Dysbiosis is characterized not only by changes in the qualitative and quantitative aspects of the microbiota but also by shifts in the production of various metabolites by the bacteria. Dysbiosis can lead to increased intestinal permeability, loss of epithelial integrity, and weakened mucus-associated defense. As a result, viable bacteria, microbial products, and host–bacteria interactions can influence normal physiology and disease susceptibility. These influences can occur locally, signaling to different cell populations of the intestinal mucosa and distant organs, including the liver. In the liver, bacterial products can directly affect hepatocytes or cells of the immune system, such as Kupffer cells or stellate cells. Recognition of pathogen-associated molecular patterns (PAMPs) and damage-associated molecular patterns (DAMPs) through TLRs induces pro-inflammatory signals and may also affect apoptosis. Thus, gut-derived factors and alterations in microbial input can influence hepatic inflammation and injury during liver disease. Advancements in metabolomics and metagenomics have helped shed light on the mechanisms through which dysbiosis and altered metabolic output in the gut affect liver diseases. Increased intestinal permeability allows bacterial products and metabolites to cross the epithelial barrier and reach the liver through the portal vein, potentially triggering an inflammatory response. Examples of bacterial metabolites that have been implicated in disease development include ethanol (produced by the intestinal microbiome) in obesity and non-alcoholic fatty liver disease (NAFLD), short-chain fatty acids derived from dietary fibers, secondary bile acids (BA), trimethylamine from dietary choline during NAFLD, and acetaldehyde during alcoholic liver disease ^{[1][2][3][4][5][6][7][8][9][10]}[86,87,88,89,90,91,92,93,94,95]. The role of the gut microbiota in influencing health and disease is becoming more recognized. Studies have shown that the human gut microbiota plays a significant role in host metabolism. This understanding stems from initial observations that germ-free (GF) mice have lower levels of body fat, a characteristic that can be reversed when these mice are colonized with a normal gut microbiota ^[11][96]. Emerging evidence from both preclinical and clinical research indicates that the gut microbiota plays a significant role in the development of NAFLD. This involvement primarily occurs through its contribution to obesity, metabolic changes such as insulin resistance, and the promotion of liver inflammation. Additionally, certain bacterial byproducts, including ethanol, can exhibit hepatotoxic effects by stimulating Kupffer cells to produce and release nitric acid and cytokines ^[12][97]. Multiple pre-clinical and clinical studies have highlighted the key role of gut microbiota in NAFLD pathogenesis through its impact on obesity, metabolic alterations, and liver inflammation. The composition of the gut microbiota, including the abundance of specific microorganisms such as Akkermansia muciniphila, may have implications for the development and progression of NAFLD. Several studies conducted over a decade ago demonstrated the significant influence of the gut microbiota on weight gain and obesity. Germ-free mice were found to gain 42% less weight than mice with gut microbiota, even when consuming more calories. Transplanting the cecal microbiota from normal mice into germ-free mice resulted in a 57% increase in weight gain. Furthermore, germ-free mice could not gain weight even on a high-calorie diet. In another study, gut microbiota transplanted from obese mice led to greater fat gain in recipient mice compared to microbiota from lean mice. This suggested that obesity-associated gut microbiota extracted more energy from the diet by digesting indigestible polysaccharides into SCFAs. Similar findings were observed in human studies, with obese individuals having higher concentrations of short-chain fatty acids in their feces compared to lean individuals. Specific gut microbiota composition has been associated with obesity and subsequent NAFLD development. Obese mice had lower levels of Bacteroidetes and higher levels of Firmicutes and Archaea compared to lean controls. Similar alterations in the Bacteroidetes/Firmicutes ratio were observed in obese humans. Enterotype 1, characterized by the consumption of animal proteins and saturated fat, was associated with obesity, while Enterotype 2 was associated with a diet high in carbohydrates. However, the recently discovered bacterium A. muciniphila has been associated with a non-obese phenotype. Low fecal concentrations of A. muciniphila were found in pregnant women who gained excess weight during pregnancy and in obese and overweight preschool children. Conversely, A. muciniphila counts were increased in obese mice that underwent gastric bypass surgery. Experimental models have shown that A. muciniphila modulates weight gain, type 2 diabetes, and NAFLD. Administration of A. muciniphila in high-fat diet-fed mice induced Treg cells in adipose tissue, reducing inflammation and improving glucose tolerance. Similar improvements in insulin resistance, adipose tissue inflammation, fat gain, and endotoxemia were observed in mice with type 2 diabetes