Bisphenol a induces steatosis in HepaRG cells using a model of perinatal exposure

Abstract

Human exposure to bisphenol A (BPA) could favor obesity and related metabolic disorders such as hepatic steatosis. Investigations in rodents have shown that these deleterious effects are observed not only when BPA is administered during the adult life but also with different protocols of perinatal exposure. Whether perinatal BPA exposure could pose a risk in human is currently unknown, and thus appropriate in vitro models could be important to tackle this major issue. Accordingly, we determined whether long-term BPA treatment could induce steatosis in human HepaRG cells by using a protocol mimicking perinatal exposure. To this end, the kinetics of expression of seven proteins differentially expressed during liver development was determined during a 4-week period of cell culture required for proliferation and differentiation. By analogy with data reported in rodents and humans, our results indicated that the period of cell culture around day 15 and day 18 after seeding could be considered as the "natal" period. Consequently, HepaRG cells were treated for 3 weeks with BPA (from 0.2 to 2000 nM), with a treatment starting during the proliferating period. BPA was able to induce steatosis with a nonmonotonic dose response profile, with significant effects on neutral lipids and triglycerides observed for the 2 nM concentration. However, the expression of many enzymes involved in lipid and carbohydrate homeostasis was unchanged in exposed HepaRG cells. The expression of other potential BPA targets and enzymes involved in BPA biotransformation was also determined, giving answers as well as new questions regarding the mechanisms of action of BPA. Hence, HepaRG cells provide a valuable model that can prove useful for the toxicological assessment of endocrine disruptors on hepatic metabolisms, in particular in the developing liver. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 1024-1036, 2017.

Possible Involvement of Mitochondrial Dysfunction and Oxidative Stress in a Cellular Model of NAFLD Progression Induced by Benzo[a]pyrene/Ethanol CoExposure

“…First, wild-type (WT) and AhR-deficient HepaRG cells were characterized in nonsteatotic condition ( Figure 6 ). Compared to WT HepaRG cells, mut AhR HepaRG cells expressed very low levels of the AhR protein but presented normal (or subnormal) levels of CYP2E1, CYP3A4, albumin, and aldolase B ( Figure 6(a) ), which are specifically expressed in differentiated HepaRG cells [ 26 ]. Importantly, the mut AhR HepaRG cells were fully irresponsive to the dose-dependent effect of ITE on EROD activity ( Figure 6(b) ), thus indicating that the remaining low expression of AhR was unable to activate the expression of CYP1A1/2 and CYP1B1.…”

“…A clonal selection was performed by seeding 1 cell/well in a 96-well plate. After growth and expansion, the cell clone of interest (henceforth referred to as mut AhR -HepaRG cells) was selected among about 60 different clones on four parameters: (1) cell morphology, (2) AhR mRNA and protein expression, (3) EROD activity after a 2-day treatment with the potent AhR activator ITE, and (4) mRNA and protein expression of aldolase B, albumin, CYP3A4, and CYP2E1, which are bona fide markers of hepatocellular differentiation [ 26 ].…”

Drug-Induced Alterations of Mitochondrial DNA Homeostasis in Steatotic and Nonsteatotic HepaRG Cells

“…Interestingly, there was a significant correlation between the PGC1a mRNA expression and the phospho-AMPKa/total AMPK ratio in treated HepaRG cells. Despite these advantages, HepaRG cells present some limitations, as previously pointed out (Michaut et al, 2016;Bucher et al, 2017). For instance, this cell line derives from one female patient and thus does not allow identifying gender differences or genetic susceptibility.…”

Regulation of Cytosolic Sulfotransferases in Models of Human Hepatocyte Development

“…Undifferentiated and differentiated HepaRG cells have distinct gene expression profiles that tend to reflect the patterns seen in fetal and adult human liver, respectively. Genes that are primarily expressed in fetal hepatocytes, such as CYP3A7 and pyruvate kinase muscle isozyme, are more highly expressed in undifferentiated HepaRG cells, whereas genes that are predominantly expressed in adult hepatocytes, such as CYP3A4 and CYP2E1, are more highly expressed in differentiated HepaRG cells (Tsuji et al, 2014;Bucher et al, 2017). HepaRG cells are becoming established as a useful model for studying hepatocellular differentiation, xenobiotic metabolism and toxicity, and development of liver diseases (Sharanek et al, 2015;Nunn et al, 2016;Rodrigues et al, 2016;Sayyed et al, 2016;Xia et al, 2016).…”

“…Most studies using HepaRG cells have used differentiated cells as a model to complement the use of primary cultured human hepatocytes (Josse et al, 2008; Lubberstedt et al, 2011; Gerets et al, 2012;Klein et al, 2015). However, few studies have evaluated the changes in xenobiotic-metabolizing enzyme expression that occur as HepaRG cells pass through the stages of the differentiation process (Aninat et al, 2006;Hart et al, 2010;Ceelen et al, 2011;Tsuji et al, 2014;Bucher et al, 2017), and no studies have determined expression of the individual SULTs. We found that SULT1B1, SULT1C2, SULT1C3, SULT1C4, and SULT1E1 mRNA levels were highest in confluent HepaRG cells, whereas SULT2A1 RNA levels increased throughout the differentiation process.…”