Environmental electrophiles readily interact with reactive sulfur species (RSS), resulting in decline of cellular RSS levels accompanied by formation of their sulfur adducts. In the present study, we examined the effects of combined environmental electrophiles on consumption of persulfide and on cytotoxicity in HepG2 cells. A convenient assay with SSP4, a fluorometric probe for detection of per/polysulfides, indicated that each environmental electrophile caused compound-dependent consumption of persulfide. Consumption of persulfide by combined exposure to methylmercury (MeHg) and cadmium (Cd) was greater than that of the single exposure. Consistent with this finding, combined exposure to environmental electrophiles (MeHg, Cd, and 1,4-naphthoquinone) exacerbated concentration-dependent cellular toxicity in HepG2 cells compare to single exposure to each compound. As humans are exposed to environmental electrophiles daily, more attention should be paid to the study of combined exposure to environmental electrophiles, which can modulate cellular levels of RSS and disrupt redox homeostasis.

Drug-induced liver injury (DILI) is one of the serious and frequent drug-related adverse events and is classified into intrinsic and idiosyncratic types. Almost all of DILI caused in humans is known to be idiosyncratic type. The estimation of the potential risk for a drug candidate to induce idiosyncratic DILI is important to facilitate the development of new drugs, however, the estimation is difficult from the results of non-clinical toxicity studies using animals. We have previously reported the in vitro combination assay of mitochondrial function and apoptosis using human primary hepatocytes as a useful model for estimation of the risk of idiosyncratic DILI. In this study, to improve the in vitro assay for estimation of the risk of idiosyncratic DILI, we evaluate the usefulness of HepaRG cells for the mitochondrial function assay. We measured the oxygen consumption rate (OCR) as an endpoint of mitochondrial function in HepaRG cells treated with some compounds causing idiosyncratic DILI (troglitazone, leflunomide, ranitidine and diclofenac) and others known not to cause idiosyncratic DILI (acetaminophen and ethanol) and compared the results in HepaRG cells and with those for human primary hepatocytes as previously reported. As the results, HepaRG cells showed comparable or even higher sensitivity for detecting mitochondrial dysfunction than human primary hepatocytes, in all tested compounds. Taking into account these results and many other useful properties of the cells, HepaRG cells are considered to be much more suitable for this mitochondrial function assay than the human primary hepatocytes.

Maltosyltrehalose syrup (TG4 syrup) is an enzymatically derived starch hydrolysate consisting of only glucose. The main components are maltotetraose (G4) and maltosyltrehalose (TG4). G4 is a common component of essentially all starch hydrolysates, and consists of only four glucose molecules with α-1,4 linkages. TG4 also consists of four glucose units, but the terminal glycosidic bond at the reducing end of the glucose chain is inverted, which results in the final bond being α-1,1. The two terminal glucose units form a trehalose disaccharide molecule, attached to a maltose molecule. The presence of the trehalose moiety in the TG4 molecule results in unique functional and technical properties from what is found in other α-1,4 linked oligosaccharides. This paper presents results of standardized toxicity studies of TG4 syrup, including an in vitro bacterial mutagenicity test, a 90-day oral feeding study in rats, and a 90-day oral toxicity (gavage) study in rats. Treatment with TG4 syrup resulted in no microbial mutagenic activity or growth inhibition in either Salmonella typhimurium or Escherichia coli, even at the maximum dose of 5,000 μg/plate. The NOAEL in the 90-day oral feeding study was calculated as 10% of TG4 syrup in the diet, which was equal to 6,818 and 7,464 mg/kg/day (dwb) in male and female rats, respectively. The 90-day oral gavage toxicity study had a NOAEL of 5,000 mg/kg/day (dwb) in male and female rats. Taken together these data showed that TG4 syrup was safe for use in these studies, suggesting it is safe for consumption by humans.

Obesity is caused by a chronic positive energy balance, which not only increases the amount of lipid in adipose tissue, but also, in non-adipose tissue. Excessive accumulation of lipids in tissues may lead to cell dysfunction or cell death, a phenomenon known as lipotoxicity. The aim of this study was to investigate the effects of high-fat diet (HFD) feeding on the sciatic nerves of both male and female mice. HFD feeding induced increased mRNA levels of Bax and Bcl2 in HFD group of males only, although the accumulation of fatty acids in the sciatic nerves was induced by HFD feeding in the both sexes. To determine whether estrogen was involved in the inhibitory effects of HFD feeding-induced increased expression of apoptosis-related genes, ovariectomized (OVX) females were fed a normal diet (ND) or HFD with or without daily ethinylestradiol treatment for 1 week. In OVX mice, the mRNA levels of Bax and Bcl2 in HFD group were higher than in ND group. In contrast, in OVX mice treated with ethinylestradiol, there was no significant between ND and HFD groups. In conclusion, HFD induced apoptosis in the sciatic nerves of males, but not females, and estrogen had inhibitory effects on HFD-induced apoptosis in the sciatic nerves of females. Long-term feeding studies are needed to investigate the pathological effects on sciatic nerves of mice fed HFD as pathological findings were not observed under our study condition.

Epoxidized fatty acids are generated during food processing and detected in various lipid- and oil-containing foods. Although compounds with an epoxide structure have high reactivity and there is increasing concern about their potential toxic effects such as genotoxicity and cellular damage, information regarding the toxicity of epoxy fatty acids is largely unknown. Therefore, we conducted three in vitro genotoxicity studies (bacterial reverse mutation assay, in vitro micronucleus test, and p53R2-dependent luciferase reporter gene assay) and HepG2 cytotoxicity assays for epoxyoleic acid (EOA) and diepoxylinoleic acid (DELA). The in vitro genotoxicity results of EOA and DELA were uniformly negative. In the cytotoxicity assay, EOA and DELA induced weak cytotoxicity at high concentrations, but the effect was similar to those of oleic and linoleic acids at low concentrations. Considering the existing toxicity information on epoxidized soybean oil, which is a similar compound, there might be little concern concerning the health effects of epoxy fatty acids at low concentrations.