Exploring in vitro modeling in hepatocarcinogenesis research: morphological and molecular features and similarities to the corresponding human disease.

The hepatocellular carcinoma (HCC) features a remarkable epidemiological burden, ranking as the third most lethal cancer worldwide. As the HCC-related molecular and cellular complexity unfolds as the disease progresses, the use of a myriad of in vitro models available is mandatory in translational preclinical research setups. In this review paper, we will compile cutting-edge information on the in vitro bioassays for HCC research, (A) emphasizing their morphological and molecular parallels with human HCC; (B) delineating the advantages and limitations of their application; and (C) offering perspectives on their prospective applications. While bidimensional (2D) (co) culture setups provide a rapid low-cost strategy for metabolism and drug screening investigations, tridimensional (3D) (co) culture bioassays - including patient-derived protocols as organoids and precision cut slices - surpass some of the 2D strategies limitations, mimicking the complex microarchitecture and cellular and non-cellular microenvironment observed in human HCC. 3D models have become invaluable tools to unveil HCC pathophysiology and targeted therapy. In both setups, the recapitulation of HCC in different etiologies/backgrounds (i.e., viral, fibrosis, and fatty liver) may be considered as a fundamental guide for obtaining translational findings. Therefore, a "multimodel" approach - encompassing the advantages of different in vitro bioassays - is encouraged to circumvent "model-biased" outcomes in preclinical HCC research.

Past, present, and future of chemically induced hepatocarcinogenesis rodent models: Perspectives concerning classic and new cancer hallmarks.

Hepatocellular carcinoma (HCC), the main primary liver cancer, accounts for 5 % of all incident cases and 8.4 % of all cancer-related deaths worldwide. HCC displays a spectrum of environmental risk factors (viral chronic infections, aflatoxin exposure, alcoholic- and nonalcoholic fatty liver diseases) that result in molecular complexity and heterogeneity, contributing to a rising epidemiological burden, poor prognosis, and non-satisfactory treatment options. The emergence of HCC (i.e., hepatocarcinogenesis) is a multistep and complex process that addresses many (epi)genetic alterations and phenotypic traits, the so-called cancer hallmarks. "Polymorphic microbiomes", "epigenetic reprogramming", "senescent cells" and "unlocking phenotypic plasticity" are trending hallmarks/enabling features in cancer biology. As the main molecular drivers of HCC are still undruggable, chemically induced in vivo models of hepatocarcinogenesis are useful tools in preclinical research. Thus, this narrative review aimed at recapitulating the basic features of chemically induced rodent models of hepatocarcinogenesis, eliciting their permanent translational value regarding the "classic" and the "new" cancer hallmarks/enabling features. We gathered state-of-art preclinical evidence on non-cirrhotic, inflammation-, alcoholic liver disease- and nonalcoholic fatty liver-associated HCC models, demonstrating that these bioassays indeed express the recently added hallmarks, as well as reflect the interplay between classical and new cancer traits. Our review demonstrated that these protocols remain valuable for translational preclinical application, as they recapitulate trending features of cancer science. Further "omics-based" approaches are warranted while multimodel investigations are encouraged in order to avoid "model-biased" responses.

Western diet-induced mouse model of non-alcoholic fatty liver disease associated with metabolic outcomes: Features of gut microbiome-liver-adipose tissue axis.

OBJECTIVES: Non-alcoholic fatty liver disease (NAFLD) has a growing epidemiologic and economic burden. It is associated with Western diet (WD) patterns, and its pathogenesis involves metabolic disorders (obesity, dyslipidemia, hyperglycemia, and diabetes) and gut dysbiosis, features that are usually neglected or not reproduced by most animal models. Thus, we established a 6-mo WD-induced NAFLD mouse model associated with metabolic disorder, investigating its main features at the gut microbiome-liver-adipose tissue axis, also evaluating the correlations of gut dysbiosis to the other disease outcomes. METHODS: Male C57 BL6 mice received a high-fat (30% lard and 0.2% cholesterol, ∼57% calories) and sucrose-rich (20%) chow, and a high-sugar solution (23.1 and 18.9 g/L of D-fructose and D-glucose) for 6 mo. RESULTS: The model featured high serum cholesterol levels, glucose intolerance, and hyperinsulinemia. WD intervention resulted in extensive macro/microvesicular liver steatosis and pericellular fibrosis-resembling human disease-accompanied by hepatic stellate cell activation and CD68+ macrophage infiltration, increased protein levels of proinflammatory p65-nuclear factor-κB, interleukin-6 and tumor necrosis factor-α, with decreased antioxidant regulator Nrf2. Mice showed clear obesity with adipocyte hypertrophy, and CD68+macrophage/mast cell infiltration in adipose tissue while a reduction in number of goblet cells was also observed in the small intestine. Moreover, the pyrosequencing of the 16 S ribosomal RNA of gut cecal content showed decreased bacterial diversity, enriched Firmicutes and Proteobacteria, decreased Bacteroidetes and Fusobacteria, and increased ratio of Firmicutes to Bacteroidetes. Bacteroidetes and Bacteroides had the highest number of significant correlations with liver-adipose tissue axis outcomes. In silico analysis of gut microbiome in NAFLD obese patients revealed a depletion in Bacteroides, which also correlated to disease outcomes. CONCLUSION: This mice model gathered suitable phenotypical alterations in gut-liver-adipose tissue axis that resembled NAFLD associated with metabolic disorders in humans and may be considered for preclinical investigation.