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Introduction: Arylamine N -acetyltransferase 2 (NAT2) expresses a well-defined genetic polymorphism in humans that modifies drug and xenobiotic metabolism. Recent studies and genome wide association studies have reported that genetic variants of NAT2 are associated with differential risks of developing dyslipidemia and cardiometabolic disorders, suggesting a previously unrecognized role of NAT2 in pathophysiology of metabolic disorders. In support of this notion, we recently showed that human NAT2 expression is differentially regulated by glucose and insulin. Moreover, our in silico analysis showed that NAT2 is co-expressed with nuclear receptors enriched in the liver, e.g., NR1H4 (FXR) and NR1I2 (PXR), that have been previously implicated in regulation of hepatic glucose and lipid homeostasis. Identification of transcriptional regulator(s) of human NAT2 would aid in understanding novel functions that it may play in the liver. Thus, the present study was designed to investigate if NAT2 is transcriptionally regulated by hepatic nuclear receptors. Methods: To test this, we treated cryopreserved human hepatocytes with agonists towards four different hepatic transcription factors/nuclear hormone receptors, namely FXR (NR1H4), PXR (NR1I2), LXR (NR1H3), and PPARα (PPARA), and measured their effects on the level of NAT2 mRNA. Results: While the treatment with a FXR, PXR, or LXR agonist (i.e., GW-4064, SR-12813, or GW-3965) significantly induced their respective target genes, treatment with these agonists did not significantly alter the transcript level of NAT2 in human hepatocytes. PPARα agonist, GW-7647, treatment resulted in a statistically significant decrease in the NAT2 transcript level. However, its magnitude was marginal. Conclusion: In summary, hepatic nuclear receptors we examined in the present study (FXR, PXR, LXR, and PPARα) did not significantly alter NAT2 expression in cryopreserved human hepatocytes. Additional studies are needed to identify transcriptional regulators of hepatic NAT2 expression. [ABSTRACT FROM AUTHOR]

Metabolic dysfunction associated-steatotic liver disease (MASLD)/metabolic dysfunction-associated steatohepatitis (MASH) is the liver manifestation of metabolic syndrome, which is characterized by insulin resistance, hyperglycemia, hypertension, dyslipidemia, and/or obesity. Environmental pollutant exposure has been recently identified as a risk factor for developing MASH. Heterocyclic amines (HCAs) are mutagens generated when cooking meat at high temperatures or until well-done. Recent epidemiological studies reported that dietary HCA exposure may be linked to insulin resistance and type II diabetes, and we recently reported that HCAs induce insulin resistance and glucose production in human hepatocytes. However, no previous studies have examined the effects of HCAs on hepatic lipid homeostasis. In the present study, we assessed the effects of two common HCAs, MeIQx (2-amino-3, 8-dimethylimidazo [4, 5-f] quinoxaline) and PhIP (2-amino-1-methyl-6-phenylimidazo[4, 5-b] pyridine), on lipid homeostasis in cryopreserved human hepatocytes. Exposure to a single concentration of 25 μM MeIQx or PhIP in human hepatocytes led to dysregulation of lipid homeostasis, typified by significant increases in lipid droplets and triglycerides. PhIP significantly increased expression of lipid droplet-associated genes, PNPLA3 and HSD17B13, and both HCAs significantly increased PLIN2. Exposure to MeIQx or PhIP also significantly increased expression of several key genes involved in lipid synthesis, transport and metabolism, including FASN, DGAT2, CPT1A, SCD, and CD36. Furthermore, both MeIQx and PhIP significantly increased intracellular cholesterol and decreased expression of PON1 which is involved in cholesterol efflux. Taken together, these results suggest that HCAs dysregulate lipid production, metabolism, and storage. The current study demonstrates, for the first time, that HCA exposure may lead to fat accumulation in hepatocytes, which may contribute to hepatic insulin resistance and MASH. [ABSTRACT FROM AUTHOR]

Purpose: Arylamine N-acetyltransferase 1 (NAT1), a phase II metabolic enzyme, is frequently upregulated in breast cancer. Inhibition or depletion of NAT1 leads to growth retardation in breast cancer cells in vitro and in vivo. A previous metabolomics study of MDA-MB-231 breast cancer cells suggests that NAT1 deletion leads to a defect in de novo pyrimidine biosynthesis. In the present study, we observed that NAT1 deletion results in upregulation of cytidine deaminase (CDA), which is involved in the pyrimidine salvage pathway, in multiple breast cancer cell lines (MDA-MB-231, MCF-7 and ZR-75-1). We hypothesized that NAT1 KO MDA-MB-231 cells show differential sensitivity to drugs that either inhibit cellular pyrimidine homeostasis or are metabolized by CDA. Methods: The cells were treated with (1) inhibitors of dihydroorotate dehydrogenase or CDA (e.g., teriflunomide and tetrahydrouridine); (2) pyrimidine/nucleoside analogs (e.g., gemcitabine and 5-azacytidine); and (3) naturally occurring, modified cytidines (e.g., 5-formyl-2′-deoxycytidine; 5fdC). Results: Although NAT1 KO cells failed to show differential sensitivity to nucleoside analogs that are metabolized by CDA, they were markedly more sensitive to 5fdC which induces DNA damage in the presence of high CDA activity. Co-treatment with 5fdC and a CDA inhibitor, tetrahydrouridine, abrogated the increase in 5fdC cytotoxicity in NAT1 KO cells, suggesting that the increased sensitivity of NAT1 KO cells to 5fdC is dependent on their increased CDA activity. Conclusions: The present findings suggest a novel therapeutic strategy to treat breast cancer with elevated NAT1 expression. For instance, NAT1 inhibition may be combined with cytotoxic nucleosides (e.g., 5fdC) for breast cancer treatment. [ABSTRACT FROM AUTHOR]

