In-Depth Proteome Analysis Highlights HepaRG Cells as a Versatile Cell System Surrogate for Primary Human Hepatocytes

Abstract

Of the hepatic cell lines developed for in vitro studies of hepatic functions as alternatives to primary human hepatocytes, many have lost major liver-like functions, but not HepaRG cells. The increasing use of the latter worldwide raises the need for establishing the reference functional status of early biobanked HepaRG cells. Using deep proteome and secretome analyses, the levels of master regulators of the hepatic phenotype and of the structural elements ensuring biliary polarity were found to be close to those in primary hepatocytes. HepaRG cells proved to be highly differentiated, with functional mitochondria, hepatokine secretion abilities, and an adequate response to insulin. Among differences between primary human hepatocytes and HepaRG cells, the factors that possibly support HepaRG transdifferentiation properties are discussed. The HepaRG cell system thus appears as a robust surrogate for primary hepatocytes, which is versatile enough to study not only xenobiotic detoxification, but also the control of hepatic energy metabolism, secretory function and disease-related mechanisms.

NADPH-cytochrome P450 reductase expression and enzymatic activity in primary-like human hepatocytes and HepG2 cells for in vitro biotransformation studies

“…To overcome these limitations, several liver cancer-derived cell lines such as HepaRG and HepG2 were developed to serve as surrogate for pHHs. Advantages are their unlimited availability, convenient handling and proliferative capacity [20][21][22][23][24]. A clear disadvantage is their genetic instability due to their cancer origin, which makes them almost unusable for clinical applications such as diseaserelated liver repopulation.…”

The GOLIATH Project: Towards an Internationally Harmonised Approach for Testing Metabolism Disrupting Compounds

“…Of note, the differentiated HepaRG cells present mitochondrial function close to primary human hepatocytes [45,46]. More generally, recent investigations reported that the deep proteome of HepaRG cells is overall similar to that of primary human hepatocytes [47].…”

The application of omics-based human liver platforms for investigating the mechanism of drug-induced hepatotoxicity in vitro

“…Even though the activity levels of cytochrome oxidase are higher in HepaRG than in HepG2 cells, the levels of their cytochrome activities are markedly decreased in both cell lines compared to those in PHH (reduced by 60% and 90% in HepaRG and HepG2 cells, respectively) (Sison-Young et al 2015). At the proteome level, there are detectable differences in the proteins regulating cell senescence and proliferation between HepaRG and PHH, probably due to the transdifferentiation features of HepaRG, reminding their similarity to stem cells and liver cancer cells (Tascher et al 2019), which raise concerns about the applicability of these in vitro models in DILI studies.…”

“…Recently, studies have demonstrated that compared to HepG2 cells, the baseline transcriptomic (Hart et al 2010;Jennen et al 2010;Jetten et al 2013) and proteomic (Sison-Young et al 2015) profiling of differentiated HepaRG cells indicates a higher resemblance to PHH. Tascher et al also showed that the amounts of proteins related to bile acid synthesis, conjugation, detoxification and transport expressed by HepaRG are comparable to PHH, encouraging the use of these cells in studies of drug metabolism, compound-induced liver injury and pathogenesis of cholestatic liver diseases in humans (Tascher et al 2019). The molecular mechanisms for the hepatic cholestasis (Rodrigues et al 2018), steatosis (Mesnage et al 2018) and cytotoxicity have been studied by means of multi-omicsbased methods after exposure of HepaRG cells to a number of compounds (Seeger et al 2019a;Smith et al 2018).…”