Clinical and biological significance of the relationship between gut microbiota and liver disease

Abstract

The gut microbiota is of growing interest to clinicians and researchers due to its elucidating extensive role in metabolic and immune mechanisms, not only in the gut but also in other organs. The liver shares a close bidirectional relationship with the intestine and the gut microbiota. Disturbances in the composition of the gut microbiota can affect the immune systems of both the intestine and liver. In turn, bile composition also influences the gut microbiota. Disruption of this balance can arise from various causes and may significantly impact intestinal and liver health. Therefore, the aim of the current review is to discuss the biological relationships between the gut microbiota and liver function as well as the clinical significance of their disturbances.

Key Words: Gut microbiota; Liver disease; Immunometabolism; Nonalcoholic fatty liver disease; Bile; Innate immune system; Short-chain fatty acids

Core Tip: The gut microbiota has numerous metabolic and immune connections not only to the gut itself but also to other organs, including the lungs, brain, and liver. The gut-liver connection is bidirectional, and disruptions in this relationship contribute to the development of various liver diseases. Modulation of gut microbiota composition and function forms the basis of therapeutic strategies for conditions such as non-alcoholic fatty liver disease, cirrhosis, and hepatocellular carcinoma.

INTRODUCTION

The gut microbiota is of growing interest to clinicians and researchers due to its extensive role in many physiological and pathophysiological processes. There is increasing evidence that the gut microbiota functions as a complex, regulated, multicellular “organ” involved in maintaining the body’s metabolic and immune homeostasis[1,2]. It is believed that the number of microorganisms inhabiting the intestine may even exceed the number of cells in the human body, and that the total number of genes possessed by gut microorganisms may greatly surpass the number of human genes[3,4]. The gut microbiota is composed of bacteria, archaea, fungi, and viruses, which are species-specific and have a complex composition influenced by various internal and external factors[5-7].

Growing evidence suggests that the gut microbiota is an integral part of the intestine’s complex interactions with other organs. Indeed, the gastrointestinal tract is one of the most evolutionarily ancient systems in the body, and its role in regulating the functions of other organs and systems is well established. In recent years, clinicians and researchers have focused on the “gut-lung”, “gut-brain”, and “gut-liver” axes, with the gut microbiota serving as a key component of these connections (Figure 1).

Figure 1 Gut microbiota and liver connections. The gut microbiota can have both positive and negative effects on liver function. The gut microbiota has direct and indirect effects on liver function and is involved in the regulation of metabolism and immunity through metabolite production, bile acid metabolism, eating behavior, and regulation of inflammation. The artwork used in this figure was adapted from Servier Medical Art (http://smart.servier.com/) (Supplementary material).

There is compelling evidence that the liver has a bidirectional relationship with the gut microbiota, and that disruptions in this relationship are involved in the development of liver disease[8,9]. Liver diseases remain a significant issue in modern medicine. They are widespread and contribute to disability and premature mortality[10-13]. A major concern is that patients are often unaware of the presence of liver disease, may not recognize the symptoms as serious, and therefore delay seeking medical attention[14]. Non-alcoholic fatty liver disease (NAFLD), for example, is very common but frequently underdiagnosed. These patients often have metabolic risk factors such as obesity, insulin resistance, or diabetes mellitus, and atherosclerotic cardiovascular disease[15-17]. Because hepatic symptoms are often absent or mild in the early stages, patients may focus on other conditions with more overt manifestations. Diagnostic challenges, including the need for specialized equipment and trained personnel, further complicate the situation. In this context, investigating the mechanisms underlying liver disease and identifying new diagnostic and therapeutic approaches remains a pressing clinical priority. Therefore, the current review aims to discuss the biological links between gut microbiota and liver function, as well as the clinical significance of their impairment.

STRUCTURE AND FUNCTION OF THE GUT MICROBIOTA

Composition and core characteristics of the gut microbiota

The gut microbiota is a complex ecosystem of microorganisms inhabiting the human digestive tract[7]. It is mainly composed of bacteria, with more than 1500 species belonging to over 50 different phyla. However, 99% of these bacteria are represented by approximately 30-40 species. The most common bacterial phyla in the human gut are Bacteroidetes and Firmicutes, followed by Proteobacteria and Actinobacteria[18-21]. In addition to bacteria, the gut microbiota also includes archaea, viruses (phages), and fungi, all of which contribute to its complexity and functionality[22,23].