mellitus following A. muciniphila administration. Overall, these studies highlight the vital role of the gut microbiota in NAFLD pathogenesis through its impact on obesity, metabolic alterations, and liver inflammation. The composition of the gut microbiota, including the abundance of specific microorganisms such as A. muciniphila, may have implications for the development and progression of NAFLD ^{[11][13][14][15]}[96,98,99,100]. Endogenous ethanol is a byproduct of certain gut microbiota species and is absorbed into the bloodstream and transported to the liver through the portal vein. In the liver, alcohol dehydrogenase enzymes metabolize ethanol, resulting in the formation of acetate and acetaldehyde. Acetate can contribute to fatty acid synthesis, while acetaldehyde produces reactive oxygen species, leading to oxidative stress. This process contributes to the accumulation of triglycerides in the liver and fulfills both steps of the “two hits” hypothesis for NAFLD development. Elevated levels of ethanol have been found in obese patients and even in non-alcohol-consuming children with non-alcoholic steato-hepatitis (NASH), suggesting its role in the development of NAFLD/NASH. Moreover, studies have shown increased expression of alcohol-metabolizing enzymes such as alcohol dehydrogenase, catalase, and aldehyde dehydrogenase in NASH livers. Ethanol may also promote NAFLD by increasing the permeability of the gut mucosa, which can lead to endotoxemia, a condition characterized by the presence of endotoxins in the bloodstream. In summary, endogenous ethanol produced by the gut microbiota can contribute to NAFLD development through its metabolism in the liver, triglyceride accumulation, oxidative stress, and promotion of gut mucosal permeability and endotoxemia ^[9][16][17][94,101,102]. Endotoxin is a component of the cell membrane of Gram-negative bacteria. The active endotoxin component is called lipopolysaccharide, which binds to the LPS-binding protein and CD14 receptor to form a complex. This complex interacts with Toll-like receptors (TLRs) and triggers an inflammatory cascade. Genetically obese mice develop steatohepatitis (inflammation and fat accumulation in the liver) when low doses of LPS are infused into their bodies. In NAFLD mice, injection of LPS further promotes liver injury by enhancing the production of proinflammatory cytokines. A high-fat diet also leads to increased circulating LPS in rodents with diet-induced NAFLD. Human studies have shown that individuals with NAFLD have significantly higher circulating endotoxin levels than healthy controls. These elevated endotoxin levels are particularly pronounced in the preliminary stages of fibrosis. Activation of Toll-like receptors leads to the translocation of NF-κB (a transcription factor) into the nucleus, resulting in the transcription of proinflammatory genes such as TNF-α, IL-1β, IL-6, and IL-12. IL-1β promotes the accumulation of triglycerides in hepatocytes by enhancing the activity of diacylglycerol transferase, an enzyme that converts diglycerides into triglycerides. On the other hand, TNF-α inhibits insulin receptors and insulin receptor substrate-1, leading to increased levels of circulating insulin and insulin resistance. This insulin resistance facilitates the entry of fatty acids derived from adipose tissue into the liver. In summary, endotoxin (LPS) derived from Gram-negative bacteria activates Toll-like receptors and triggers an inflammatory response in NAFLD. This inflammation is associated with the production of proinflammatory cytokines and the promotion of triglyceride accumulation in hepatocytes. Additionally, the activation of proinflammatory genes and the development of insulin resistance contribute to the progression of NAFLD ^{[18][19][20][21][22][23][24]}[103,104,105,106,107,108,109]. Choline, an essential component of cell membranes, plays a vital role in lipid transport from the liver. The gut microbiota regulates choline metabolism by producing enzymes that convert choline into methylamines. When the liver absorbs these methylamines, they have the potential to induce inflammation. Studies using a mouse model of high-fat diet-induced steatosis have shown that there is a decrease in circulating phosphatidylcholine (a form of choline) and an increase in the excretion of choline metabolites in urine. These findings support the presence of a gut microbiota phenotype that leads to choline deficiency and contributes to liver injury. Choline deficiency contributes to the accumulation of triglycerides in the liver and a decrease in the liver’s secretion of very-low-density lipoprotein (VLDL). Animal studies have demonstrated that a choline-deficient diet can result in liver steatosis, which is reversible upon choline supplementation. It is important to note that the data regarding choline deficiency and its association with NAFLD development are derived from animal models and choline-deficient conditions, which may not fully represent the complexity of NAFLD in humans. However, decreased choline levels and increased levels of toxic choline metabolites may represent a potential mechanism through which gut microbiota-mediated choline deficiency contributes to the development of NAFLD ^{[9][25][26][27][28]}[94,110,111,112,113].

