2.3.1. Chemical-Based Injury Models

Most common are models in which a toxic chemical substance (hepatotoxin), such as carbon tetrachloride (CCl4) or thioacetamide (TAA), is repeatedly administrated over a longer period [75,77]. At early time points, the applied toxins induce acute inflammation, while long-term intoxication results in robust and highly reproducible fibrosis, cirrhosis, or even HCC. During the acute phase, Kupffer cells induce an inflammatory response associated with secretion of chemokines, cytokines, and many other pro-inflammatory factors. This milieu attracts phagocytic active white blood cells (monocytes, neutrophils, and lymphocytes), stimulating parenchymal necrosis. While short-term injections of toxins are often used to study liver regeneration after toxic injury, the prolonged application of these compounds are the most widespread models for analyzing hepatic fibrogenesis. Toxic drugs much less frequently used in liver fibrosis research are dialkyl nitrosamines, such as dimethylnitrosamine (DMN) and diethylnitrosamine (DEN). These are hepatocarcinogens that provoke severe liver damage in mice when given parenterally or orally [81]. In the acute phase, these compounds provoke intense neutrophilic infiltration and extensive centrilobular haemorrhagic necrosis. When applied for longer periods, both DMN and DEN induce massive bile duct proliferation, ending in fibrosis, bridging necrosis, and in liver cancer [81]. Based on their mutagenic and carcinogenic properties, DMN and DEN are frequently used in translational research by investigators interested in recapitulating the multi-stage process of human liver carcinogenesis or the formation of HCC.

The application of LPS alone or in combination with other substances such as D-galactosamine (D-GalN), *Mycobacterium bovis* bacille Calmette–Guérin (BCG), or the diptheroid *Corynebacterium parvum* is frequently used to induce severe hepatic inflammation or lethal hepatitis [79]. LPS is composed of lipid- and polysaccharide-containing moieties found in the outer membrane of Gram-negative bacteria. It acts as a PAMP that is recognized by TLR4 after binding to a special LPS-binding protein in the serum. Among others, the NF-κB pathway is activated and provokes the strong activation of Kupffer cells [79]. The simultaneous application of substances, such as the amino sugar D-GalN, that inhibits synthesis of different macromolecules and induces hepatocyte damage and formation of intracellular ROS leads to a several-thousand-fold increased susceptibility toward LPS. Single injection of this endotoxin is the most commonly used toxemia model and is suitable for inducing a shock-like state. However, compared with mice, humans are more sensitive to LPS. In humans, small traces of gut-derived LPS resulting from leakages in the gut epithelium entering the portal circulation are already adequate to establish severe hepatic inflammation.