The composition of the human gut microbiota changes significantly throughout life due to various factors such as diet, environment, and aging. During the fetal period, the gut is sterile but is rapidly colonized with microorganisms after birth. The gut microbiota undergoes substantial changes during the first two years of life and generally stabilizes by around three years of age. This period is critical for the development of a diverse microbial community[6,24,25]. The gut microbiota has species-specific characteristics and is strongly influenced by dietary patterns. For example, studies have shown that captivity and artificial feeding can significantly alter the gut microbiota of Sichuan golden monkeys, often resulting in an increase in Bacteroidetes and a decrease in Firmicutes, Actinobacteria, Verrucomicrobia, and Tenericutes[26]. Another study found that captive primates tend to lose their natural gut microbiota and are colonized predominantly by Prevotella and Bacteroides, species commonly found in the human gut[27]. Dietary components such as fiber, amino acids, organic acids, and polyphenolic compounds can significantly affect the gut microbiota’s composition. Excessive intake of carbohydrates and fats is a key factor influencing microbiota balance. Obese individuals often have a higher Firmicutes/Bacteroidetes ratio, which tends to decrease with weight loss, suggesting its potential role in obesity[28-30]. However, some studies have reported conflicting results, indicating that other factors, such as lifestyle and genetic predisposition, may also influence the relationship between the Firmicutes/Bacteroidetes ratio and obesity[31,32]. Alterations in bacterial ratios are also characteristic of various diseases. For instance, an elevated Firmicutes/Bacteroidetes ratio is associated with ulcerative colitis. This imbalance may contribute to the onset and progression of the disease, highlighting its relevance in inflammatory responses[33,34].

Archaea are another important component of the gut microbiota. The most common archaea in the human gastrointestinal tract are Methanobacteriales and Methanomassiliicoccales[35,36]. Methanogenic archaea, such as Methanobrevibacter, participate in methanogenesis by converting hydrogen and carbon dioxide into methane. This process is essential for maintaining microbial community balance by removing excess hydrogen, which can interfere with other microbial activities[37,38]. The methane produced by archaea also serves several physiological functions. For example, it is associated with slowed intestinal transit, particularly in the ileum and colon. Methane reduces the rate of peristalsis and increases the amplitude of contractions in the ileum, leading to slower transit[39-41]. Changes in the ratio of methanogenic archaea may be linked to certain diseases, making this a promising area of research. For example, the presence of methane in exhaled air has been associated with NAFLD, suggesting that elevated methane levels in exhaled air could serve as a potential biomarker for NAFLD diagnosis[42].

Fungi make up a relatively small proportion of the human gut microbiota, constituting approximately 0.03% of the fecal microbiota. They are composed mainly of three groups: Ascomycetes, Basidiomycetes, and Zygomycetes[35,43]. The role of intestinal fungi in liver disease development remains largely unclear. However, altered fungal microbiota composition has been reported in patients with NAFLD. In these patients, the relative abundance of Talaromyces, Paraphaeosphaeria, Lycoperdon, Curvularia, Phialemoniopsis, Paraboeremia, Sarcinomyces, Cladophialophora, and Sordaria, as well as Leptosphaeria, Pseudopithomyces, and Fusicolla, was increased[44]. Intestinal fungi also produce various bioactive substances. For instance, the contribution of prostaglandin E2 produced by fungi to the development of alcoholic hepatic steatosis has been described[35,45]. Patients with hepatocellular carcinoma (HCC) exhibit significant alterations in the gut mycobiome, including reduced fungal diversity and increased presence of pathogenic fungi such as Candida albicans and Malassezia spp. These changes are associated with worsened clinical parameters and increased tumor size[46-48]. Malassezia species residing in tumors may contribute to HCC development by inhibiting bile acid synthesis and modulating the tumor microenvironment[46].

In recent years, interest in the virome, the community of intestinal viruses, has increased. The intestinal virome consists of eukaryotic viruses capable of replicating in human cells and bacteriophages, which replicate in intestinal bacteria and are the most abundant viral entities in the gut[49]. It is estimated that the human intestine contains more than 10¹² viruses, a number that may be comparable to that of intestinal bacteria[50]. The community of intestinal bacteriophages, known as the gut “phageome”, is of particular clinical interest. Bacteriophages play a central role in the gut microbiota by infecting and transforming local intestinal bacteria, and they also interact directly with the human immune system. For example, intestinal viruses can stimulate immune responses via toll-like receptor (TLR) 3. Additionally, viruses can influence the human body indirectly by affecting bacterial communities that produce various metabolites and signaling molecules. Most commonly, bacteriophages replicate within bacterial host cells and lyse them upon release. However, some phages can also influence bacterial physiology by altering gene expression, modulating bacterial gene transcription, or transferring new genetic material[51].

The clinical significance of viromes remains largely unknown. However, there is growing evidence of a link between intestinal viruses and metabolic diseases such as type 1 diabetes mellitus and various intestinal disorders[52-54]. Liver disease has also been associated with intestinal viruses. In patients with more severe forms of NAFLD, a decreased diversity of intestinal viruses and a lower proportion of bacteriophages compared to other intestinal viruses have been observed[55]. Changes in the composition of intestinal viruses have also been reported in patients with alcoholic hepatitis. These alterations were associated with disease severity and mortality. Specifically, increased levels of Escherichia-, Enterobacteria-, and Enterococcus-specific phages, as well as significant increases in viruses such as Parvoviridae and Herpesviridae, were found in fecal samples from patients with alcoholic hepatitis[56]. Given these findings, bacteriophages are being explored as promising therapeutic agents for several diseases through their ability to modulate the composition of the gut microbiota[57].

