3.1. NAFLD/NASH Pathogenesis
NAFLD is a complex disease trait where inter-patient genetic and epigenetic variations and environmental factors are combined to define development and disease progression [34] (Figure 2).
The generally accepted dogma in the pathogenesis of NAFLD is that insulin resistance—commonly associated with obesity—leads to the hepatic accumulation of triglycerides, a process that usually results from increased FFAs flux from adipose tissue to the liver, dietary fat via chylomicron metabolism, and increased de novo lipogenesis [35,36,37]. Obesity generates a state of low-grade inflammation characterized by the accumulation of immune cells in adipose tissue (particularly macrophages), and the production of proinflammatory cytokines by adipocytes contributes to the development of systemic and hepatic insulin resistance [35,36,37]. However, multiple pathways (multiple hits) are involved in the development of NASH and fibrosis. “Hits” that may contribute include oxidative stress, endotoxins, changes in the gut-liver axis, and mitochondrial dysfunction [34].
Oxidative stress and mitochondrial dysfunction were proposed as main triggers for the progression of steatosis to steatohepatitis [38]. FFAs catabolism in the liver takes place mainly via mitochondrial β-oxidation—a process that can lead to the generation of ROS, including superoxide, hydrogen peroxide, and hydroxyl radicals, in the case of increased FFAs delivery [38].
In the same context, impaired mitochondrial activity in NASH patients due to reduced enzymatic activities of mitochondrial electron transport chain and excessive in FA oxidation results in hepatic ATP depletion and may cause structural mitochondrial abnormalities consisting of enlargement (megamitochondria), paracrystalline inclusions, and loss of cristae [39,40].
In NAFLD, enhanced cytochrome P450 2E1 (CYP2E1) expression and activity seem to be an important source of ROS which trigger oxidative stress and perpetuate the hepatic mitochondrial dysfunction [41]. Moreover, it has been reported that upregulated microsomal CYP4A enzymes ω-hydroxylate fatty acids into dicarboxylic acids that are then preferentially oxidized by peroxisomes, thus promoting ROS production in NAFLD [42]. The ablation of a homolog of human CYP4A gene (CYP4A14) in animal models of steatohepatitis has shown to attenuate hepatic inflammation and fibrosis [43]. The abundant production of ROS induces the peroxidation of hepatic triglycerides with the release of reactive aldehydes such as 4-hydroxynonenal (4-HNE) and malondialdehyde (MDA) which can damage mitochondrial components [44].
Accumulating data have implicated the disruption of endoplasmic reticulum (ER) homeostasis (i.e., ER stress) in the development of NASH [45]. Factors that disturb ER folding capacity (e.g., excessive protein synthesis, mitochondrial dysfunction, oxidative stress) will lead to the activation of a physiologic mechanism called the “unfolded protein response” (UPR) in hepatocytes. This adaptive mechanism aims to increase the folding capacity of the ER, thus bringing the organelle and the cell into a state of equilibrium. When the activation of the UPR fails to promote cell survival, the cell is taken down the pro-apoptotic ER stress response pathway, which can ultimately lead to apoptotic cell death, inflammation, and/or fat accumulation [45].
Several studies have suggested a role of gut microbiome in NASH pathogenesis [46]. Intestinal barrier alterations cause increased intestinal permeability of bacteria, viruses, or microbial products such as lipopolysaccharide (LPS). These pathogens are recognized through specialized recognition receptors that include toll-like receptors (TLRs) and inflammasomes, inducing a signaling cascade leading to the production of inflammatory cytokines [46].