Heterocyclic amines (HCAs) are well-known for their mutagenic properties. One of the major routes of human exposure is through consumption of cooked meat, as certain cooking methods favor formation of HCAs. Recent epidemiological studies reported significant associations between dietary HCA exposure and insulin resistance and type II diabetes. However, no previous studies have examined if HCAs, independent of meat consumption, contributes to pathogenesis of insulin resistance or metabolic disease. In the present study, we have assessed the effect of three HCAs commonly found in cooked meat (2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline [MeIQ], 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline [MeIQx], and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine [PhIP]) on insulin signaling and glucose production. HepG2 or cryopreserved human hepatocytes were treated with 0–50 μM of MeIQ, MeIQx, or PhIP for 3 days. Treatment of HepG2 cells and hepatocytes with MeIQ and MeIQx resulted in a significant reduction in insulin-induced AKT phosphorylation, suggesting that HCA exposure decreases hepatic insulin signaling. HCA treatment also led to significant increases in expression of gluconeogenic genes, G6PC and PCK1, in both HepG2 and cryopreserved human hepatocytes. Additionally, the level of phosphorylated FOXO1, a transcriptional regulator of gluconeogenesis, was significantly reduced by HCA treatment in hepatocytes. Importantly, HCA treatment of human hepatocytes led to increases in extracellular glucose level in the presence of gluconeogenic substrates, suggesting that HCAs induce hepatic glucose production. The current findings suggest that HCAs induce insulin resistance and promote hepatic glucose production in human hepatocytes. This implicates that exposure to HCAs may lead to the development of type II diabetes or metabolic syndrome. [ABSTRACT FROM AUTHOR]

Arylamine N-acetyltransferase 2 (NAT2) is a phase II metabolic enzyme, best known for metabolism of aromatic amines and hydrazines. Genetic variants occurring in the NAT2 coding region have been well-defined and are known to affect the enzyme activity or protein stability. Individuals can be categorized into rapid, intermediate, and slow acetylator phenotypes that significantly alter their ability to metabolize arylamines, including drugs (e.g., isoniazid) and carcinogens (e.g., 4-aminobiphenyl). However, functional studies on non-coding or intergenic variants of NAT2 are lacking. Multiple, independent genome wide association studies (GWAS) have reported that non-coding or intergenic variants of NAT2 are associated with elevated plasma lipid and cholesterol levels, as well as cardiometabolic disorders, suggesting a novel cellular role of NAT2 in lipid and cholesterol homeostasis. The current review highlights and summarizes GWAS reports that are relevant to this association. We also present a new finding that seven, non-coding, intergenic NAT2 variants (i.e., rs4921913, rs4921914, rs4921915, rs146812806, rs35246381, rs35570672, and rs1495741), which have been associated with plasma lipid and cholesterol levels, are in linkage disequilibrium with one another, and thus form a novel haplotype. The dyslipidemia risk alleles of non-coding NAT2 variants are associated with rapid NAT2 acetylator phenotype, suggesting that differential systemic NAT2 activity might be a risk factor for developing dyslipidemia. The current review also discusses the findings of recent reports that are supportive of the role of NAT2 in lipid or cholesterol synthesis and transport. In summary, we review data suggesting that human NAT2 is a novel genetic factor that influences plasma lipid and cholesterol levels and alters the risk of cardiometabolic disorders. The proposed novel role of NAT2 merits further investigations. [ABSTRACT FROM AUTHOR]

Arylamine N -acetyltransferase 2 (NAT2) is well-known for its role in phase II metabolism of xenobiotics and drugs. More recently, genome wide association studies and murine models implicated NAT2 in regulation of insulin sensitivity and plasma lipid levels. However, the mechanism remains unknown. Transcript levels of human NAT2 varied dynamically in HepG2 (hepatocellular) cells, depending on the nutrient status of the culture media. Culturing the cells in the presence of glucose induced NAT2 mRNA expression as well as its N -acetyltransferase activity significantly. In addition, insulin or acetate treatment also significantly induced NAT2 mRNA. We examined and compared the glucose- and acetate-dependent changes in NAT2 expression to those of genes involved in glucose and lipid metabolism, including FABP1 , CPT1A , ACACA , SCD , CD36 , FASN , ACLY , G6PC , and PCK1. Genes that are involved in fatty acid transport and lipogenesis, such as FABP1 and CD36 , shared a similar pattern of expression with NAT2. In silico analysis of genes co-expressed with NAT2 revealed an enrichment of biological processes involved in lipid and cholesterol biosynthesis and transport. Among these, A1CF (APOBEC1 complementation factor) showed the highest correlation with NAT2 in terms of its expression in normal human tissues. The current study shows, for the first time, that human NAT2 is transcriptionally regulated by glucose and insulin in liver cancer cell lines and that the gene expression pattern of NAT2 is similar to that of genes involved in lipid metabolism and transport. [ABSTRACT FROM AUTHOR]

The correct definition of MeIQ is 2-amino-3,4-dimethylimidazo[4,5-f]quinoline, as correctly indicated in Fig. In both instances the abbreviation MeIQ was incorrectly defined as 2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline. [Extracted from the article]