Thus, despite the growing interest in this topic and the emerging body of research, a comprehensive understanding of the virome’s role in various diseases remains incomplete. Nevertheless, it is becoming clear that, in addition to bacterial translocation of the gut microbiota to the liver in NAFLD, viruses may also translocate and contribute to liver pathology.

Thus, the gut microbiota has a complex composition that is maintained through various mechanisms involving both the microorganisms themselves and the human immune system. Interestingly, gut bacteria engage in various “social” interactions known as quorum sensing (QS). QS is a bacterial communication mechanism that enables the coordination of group behavior based on population density. In the gut microbiota, QS plays a crucial role in maintaining microbial balance, combating pathogens, and facilitating interactions with the host organism[58-60]. QS has been observed in both pathogenic and commensal microorganisms, highlighting the universality of this mechanism. Furthermore, QS may influence the spatial distribution of bacteria throughout the gastrointestinal tract[58]. This system is based on the release of small signaling molecules (autoinducers) by bacteria, which accumulate in the environment in proportion to bacterial density and are detected by the bacterial community. Gram-negative bacteria primarily produce species-specific acyl-homoserine lactones for this purpose, while Gram-positive bacteria mainly use autoinducing peptides. A shared signaling molecule, autoinducer-2, also known as furanosyl borate diester or tetrahydroxy furan, is used by both Gram-negative and Gram-positive bacteria[59,61].

The macroorganism also plays a vital role in maintaining the proper composition of the gut microbiota through mechanisms such as mucus production, immune tolerance, and other regulatory processes. For example, the mucus that covers the intestinal wall promotes colonization by commensal microbiota. In turn, the microbiome can influence mucus production, both by directly activating various signaling cascades and through bioactive factors produced by epithelial cells[62,63]. Mucin 2 oligosaccharides provide numerous attachment sites for microbes and also serve as an energy source[63]. Another important mechanism by which the intestinal epithelium regulates immune responses is the secretion of cytokines and chemokines[64,65]. This secretion helps maintain a balanced interaction between intestinal microbes and the host immune system. Additionally, immune mechanisms involving intestinal epithelial cells include the production of antimicrobial peptides, which help control microbial populations in the gut, preventing infections and maintaining mucosal integrity[66,67]. Thus, over the course of a long evolutionary history of coexistence, the microbiota and the macroorganism have developed numerous mechanisms to support this mutually beneficial relationship - one that is essential for many physiological functions.

Functions of the gut microbiota

The most well-known function of the gut microbiota is its involvement in digestion. By fermenting non-digestible fiber, the gut microbiota enables the extraction of additional energy from food. This function is particularly prominent in herbivorous animals but is also important in humans, as it allows for the metabolization of certain carbohydrates that are not digested by human enzymes. The gut microbiota also has many other functions that are beyond the scope of this review, including the synthesis of essential vitamins such as vitamin K and several B vitamins, which play important roles in various physiological processes[68,69]. For example, decreased levels of vitamin B12 in obese mice may be due to reduced synthesis by the gut microbiota[70]. In addition, the gut microbiota is involved in the metabolism of xenobiotics and drugs, further highlighting its systemic importance[71].

Production of short-chain fatty acids

The enzymatic activity of the gut microbiota results in the production of short-chain fatty acids (SCFAs), which are important immune and metabolic regulators with effects that extend far beyond the gut. The primary substrates for SCFA formation are dietary fibers, including resistant starch, cellulose, and pectin[72]. SCFAs produced by the gut microbiota serve as energy substrates for colonocytes and are also absorbed by the intestinal epithelium into the bloodstream. After absorption via the portal vein, SCFAs first enter the liver and then circulate in the peripheral bloodstream, forming part of the “axes” that link the intestine to other organs. Acetate, propionate, and butyrate are the major SCFAs produced by the gut microbiota. Their production primarily involves Bacteroidetes and Firmicutes, which are mainly localized in the proximal colon[73-75].

Once in the systemic circulation, SCFAs exert their effects through G-protein-coupled receptors such as GPR43 and GPR41, also known as free fatty acid (FFA) receptors FFA2 and FFA3, respectively. They also act via the GPR109a receptor (also known as hydroxycarboxylic acid receptor 2 or HCA2) and olfactory receptor 78[76-81]. In addition, SCFAs mediate their effects by inhibiting histone deacetylases. For example, butyrate can promote a metabolic shift in macrophages toward an anti-inflammatory M2 phenotype by inhibiting histone deacetylase 3[82,83]. SCFAs are known to play a critical role in regulating immune responses and maintaining immune homeostasis[84,85]. They can reduce inflammation, repair the intestinal barrier, and promote the proliferation of certain immune cells, such as regulatory T cells (Tregs)[86]. In particular, butyrate supports the differentiation of Tregs, which are essential for maintaining immune tolerance and preventing autoimmune responses. It increases the expression of FoxP3, a key transcription factor for Tregs, and suppresses the differentiation of pro-inflammatory T helper 17 cells by decreasing the expression of retinoic acid-related orphan receptor γt and interleukin (IL)-17[87]. Furthermore, SCFAs influence the metabolic status of T cells, thereby affecting epigenetic modifications and T cell function[88].

