Gut-Hepatic Relationship: From Disorders of the Gut Microbiota to Hepatocellular Carcinoma

Enterohepatic relationships reflect the integrity of homeostasis regulation in the human body. Anatomical and physiological connection through the portal vein of the intestine and liver ensures the transport of products derived from the intestine directly to the liver and through the liver ensures the connection of bile and the secretion of antibodies into the intestine. However, impairment of intestinal barrier integrity, endotoxemia, increase of lipopolysaccharides in hepatocytes, Kupffer and stellate cells with increasing content of pro-inflammatory cytokines and active oxygen forms creates inflammatory environment with sensitization of liver cells to injury and profibrotic processes. The goals of experimental and clinical research in this course are multifaceted, ranging from regulating human metabolism, immune and inflammatory reactions to preventing carcinogenesis, inhibition of liver cancer progression and improving the efficiency of liver cancer treatment. Here we will discuss epidemiological issues of hepatocellular carcinoma, the role of intestinal microbiota when enterohepatic relationships are impaired, and the intestinal-associated mechanisms of carcinogenesis in this form of liver cancer.


Introduction
One of the leading causes of death worldwide is hepatocellular carcinoma. Research in recent years shows the important role of enterohepatic relationships in the pathophysiological mechanism responsible for the development and progression of HCC. The intestinal microflora is a positive factor of human homeostasis and immune reactions due to precise control and immunosensory ability to distinguish between commensal and pathogenic bacteria.

