**3.1 Intestinal dysbiosis**

*Human Microbiome*

and currently.

**3. The gut microbiota**

which remain for life [13]**.**

tissue shows the presence of perilobular inflammation, hepatocellular ballooning, Mallory's hyaline, and acidophil bodies with or without fibrosis. Although there are other noninvasive tests, such as the fatty liver index, NAFLD fibrosis score, and FibroMeter, and elastographic techniques, such as FibroScan, that can suggest the presence of NASH and detect fibrosis [15], a precise histological diagnosis of NASH is commonly based on liver biopsy [26]. The development of alternative noninvasive strategies has been an area of intensive research over the past decade

Regarding NAFLD therapeutics, all forms of treatment of metabolic disorders

Millions of symbiotic microorganisms live on and within human beings and play an important role in human health and disease. Initial colonization occurs at the time of birth, and humans progressively acquire ∼1014 bacterial cells at equilibrium,

The human microbiota, especially the GM, has even been considered to be an "essential organ," carrying approximately 150 times more genes than the human genome [34]. The GM is composed of an immense number of microorganisms (bacteria, viruses, and fungi) with several functions, such as host nutrition, bone mineralization, immune system regulation, xenobiotic metabolism, proliferation of intestinal cells, and protection against pathogens [35, 36]. This bacterial community is dominated by anaerobic bacteria and includes 500–1000 species [37]. *Firmicutes* and *Bacteroidetes* are the most important phyla among the intestinal bacteria, with a

The duodenum and proximal jejunum normally contain small numbers of bacteria, usually lactobacilli and enterococci, which are facultative anaerobes. The distal ileum is a transition zone between sparse populations of aerobic bacteria of the proximal small intestine and very dense populations of anaerobic microorganisms in the large bowel. Occasional groups of bacteria can be found in low concentrations within the lumen of the small intestine. Bacteria do not form clusters, and the

luminal contents are separated from the mucosa by a mucus layer [13].

are able to modify liver damage. Diet and lifestyle modification and insulinsensitizing agents appear to be promisingly effective against NAFLD progression. However, these approaches may not be effective in some patients. Many other drugs are currently being studied to establish treatments for NAFLD. At present, no accepted drug treatment for NASH has been stated [24]. In this sense, it is very important to improve the knowledge of NAFLD physiopathology. Actually, the underlying precise mechanisms of NAFLD pathogenesis have just begun to be understood. The classic "multiple hit" theory states that lipid accumulation initiates hepatic steatosis and subsequently triggers multiple insults acting together (hormones/adipokines from adipose tissue, inflammation, deregulated fat metabolism, lipotoxicity, oxidative stress, mitochondrial dysfunction, and genetic and epigenetic factors), ultimately inducing NASH and cirrhosis [27]. Progression to NASH is linked to systemic inflammation, and it is associated with other pathological processes, such as innate immunity alterations, endoplasmic-reticulum stress, toll-like receptor (TLR) signaling, mitochondrial dysfunction, and intestinal dysbiosis [6, 28–32]. Regarding this last process, approximately 70–75% of blood that reaches the liver comes from the portal vein circulation that communicates the liver with the intestine [33]. The liver is continually exposed to GM-derived mediators, including bacteria and bacterial components, such as LPS, promoting

an inflammatory response that contributes to liver injury [13].

proportion of over 90% of the total community [38].

**46**

Intestinal dysbiosis is a process that may adversely impact metabolism and produce immune responses, favoring NAFLD progression. Important studies on the relationship of the GM with obesity have identified profound changes in the composition and metabolic function of the GM in subjects with obesity. Moreover, these studies demonstrated that the GM interacts with host epithelial cells to indirectly control energy expenditure and storage and activate inflammatory responses in NASH pathogenesis [44]. Qualitative or quantitative imbalances in the GM might have serious health consequences for the host, including small intestinal bacterial overgrowth (SIBO) syndrome [13]. Due to gut dysbiosis, there is an elevated production of toxic bacterial components and metabolic mediators, which consequently accumulate in the intestine. In addition, an increase in intestinal permeability and further disruption of the epithelial barrier lead to the release of these GM-derived mediators [42], which could reach the liver through portal circulation, favoring hepatic inflammation and the development of NAFLD [45, 46]. After disruption of the gut epithelial barrier, the liver is exposed to microbial products and metabolites resulting from bacterial metabolism [47, 48]. In this sense, it has been demonstrated that patients with NAFLD have gut dysbiosis, gut epithelial barrier dysfunction, and increased translocation of bacterial components to the liver [49]. For this reason, mediators derived from gut dysbiosis might also be related to the pathogenesis of the disease. Several previous studies in clinical settings have associated intestinal dysbiosis with the occurrence of NAFLD [50–52] and with the progression to NASH [10, 53].

Among the various factors, dietary habits are considered to be most influential on the gut microbiome in subjects with obesity and NAFLD patients. It is wellknown that a high-fat diet causes gut dysbiosis characterized by lowered species richness and changes in microbial composition, such as decreased *Bacteroidetes* and increased *Firmicutes* and *Proteobacteria* abundances [43]. On the other hand, *Prevotella*, a member of the phylum *Bacteroidetes*, is associated with plant-rich diets. *Prevotella*-dominated microbiotas have higher fiber utilizing capacity than *Bacteroides*-dominated microbiotas, producing higher amounts of SCFAs [54]. There are some studies that consider *Prevotella* to be a beneficial commensal bacterium [10, 55], but there are others that noted enriched fecal *Prevotella* in NASH or cirrhotic patients [56–58]. These contradictory results may be partly explained by the differences in populations, age, or NAFLD stages between the studies. In this sense, further studies on *Prevotella* should be directed to characterize properties at the species level and to evaluate these species in different stages of NAFLD.

GM-derived mediators resulting from intestinal dysbiosis could play a key role in NAFLD progression through several mechanisms: (1) enhanced energy extraction from food nutrients by formation of SCFAs; (2) modulation of BA synthesis, which is crucial for fat absorption and affects metabolism of glucose via farnesoid X receptor (FXR); (3) innate immune system activation by bacterial component translocation; (4) endogenous ethanol production; and (5) reduction in choline metabolism, which reduces efflux of very-low-density lipoprotein (VLDL) from hepatocytes, promoting inflammation. These mechanisms involve translocation

of these mediators, such as SCFAs, BAs, endogenous ethanol, and choline metabolites, which may be potentially evaluated as noninvasive blood markers of NAFLD progression [59].