SCFAs, especially butyrate and propionate, reduce the production of proinflammatory cytokines by dendritic cells. This modulation influences the activation and function of cluster of differentiation 8 T cells, which play a critical role in the adaptive immune response[89]. SCFAs also affect the localization and function of immune cells such as natural killer cells in the large intestine and regulate intestinal leukocyte activity through SCFA-specific receptors[90]. In macrophages, SCFAs modulate function by regulating the production of inflammatory mediators and enhancing phagocytic activity, thereby promoting pathogen elimination. Additionally, they increase the production of reactive oxygen species, which are essential for effective pathogen destruction[91].

Thus, SCFAs exert multiple biological effects on various organs and systems, including the liver, which is the first organ to receive SCFAs from the intestine via the portal vein. SCFAs have both direct and indirect effects on hepatic metabolic and immune functions. Directly, they exert immunomodulatory actions and influence liver cell metabolism. For instance, SCFAs have been shown to reduce hepatocyte apoptosis induced by uremic toxins bound to gut-derived proteins[92]. Through activation of FFAR3, SCFAs may enhance the metabolic functions of the liver[93]. Butyrate, in particular, offers protection against insulin resistance and fatty liver dystrophy by modulating mitochondrial function, efficiency, and dynamics in insulin-resistant obese mice. Specifically, it improves respiratory capacity and fatty acid oxidation, activates the AMP-activated protein kinase-acetyl-CoA carboxylase pathway, and promotes inefficient metabolism[94]. In a study involving ApoE^-/^- mice, butyrate treatment significantly slowed the progression of atherosclerosis and hepatic steatosis induced by a high-fat diet. Butyrate achieved this by regulating the expression of genes involved in lipid and glucose metabolism[95]. Furthermore, butyrate has been shown to increase the activity of ATP-binding cassette transporter A1, a key protein in reverse cholesterol transport, which facilitates the export of cholesterol from macrophages to form high-density lipoprotein[95]. SCFAs also stimulate the hepatic expression of ApoA-I, a major component of high-density lipoprotein particles, enabling cholesterol efflux and supporting the removal of excess cholesterol from peripheral tissues[96]. SCFAs may additionally exert effects through epigenetic mechanisms by modulating DNA methylation and histone acetylation of genes associated with NAFLD, thereby influencing lipid metabolism and immune homeostasis[97].

The indirect effects of SCFAs on the liver include their role in maintaining the integrity of the intestinal barrier through several mechanisms. For example, butyrate can increase the expression of tight junction proteins such as claudin-1 and zonula occludens-1 in epithelial cells by activating protein kinase B/mammalian target of rapamycin-mediated protein synthesis[98]. Additionally, SCFAs stimulate the secretion of the incretin hormone glucagon-like peptide-1 in mixed colon cultures in vitro[99]. In this way, SCFAs contribute to hepatic energy balance by regulating appetite and mediating systemic glucose homeostasis[100].

Disruptions in SCFA production are associated with various inflammatory conditions, including autoimmune liver diseases such as primary biliary cholangitis, primary sclerosing cholangitis, and autoimmune hepatitis[85]. Because SCFAs possess anti-inflammatory properties, they may help reduce hepatic inflammation by modulating immune responses and lowering the production of proinflammatory cytokines[98,100]. Thus, SCFAs are essential metabolic and immune regulators in the body, and the gut microbiota plays a central role in this regulation.

LINKS BETWEEN MICROBIOTA AND LIVER FUNCTION

Involvement in human nutritional behavior

Metabolic disorders play a key role in the initiation and progression of liver diseases such as non-alcoholic steatohepatitis (NASH). Excessive intake of digestible carbohydrates and trans fats, along with reduced intake of non-digestible fiber, are well-known risk factors for NAFLD across many populations. As a result, eating disorders are a pressing concern.

It has been suggested that the gut microbiota may influence human behavior, including eating behavior. The gut microbiota interacts with the “gut-brain axis”, a bidirectional communication system between the gut and the brain mediated by hormonal, immune, and neural signals. Through this interaction, the gut microbiota can influence eating behavior as well as digestive and absorptive processes (e.g., by regulating intestinal motility and barrier integrity)[101]. Experimental studies have shown that the microbiota can influence host eating behavior through several potential mechanisms. These include microbial effects on brain reward centers, the production of mood-altering toxins, alterations in taste receptors, and interference with neural signaling via the vagus nerve - the primary neural link between the gut and brain[102]. The microbiota also produces various neurochemicals such as dopamine and serotonin, which affect both mood and behavior[103]. It is estimated that nearly half of the body’s dopamine is produced in the gut[102,104].