Hepatocellular Carcinoma: Epidemiology Issues
Hepatocellular carcinoma (HCC) is the most common primary carcinoma of the liver. Liver cancer is the sixth most frequently diagnosed and the fourth in the mortality rate due to cancer in the world after lung cancer, colorectal cancer, and stomach cancer [1]. HCC is one of many types of tumors that occur against the background of chronic inflammation. HCC, which accounts for 90% of all primary liver cancers [2,3].
According to Singal [1]. Liver cancer is a tumor with high mortality, with most cases being detected at late stages and the morbidity-to-mortality ratio approaching 1. It is important to note that HCC is the result of many etiological factors, such as viral infections of hepatitis B and C, non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH), alcoholic liver disease (ALD), autoimmune hepatitis and several genetic disorders. Hepatitis B virus (HBV) is the main cause of liver cancer and mortality in the world (33%), followed by alcohol (30%), hepatitis C virus (HCV) (21%) and other causes (16%) [1]. The main risk factors for HCC, including infection with hepatitis B virus (HBV) or hepatitis C virus (HCV), lead to the formation and progression of liver cirrhosis (LC), which is observed in 80-90% of patients with HCC. The five-year cumulative risk of HCC progression in patients with liver cancer varies from 5 to 30% [2]. With the introduction of vaccination programs against hepatitis B and treatment of hepatitis C worldwide, the epidemiology of HCC shifts from a disease in which viral hepatitis prevails to the hepatic component of metabolic syndrome -NAFLD [3], where about 10-30% of cases of NAFLD progress to liver cancer. More than 65 million Americans are affected by NAFLD, which costs 103 billion dollars a year in the United States itself [1]. There are data on the influence of lipid reduction strategy, including ezetimibe, and its combination with other hypolipidemic agents on atherogenic dyslipidemia, glycemic profile, and morphological changes in liver steatosis [5].
The progression of HCC in NASH occurs in the absence of LC [6] in more than 25%, which is significantly higher than the frequency observed in other liver diseases [7]. HCC has a strong prevalence of men in morbidity and mortality, with a ratio of men to women exceeding 2.5 for both epidemiological indicators [8].
It is believed that this differential distribution by gender is due to the clustering of risk factors among men, as well as the potential impact of androgens on the risk of HCC. There are also essential environmental risk factors for HCC. For example, in some parts of Africa and Asia, an important factor for the progression of HCC is the consumption of Aflatoxin B1 with food, which occurs due to fungal contamination of staple foods. The impact of Aflatoxin B1 is closely related to the mutations of TR53 (codon 249) and the progression of HCC in people infected with HBV [9]. Several epidemiological studies have identified an increased risk of HCC progression among smokers, with meta-analysis reporting an adjusted OR 1.5 (95% of CI 1.37-1.67) compared to non-smokers [10]. Study of HCC prevention in the general population and in patients with chronic liver disease, coffee consumption, aspirin intake and metformin treatment consistently reduce the incidence of HCC in diabetics [11][12][13].
which products are transported to the liver through the portal vein and affect liver function [14,15]. An important factor in liver injury is disturbance of the gut microbiota (GM) system. The human gastrointestinal tract (GI tract) colonizes a large number of microbial microflora, forms a system of metabolism of the microbiota with the macroorganism, participating in various metabolic processes of the body, and plays an essential role in the human immune response. More than 2,000 different types of bacteria live in the intestines, and their number exceeds 100 trillion microorganisms, which is 10 times more than the total number of human cells [16]. Gut microbiome, which refers to the collective genomes of all microorganisms that make up the gut microflora, contains 150 times more genes than the human genome [17]. The number of different microorganisms is gradually increased during the intestine. This can be explained by the presence of a more aggressive environment in the upper parts of the intestine due to the incoming acidic content of the stomach, the action of digestive enzymes, rapid progression of chyme. The density of microbes increases from proximal to distal end of the intestine and includes a biomass of 1.5-2.0 kg, which is dominated by strictly anaerobic bacteria [18]. Aerobia predominating in the small intestine, as they progress down the GI tract, are replaced by facultative and then strict anaerobes. These microorganisms are collectively referred to as gut microbiota (IM), which consists of commensals, beneficial bacteria, and opportunistic pathogenic bacteria and pathogenic bacteria in a complex and dense microenvironment [19,20].
The GI tract, which functions as an effective barrier against endotoxin and intestinal bacteria, can protect the body [21].
In addition, the liver also plays an essential physiological role  and increased of endotoxin levels [42].
The antibiotic treatment led to a reduction of tumor number and size compared with mice that did not receive antibiotics.
Moreover, mice that were grown in specific germ-free conditions demonstrated fewer and smaller tumors in this model compared with mice that were grown under non-germ free (SPF) conditions [42]. Human HCC is often associated with chronic inflammation of the liver and liver cirrhosis, pathophysiological processes that are the result of chronic viral infection, metabolic disorders or exposure to chemical toxins that can develop against the background of inflammatory environment in patients with advanced liver disease.
Experimental data from rodent models as well as robust clinical data suggest a role of inflammation in the pathophysiology of HCC development. The liver is constantly exposed to microbial products from the enteric microflora, such as endotoxin, which activates Inflammasoms, as a multi-protein oligomeric complex containing leucine and nucleotide binding domains, are responsible for the activation of the inflammatory response, controlling the splitting of pro-inflammatory cytokines. It was shown that dysbiosis leads to increased TNF-a expression [46]. The increased activation and production of TLR4 and pro-inflammatory cytokines in dysbiosis may also lead to the recruitment and activation of hepatic immune cells, and the pro-inflammatory transmission of TLR4 signals is involved in sensitizing cells to the profibrotic signaling pathways and thus leads to liver fibrosis [43] contributing to the progression of liver disease [46]. Kupffer cells (KC), as hepatic immunological cells, are critical components of the innate immune system located in the sinusoidal vascular space [47].

KC can be activated by a variety of endogenous and exogenous
stimuli, including endotoxins in the impairment of GM [47]. KC activation triggers the production of inflammatory cytokines such as TNF-α as well as active oxygen forms [47], which may also lead to cholesterol, and fat-soluble vitamins [49]. In addition, bile acids also function as signal molecules that affect physiological processes [49], which include regulation of glucose and lipid metabolism through activation of the farnesoid X-receptor (FXR) and binding to the G-protein of the bile acid receptor [50][51][52]. Bile acids may also affect the gut microbiota, as they are directly related to the integrity of the intestinal mucosa and the synthesis of antibacterial peptides [53]. When bile acids bind to FXR, antimicrobial peptides such as angiogenin 1 are produced. These peptides can inhibit excess gut microbiota growth by increasing the potential of intestinal epithelial cells to prevent bacterial uptake, improving intestinal barrier function [53]. Gut microbiota in turn may affect the size and composition of the bile acid complex by converting primary bile acids to secondary ones [54,55]. This may subsequently change the metabolism of lipids and glucose, especially in NAFLD-predisposed people [54,55]. Another mechanism through which gut microbiota can contribute to liver disease is the production of short-chain fatty acids (SCFAs).

