siRNA, designed for the targeted delivery of BRCC3 siRNA to hepatic Kupffer cells. This targeted approach effectively suppresses the BRCC3/NLRP1 signaling pathway, thereby mitigating sepsis-induced liver injury. Luo et al. (74) showed that sodium butyrate can reduce the inflammatory response through the GPR43/NF-κB signaling pathway, significantly alleviating LPS-induced liver injury. The opioid agonist remifentanil has been shown to exert substantial hepatoprotective effects during HIRI. Yang et al. (75) reported that pretreatment with remifentanil can suppress TLR4 expression and attenuate inflammatory responses associated with HIRI; however, this effect is not observed in TLR4 knockout mice. Chen et al. (76) found that both protein and mRNA levels of FXR and glutathione peroxidase 4 (GPX4) are significantly reduced following HIRI. They were the first to propose a potential co-regulatory relationship between FXR and GPX4, suggesting their joint involvement in the ferroptosis process during HIRI. FXR potentially regulates GPX4 transcription, increasing its expression to inhibit ferroptosis during HIRI, thereby reducing liver injury. This study uncovers a novel intrinsic signaling axis for combating HIRI and establishes a direct connection between nuclear receptor signaling pathways and ferroptosis, thereby advancing the understanding of the pathological mechanisms underlying HIRI. FXR agonists like obeticholic acid are clinically used to treat primary biliary cholangitis and have a well-established safety profile. The FXR/GPX4 pathway presents a novel therapeutic strategy for addressing HIRI.

Furthermore, molecularly targeted interventions pose risks of secondary resistance and adverse imbalances. Resistance to drugs targeting specific molecular pathways typically emerges due to mutations in the target, activation of alternative signaling pathways, and disruptions in receptor transport and degradation (77,78). Focusing on a single bacterial resistance pathway can inadvertently activate compensatory mechanisms, enabling bacteria to develop resistance through target mutations, alterations in membrane structure, and other mechanisms. Concurrently, imbalanced interventions may exacerbate hepatic inflammation, thereby compromising the gut barrier and indirectly facilitating the translocation of resistant bacteria and dissemination of resistance genes within the liver. Therefore, targeted therapies for HIRI require further comprehensive investigation. Future research may offer novel theoretical frameworks and identify potential drug targets for modulating gut microecology or its metabolites in the treatment of HIRI.

Conclusions

A complex bidirectional regulatory relationship exists between HIRI and the gut microbiota. The gut-liver axis functions as a communication bridge between the gut and liver, playing a pivotal role in the regulation of host physiology and the progression of HIRI. HIRI can induce an imbalance in the gut microbiota by compromising the integrity of the gut barrier and altering the gut environment. Conversely, dysbiotic microbiota and their metabolic products can exacerbate liver inflammation, oxidative stress, and cellular damage by activating hepatic innate immune receptors such as TLR4, modulating inflammatory responses, and influencing immune cell function and metabolic homeostasis, thereby creating a vicious cycle. It is important to note that the majority of research on the interactions and molecular pathways between HIRI and the gut microbiota is derived from animal studies. Although some clinical data indicate a clear association between disturbances in the gut microbiota, key inflammatory factors and pathway molecules, and the severity of liver injury in HIRI patients, there remains a paucity of systematic studies addressing the complex clinical scenarios in humans. In contrast to the controlled conditions of standardized animal experiments, patients undergoing liver transplantation or hepatic surgery demonstrate considerable variability in dietary habits, individual gut microbiome composition, pre-existing comorbidities (including hepatobiliary diseases, metabolic disorders, and chronic inflammation), and the administration of postoperative immunosuppressive therapies. The mechanisms through which these human-specific variables affect the gut microbiome-signaling pathway axis in the context of HIRI remain inadequately understood. Consequently, the extrapolation of current basic research findings to human clinical pathology is challenging, as these findings are difficult to replicate precisely or apply directly in a clinical setting. Despite the current challenges in this field, including notable individual variability, an incomplete comprehension of underlying mechanisms, and a lack of sufficient clinical evidence, the advancing understanding of gut-liver axis interactions, along with progress in microbiome technologies and the development of innovative therapies such as optimized FMT protocols and engineered bacteria, suggests that targeted intervention of the gut microbiota may become an effective strategy for the prevention and treatment of HIRI. This approach holds promise for enhancing the clinical outcomes of patients undergoing liver surgery and presents extensive translational prospects for future research.

Acknowledgments

None.

Footnote

Peer Review File: Available at https://tgh.amegroups.com/article/view/10.21037/tgh-2026-0095/prf

Funding: This study was supported by the Natural Science Foundation of Hubei Province (No. 2024AFB1027).

Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://tgh.amegroups.com/article/view/10.21037/tgh-2026-0095/coif). All authors report that this study was supported by grant 2024AFB1027 from the Natural Science Foundation of Hubei Province. The authors have no other conflicts of interest to declare.

Ethical Statement: The authors are accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved.

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