Studies in germ-free mice have shown an increased preference for fat and higher caloric intake from fat[105]. This was associated with enhanced sensitivity of taste receptors to fat and a marked reduction in the expression of intestinal satiety peptides and fatty acid receptors[105]. In addition, the absence of gut microbiota in mice altered the expression of sweet taste receptors and glucose transporters in the proximal small intestine, correlating with increased consumption of energy-rich sweet solutions[106].

By participating in energy extraction from the diet, the gut microbiota may contribute to energy storage in the host[107]. In obesity, the gut microbiota shows an enhanced capacity to extract energy from food and may pass this trait to the host. For example, colonization of germ-free mice with microbiota from obese mice led to a significantly greater increase in total body fat than colonization with microbiota from lean mice[108]. Thus, alterations in the composition of the gut microbiota may contribute to obesity and, consequently, to the development of metabolic risk factors for liver disease.

Bile formation and metabolism

Cholesterol is a critical metabolite essential for various cellular functions. One of its key roles is in reverse cholesterol transport, the process by which excess cholesterol is removed from macrophages and transported to the liver for utilization. In the liver, hepatocytes convert cholesterol into primary bile acids, which are then secreted into the gallbladder as part of bile. The primary function of bile acids is to emulsify and aid in the absorption of fats in the small intestine. Bile acids undergo enterohepatic recirculation, a highly efficient process in which approximately 95% of bile acids are reabsorbed. This occurs through both passive absorption of conjugated and unconjugated bile acids in the small intestine and colon, and active transport in the distal ileum. Once absorbed, bile acids enter the portal bloodstream, are taken up by hepatocytes, and re-secreted into bile (Figure 2). A small portion of bile acids may enter the systemic circulation, allowing them to exert effects on other organs and tissues[109-111].

Figure 2 Metabolism and functions of bile acids. The artwork used in this figure was adapted from Servier Medical Art (http://smart.servier.com/) (Supplementary material). CYP7A1: Cholesterol 7α-hydroxylase; CYP8B1: Sterol 12α-hydroxylase; CYP27A1: Sterol 27-hydroxylase.

The composition of bile acids has a significant impact on the intestinal microbiota, contributing to the bidirectional communication of the “gut-liver axis”. Once primary bile acids enter the gastrointestinal tract, they are chemically transformed into secondary bile acids by the gut microbiota. These transformations involve oxidation, dehydroxylation, and conjugation with amino acids, which greatly expand the diversity of the bile acid pool[112,113]. The gut microbiota produces several enzymes essential for bile acid metabolism, including bile salt hydrolase, 7α-dehydroxylase, and hydroxysteroid dehydrogenase. These enzymes facilitate deconjugation and dehydrogenation reactions, converting primary bile acids into secondary forms[114,115].

Secondary bile acids act as signaling molecules, influencing host metabolism and shaping the composition of the gut microbiota[112,113,116]. Bile acids also possess antimicrobial properties, which affect the growth and survival of various bacterial strains. They may accumulate within bacterial cells and disrupt metabolic functions, thereby limiting bacterial proliferation[113,117,118]. For example, deoxycholic acid can significantly inhibit bacterial growth by interfering with ribosomal transcription and amino acid metabolism[118]. Thus, the gut microbiota can significantly influence bile acid metabolism. Moreover, the gut microbiota not only transforms bile acids but may also regulate bile acid synthesis in the liver by modulating key enzymes such as cholesterol 7α-hydroxylase, sterol 12α-hydroxylase, and sterol 27-hydroxylase[119].

Interestingly, bile itself harbors a complex microbiota. In humans, its composition includes bacteria from eight different phyla, including Actinobacteria, Bacteroidetes, Firmicutes, Fusobacteria, Gemmatimonadetes, Proteobacteria, Saccharibacteria, and Tenericutes. This composition is often disrupted in various biliary tract diseases. For example, patients with primary sclerosing cholangitis exhibit a decreased abundance of Actinobacteria, Bacteroidetes, Firmicutes, and Fusobacteria, along with an increased presence of Proteobacteria, compared to healthy controls[120].