Gut microbiota splits non-digestible carbohydrates, releasing
SCFAs in the human intestine [56]. The main SCFAs are acetate, propionate, and butyrate, which are metabolized by muscles, liver, and epithelium as such [56]. Studies on the role of SCFAs mainly focus on butyrate, the main energy source for colonocytes, which improves the barrier function of the large intestine [56] and therefore has a positive effect on intestinal permeability. Butyrate has been shown to improve intestinal barrier by induction of dense compound proteins and mucin mucin type 2 [57][58][59] and enhanced expression of claudin-1 [60]. Butyrate can induce apoptosis in the liver and inhibit cell proliferation in hepatocytes, suppressing the expression of type 1 sirtuin, while increasing the expression of miR-22, as a tumor suppressor [61]. In other words, butyrate can inhibit liver cancer cells. It has also been shown that butyrate increases the feeling of satiety, reduces food intake, and delays gastric emptying

American Journal of Biomedical Science & Research
Copy@ Ilkham Murkamilov by activating free fatty acid receptors type 2 and 3 [62]. Normal body weight and glucose homeostasis were more often found in mice with a deficit of free fatty acid receptors of type 2 and 3.

Stimulation of intestinal hormones and inhibition of food ingestion
by butyrate and propionate may represent a new mechanism by which gut microbiota regulates host metabolism. Finally, butyrate can also affect inflammation. Studies have shown that butyrate in the intestinal tract binds and activates the gamma receptor activated by the proliferator peroxis (PPAR-y), which counteracts the transduction of nuclear factor-kappa B (NF-kB), thus causing an anti-inflammatory effect [63]. Therefore, the presence or excess of gut microbiota-produced butyrate can affect the pathogenesis of liver diseases through several mechanisms [63].
Choline deficiency also plays the most important role in liver injury with gut micriobiota imbalance. Choline is an essential nutrient and phospholipid component of the cell membrane [64].
There are several mechanisms through which choline deficiency can affect the liver, including [64] the reduction of very lowdensity lipoprotein formation (VLDL), dysfunction, mitochondrion, and endoplasmic reticulum stress [64,65]. Phosphatidylcholine, which is a phospholipid, is a key component of the VLDL shell.
Choline deficiency caused by a diet or gut microbiota metabolism disorder leads to a decrease formation of VLDL and the export of triglycerides from the liver, resulting in fatty hepatosis. Choline deficiency reduces the concentration of phosphatidyl ethanolamine and phosphatidylcholine in mitochondrial membrane, leads to a decrease in membrane potential, which in turn causes oxidative damage [64] of cell membrane. Gut microbiota can help reduce the bioavailability of choline [66] contained in eggs, milk, and red meat.
This in turn increases the conversion of choline into trimethylamine (TMA) [67], which is absorbed into the blood, increasing the risk of cardiovascular disease [67]. TMA reaching the liver is further metabolized by flavin-containing monoxygenases of types 1 and 3 to form trimethylamine-N-oxide (TMAO) [67][68][69]. This can lead to an increase in the accumulation of hepatic triglycerides, as TMAO inhibits key enzymes and limits the enterohepatic circulation of bile acids [70][71][72]. Among men and women, 33% (11% -54%) and 18% (3% -38%) of the total number of HCC was due to past and present alcohol consumption [74]. The causal link between alcohol consumption and HCC development may be due to direct (hepotoxic) and indirect factors (cirrhosis development) [75]. Obesity and high fat diet have been identified as major risk factors for HCC [91,92]. In a prospective study of more than 900,000 adult American patients (404,576 males and 49,547 females) it was found that overweight and obesity have been accompanied by a significant increase in esophagus cancer, colorectal cancer, liver cancer, gall bladder cancer, pancreatic and kidney cancer mortality rates. For liver carcinoma, the odds ratio was 4.52 for men and 1.68 for women [93]. Yoshimoto S and colleagues demonstrated that administration of antibiotics and gut sterilization lead to a significant decrease in HCC development in an obesity related model of hepatocarcinogenesis in mice [36].