Gut microbiotas play significant roles in vital functions such as maintaining homeostasis, preventing pathogen colonization, producing vitamins, and maintaining a mature immune system. Gut microbiota disruption is highly related to environmental factors such as sex, diet, antibiotic use, and medications. Mitochondria is the main source of energy in the human body and plays a significant role in intestinal homeostasis. Mitochondria is a sensitive organelle that responds to environmental alterations and energy requirements. According to endosymbiosis theory, mitochondria originate from the fusion of archaebacteria and rickettsia alpha-proteobacterium. Mitochondria generate energy via oxidative phosphorylation (OXPHOS). Mitochondria that reside in the gut provide a hypoxic environment that allows obligate anaerobes to remain in the gut to maintain their homeostasis. SCFAs give energy to the epithelial cells of the colon, which affects the metabolism of mitochondria ^[29][85]. BA can influence mitochondrial metabolism. Bifidobacterium and Bacteroides are the main gut microbiota. They modify conjugated BA into secondary BA. Secondary BAs control transcription factors that act on carbohydrate and lipid metabolism, which are regulated by the mitochondria ^[30][114]. Moreover, peroxisome proliferator-activated receptor-gamma coactivator (PGC)-1alpha upregulation increases oxidative phosphorylation activity. For example, butyrate, a short-chain fatty acid, is metabolized by the colon to produce NADH, which participates in the process of OXPHOS. Furthermore, butyrate could additionally up-regulate uncoupling protein 2 (UCP2) of the mitochondria, leading to decreased production of ROS ^[30][31][114,115]. Moreover, mitochondria and gut microbiome are considered highly dynamic functionally related entities with variations between individuals and within the individual body. It has become increasingly evident that the composition and activity of the gut microbiota profoundly affect human physiology, including immune function and inflammation ^[32][116]. Pro-inflammatory cytokines play a pivotal role in initiating and amplifying the inflammatory cascade. They are involved in the recruitment and activation of immune cells, vascular permeability regulation, and tissue damage induction ^[33][117]. Emerging evidence suggests that the gut microbiota influences the production and release of proinflammatory cytokines, thereby impacting the immune response and contributing to the development and progression of inflammatory diseases ^[34][118]. One of the mechanisms through which dysbiosis influences liver inflammation involves increased intestinal permeability and the subsequent translocation of microbial products, such as lipopolysaccharides, into the liver ^[35][30]. Hepatic Kupffer cells interact with lipopolysaccharides via Toll-like receptor 4 (TLR-4) and activate intracellular signaling pathways, leading to the release of pro-inflammatory cytokines such as tumor necrosis factor-alpha (TNF-α), interleukin-1beta (IL-1β), and IL-6 ^[36][119].

Furthermore, the metabolic activities of the gut microbiota can affect pro-inflammatory cytokine production. The gut microbiota ferments dietary fibers and complex carbohydrates, producing SCFAs as byproducts. SCFAs, such as acetate, propionate, and butyrate, have immunomodulatory effects and can influence the production of pro-inflammatory cytokines ^[37][120]. For instance, butyrate has been shown to inhibit the production of pro-inflammatory cytokines while promoting the release of anti-inflammatory cytokines, contributing to maintaining immune homeostasis. The dysbiosis-induced alterations in the gut microbiota composition and function can disrupt these regulatory mechanisms, leading to an imbalanced production of pro-inflammatory cytokines and chronic inflammation ^[38][39][121,122].

Understanding the interactions between the gut microbiota and pro-inflammatory cytokines is important for unraveling the mechanisms underlying immune dysregulation and inflammation-associated diseases. Targeting the gut microbiota through interventions such as probiotics, prebiotics, and fecal microbiota transplantation (FMT) holds promise as a therapeutic approach to modulate pro-inflammatory cytokine production and mitigate inflammation-driven pathologies ^[40][123].