Abnormal profiles of circulating bile acids are early indicators of the development and progression of metabolic and neurodegenerative diseases, including obesity, type 2 diabetes mellitus, HCC, and Alzheimer’s disease[121]. An increase in total bile acid concentration is frequently accompanied by a shift toward a higher proportion of taurine-conjugated bile acids, which has been observed as an early metabolic alteration in the development of HCC[122]. Some studies suggest that secondary bile acid levels may serve as useful biomarkers for diagnosing and predicting the progression of liver diseases[123]. For instance, patients with NAFLD and mild fibrosis (stage F1) show significantly higher levels of secondary bile acids than those without fibrosis[124]. Another study found elevated levels of cholic acid, deoxycholic acid, chenodeoxycholic acid, ursodeoxycholic acid, and lithocholic acid in the fecal bile acid fraction of NAFLD patients compared to healthy controls. Additionally, the total serum bile acid concentration was higher in individuals with NAFLD and advanced fibrosis than in the control group[125]. A recent study also demonstrated a clear association between altered gut microbiota and changes in the primary composition of conjugated bile acids in patients with NASH without cirrhosis[126]. It has been suggested that dysregulation of bile acid homeostasis, and signaling via the farnesoid X receptor (FXR) and fibroblast growth factor 19, mediated by the gut microbiota may contribute to fibrogenesis, liver injury, and oncogenesis in NASH[126]. Given these clinical findings, bile acid metabolism, and the role of the gut microbiota in regulating it, are both areas of growing interest. Thus, the gut microbiota emerges as a key player in the gut-liver axis, influencing both metabolic and immune mechanisms.

CLINICAL SIGNIFICANCE OF THE GUT MICROBIOTA IN LIVER DISEASE

Role of gut microbiota in NAFLD

Key mechanisms in the pathogenesis of NAFLD include insulin resistance, which is a hallmark of the condition. Insulin resistance leads to increased lipolysis in adipose tissue, elevated plasma FFA levels, and promotes fat accumulation in the liver. The buildup of triglycerides and FFAs in hepatocytes causes lipotoxicity, contributing to liver inflammation and fibrosis. Chronic inflammation and oxidative stress are critical factors in the progression from simple steatosis to NASH. These pathological processes are driven by mitochondrial dysfunction, endoplasmic reticulum stress, and the release of pro-inflammatory cytokines. Both nutritional factors and the composition of the gut microbiota influence NAFLD development. A high-fat diet and gut dysbiosis can exacerbate hepatic fat accumulation and inflammation[127-129].

The gut microbiota plays an important role in the development and progression of NAFLD. It affects intestinal permeability, leading to the entry of bacterial endotoxins, such as lipopolysaccharides (LPSs), into the liver via the portal vein. This process causes hepatic inflammation and contributes to the progression of NAFLD[130-132]. LPSs trigger an inflammatory response in the liver by activating TLR4 in hepatocytes and Kupffer cells (liver macrophages). This activation leads to the release of pro-inflammatory cytokines such as tumor necrosis factor alpha, IL-1β, and IL-6, which contribute to hepatic inflammation and the progression of NASH[133-135]. Chronic exposure to LPSs can lead to hepatic fibrosis through the activation of hepatic stellate cells via the TLR4 signaling pathway. This fibrogenic response is an important step in the transition from simple steatosis to more serious liver diseases such as cirrhosis[135]. Given that the gut microbiota influences energy metabolism, insulin sensitivity, and lipid metabolism, dysbiosis may disrupt these processes, leading to increased fat accumulation in the liver[136,137].

As noted previously, the gut microbiota converts primary bile acids into secondary bile acids that interact with receptors such as FXR and the G protein-coupled receptor Takeda G protein-coupled receptor 5. These interactions influence lipid metabolism and glucose homeostasis, contributing to the progression of NAFLD[130,138]. FXR activation reduces fat absorption and thereby decreases lipid accumulation in the liver, while Takeda G protein-coupled receptor 5 activation promotes the release of glucagon-like peptide-1 into the bloodstream[130]. Microbial metabolites such as SCFAs, trimethylamine-N-oxide, and indole play important roles in regulating inflammation, lipid metabolism, and insulin sensitivity, thereby influencing the development of NASH[139,140]. In addition, some gut bacteria produce endogenous ethanol, which may exacerbate liver damage and inflammation, contributing to the progression of NAFLD[130,140-142].

Therapeutic approaches in the treatment of NAFLD that target the gut microbiota include the use of probiotics and prebiotics. Probiotics containing Lactobacillus plantarum and Bifidobacterium bifidum have been shown to be effective in regulating gut microbiota, reducing lipid accumulation in the liver, and improving insulin sensitivity[143]. Prebiotics such as inulin and fructooligosaccharides also support the growth of beneficial bacteria in the gut[144-146]. A combination of probiotics and prebiotics (synbiotics) may further improve gut health and potentially alleviate symptoms of NAFLD[136]. Fecal microbiota transplantation (FMT) is an intensively studied method aimed at restoring a healthy gut microbiota composition and has shown promising results in preliminary studies[130,137]. By improving intestinal barrier function, FMT reduces the translocation of harmful bacterial components into the liver, thereby lowering inflammation. FMT has also been shown to improve metabolic parameters such as insulin sensitivity, lipid metabolism, and bile acid profiles, which are critical in the treatment of NAFLD[137,147]. An emerging and interesting potential treatment for liver disease, particularly NAFLD, is the use of bacteriophages to modulate the gut microbiota. High-alcohol-producing Klebsiella pneumoniae, may be one of the contributing factors to NAFLD[148]. In an experimental mouse model of Klebsiella pneumoniae-induced NAFLD, phage therapy targeting the gut microbiota was shown to be a potentially safe and effective alternative to antibiotics[149].

Thus, abnormalities in the composition and function of the gut microbiota play an important role in NAFLD progression. Therefore, therapeutic strategies targeting the gut microbiota are of growing interest in the treatment of NAFLD.

Role of gut microbiota in liver cirrhosis

The gut microbiota plays an important role in the pathogenesis and progression of liver cirrhosis through the gut-liver axis. In cirrhosis, a compromised intestinal barrier allows bacteria and their products to enter the liver, triggering immune responses and inflammation that exacerbate liver damage[150-152]. The gut microflora is the main source of LPSs in the portal vein. An imbalance in the composition of the gut microbiota is associated with cirrhosis and can lead to complications such as hepatic encephalopathy, spontaneous bacterial peritonitis, and visceral artery vasodilation[150,151,153]. Dysbacteriosis and bacterial translocation are involved in the development of hepatic encephalopathy, a serious complication of cirrhosis[150,151,154]. This condition arises from the accumulation of harmful microbial byproducts, such as ammonia, indoles, oxyindoles, and endotoxins, which build up due to dysbiosis and the liver’s reduced capacity to eliminate them[155,156]. Moreover, patients with cirrhosis and hepatic encephalopathy often present with a specific gut microbiota profile characterized by an increase in pathogenic bacteria such as Enterobacteriaceae, which are associated with elevated plasma ammonia levels and cognitive impairment[157]. Dysbiosis of the gut microbiota may also increase the risk of portal hypertension by promoting inflammation and immune responses[152].

Modulation of the gut microbiota with antibiotics, probiotics, prebiotics, and synbiotics has been shown to be effective in the treatment of cirrhosis and its complications[150,151,153]. Therapeutic approaches targeting the gut microbiota in cirrhosis include the use of antibiotics to treat bacteria-related complications by reducing bacterial load and translocation[151]. Probiotics and synbiotics help restore balance to the gut microbiota, improve gut barrier function, and reduce inflammation, offering a potential therapeutic approach for the management of cirrhosis[150,151,153].

Thus, disturbances in the gut microbiota play an important role in liver cirrhosis, and therapeutic strategies aimed at correcting these imbalances are of significant clinical interest.

The role of gut microbiota in HCC

HCC is a complex and multifactorial disease with a poor prognosis and a high mortality rate. The pathogenesis of HCC involves a combination of genetic, molecular, and environmental factors[158,159]. HCC often develops against a background of chronic liver diseases such as cirrhosis, hepatitis B and C, alcoholic liver disease, and NAFLD[160,161]. Chronic inflammation and hepatic fibrosis are key contributors to HCC development. Persistent oxidative stress and long-term inflammation cause DNA damage, further promoting HCC progression[161,162]. The tumor microenvironment, including immune cells, hepatic stellate cells, and an abnormal extracellular matrix, also plays a crucial role in the progression of HCC.

The gut microbiota is involved in the development and progression of HCC through various mechanisms[163-165]. It influences HCC via the gut-liver axis, whereby microbial metabolites and components impact liver function and immune responses[166,167]. An imbalance in the gut microbiota can promote chronic liver inflammation, fibrosis, and cirrhosis, which are known risk factors for HCC. This inflammation is often driven by microbial products such as LPSs and other genotoxins. Persistent microbiota-induced inflammation may lead to genetic and epigenetic changes in hepatocytes, thereby facilitating carcinogenesis[163,168-170].

The gut microbiota may also contribute to HCC through the modification of bile acids[171,172]. Taurocholic acid has been shown to promote the polarization of tumor-associated macrophages toward an immunosuppressive M2 phenotype. In addition, ω-muricholic acid reduces the accumulation of natural killer T cells in the tumor, thereby promoting tumor growth. Deoxycholic acid has been found to induce senescence in hepatic stellate cells, resulting in increased production of chemokines such as IL-8 and transforming growth factor-beta. These factors may drive epithelial-to-mesenchymal transition and contribute to HCC development, including increased metastatic potential and aggressiveness of HCC cells[173-176].

It should be noted that the microbiota can modulate the immune response, which has important implications for the microbiota-macroorganism relationship. The gut immune system faces a complex task: On the one hand, it must carry out immune surveillance and eliminate pathogens; on the other hand, it must maintain immune tolerance toward the commensal microbiota. It has been shown that, in NAFLD-associated HCC, the gut microbiota can influence the peripheral immune response by promoting an immunosuppressive phenotype, which is associated with worse clinical outcomes[164].

The gut microbiota is also associated with the intratumoral microbiota, which has a distinct profile and plays an important role in HCC progression. It may contribute by inducing DNA damage, mediating tumor-associated signaling pathways, altering the tumor microenvironment, promoting metastasis, or through other mechanisms[177]. The intratumoral microbiota in HCC is highly diverse and includes various bacterial taxa such as Enterobacteriaceae, Fusobacterium, and Neisseria, which are more prevalent in HCC tissues compared to adjacent non-tumor tissues[178,179]. The intratumoral microbiota can influence the immunosuppressive microenvironment by affecting immune cell infiltration and function. For example, in HBV-associated HCC, the presence of specific intratumoral microbiota is associated with an increase in tumor-infiltrating CD8+ T lymphocytes and myeloid-derived suppressor cells, both of which contribute to immune suppression[179,180]. Microbial communities in HCC tissues may also alter metabolic pathways, including the upregulation of fatty acid and lipid synthesis, potentially contributing to tumor progression[178,181]. Moreover, bacteria found within HCC tumors may exert both pro-tumor and anti-tumor effects, depending on their species and context.

Changes in the composition of the gut microbiota may serve as a biomarker for the early diagnosis of HCC and for predicting the efficacy of immunotherapy[167,182,183]. Given the close relationship between the gut and liver, modulation of the gut microbiota may influence HCC outcomes by improving liver function, reducing inflammation, and inhibiting tumor growth[166,167,184]. The development of therapies that specifically target the intratumoral microbiota may enhance treatment efficacy and open new avenues for HCC management[175,185]. Various strategies to modulate the gut microbiota are currently being explored, including dietary interventions, antibiotics, probiotics, prebiotics, and FMT[166,167,170,184]. Modulation of the gut microbiota may also improve the effectiveness of immune checkpoint inhibitors used in HCC treatment by enhancing immune responses[168,186].

Thus, a growing body of evidence is advancing our understanding of the complex interactions between the gut microbiota and HCC, highlighting new possibilities for diagnosis, prognosis, and therapeutic intervention.

CONCLUSION

The gut microbiota is of growing interest to clinicians and researchers across various fields of medicine. It is closely associated with multiple organs and systems of the body, including the liver. The relationship between the liver and the gut microbiota is bidirectional and involves numerous complex interactions. The gut microbiota contributes to maintaining energy homeostasis, metabolic regulation, and immune function in the liver. Disruptions in these interactions contribute to the development and progression of liver diseases. In recent years, interest in assessing gut microbiota biodiversity has increased significantly. There is abundant evidence that factors such as aging, nutrition, smoking, obesity, and drug intake can severely affect gut microbiota diversity and, consequently, its function[187-190]. Conversely, alterations in the gut microbiota may contribute to drug resistance in HCC through mechanisms such as metabolite production, gene transfer, and immune modulation[191]. In this context, therapeutic strategies targeting the gut microbiota are gaining attention (Table 1). These include the use of probiotics, prebiotics, and FMT.

Table 1 Therapeutic strategies targeting the gut microbiota.

Therapeutic strategy	Contents	Clinical benefit
Antibiotics	Prophylactic and therapeutic use of antibiotics in cirrhosis	Treatment of bacterial infections in cirrhosis, including SBP
Probiotics	Useful microbiota	Reducing gut microbiota permeability (“leaky gut”)
		Reduction of systemic inflammation
		Correction of intestinal dysbiosis
		Effect on metabolism
Prebiotics	Inulin, fructooligosaccharides, galacto-oligosaccharides, oligosaccharides derived from starch and glucose, pectin oligosaccharides, non-carbohydrate oligosaccharides	Prebiotics stimulate the growth and activity of their own beneficial microflora	Stimulation of the growth of beneficial microbiota
			Production of SCFAs
			Suppression of pathogens
			Reduction of ammonia levels (in cirrhosis)
Synbiotics	Combinations of probiotics and prebiotics	Synbiotic supplements may improve liver function, regulate lipid metabolism, and reduce the degree of liver fibrosis
Fecal microbiota transplantation	Fecal microbiota from a healthy donor is injected into the gastrointestinal tract of another patient	Clinical studies have shown the effectiveness of the procedure in infectious and non-infectious liver diseases

SBP: Spontaneous bacterial peritonitis; SCFAs: Short-chain fatty acids.

Thus, the connection between gut microbiota and liver disease is of clear clinical interest. Promising directions for future research include deeper exploration of the immune and metabolic mechanisms linking the gut microbiota and the liver, as well as evaluating the roles of other gut microbial community members, such as archaea and fungi. Identifying new biomarkers and therapeutic approaches also remains a key priority. Experimental research is underway in several areas, including genetic engineering of intestinal bacteria and the use of bacteriophages[148,192,193]. However, the clinical applicability of these methods is still not fully established.