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Application of toxicogenomics for the risk assessment of the food contaminant acetamide

ABSTRACT: Acetamide (CAS 60-35-5) is detected in common foods including milk, eggs, beef, and roasted coffee beans. Chronic rodent bioassays using high doses (≥1000 mg/kg/day) suggest acetamide is a group 2B possible human carcinogen due to the induction of liver tumors. Weight-of-evidence indicates acetamide is not genotoxic, and therefore a threshold response is expected. We used a toxicogenomics approach in Wistar rats gavaged daily for 7 and 28 days to determine the benchmark dose (BMD), and investigate its mode of action from differential gene expression. Two exposure experiements were carried out in succsession. An initial dose range finding experiment used male Wistar rats gavaged daily for 7 days at 0, 30, 100, 300 and 1000 mg/kd/day. A second experiment used both male and female Wistar rats exposed for 7 and 28 days to 0, 300,500,750,1000 and 1500 mg/kg/day. No treatment related changes in terminal body weight, clinical chemistry, or histopathology were observed at doses of 30, 100, or 300 mkd. At 1000 mkd, the dose reported to elicit carcinogenesis, liver weights decreased 1.3-fold with the presence of single cell necrosis, hepatocyte vacuolization, and increased mitotic activity consistent with a 4.2 fold increase in the cell proliferation index. Accordingly, plasma ALT and AST were increased 2.0- and 2.2-fold, respectively. RNA-Seq analysis identified 1 differentially expressed gene at 300 mkd, and 2,685 at 1000 mkd (|fold-change| ≥ 1.5 and FDR ≤ 0.05). Down-regulated genes were associated with lipid metabolism while up-regulated genes included cell signaling, immune response, and cell cycle functions. Collectively, these results revealed a no-observable adverse effect level (NOAEL) and no-observed-transcriptional effect level (NOTEL) at 300 mkd, warranting further investigation at doses between 300 and 1000 mkd to identify the benchmark dose (BMD). Functional enrichment indicates perturbation of cell cycle and lipid metabolism, though a more refined dose-response evaluation will be needed to demonstrate dose-dependent effects related to the MOA of acetamide. Funding by the Bill and Melinda Gates Foundation (OPP1142801).

ORGANISM(S): Rattus norvegicus

PROVIDER: GSE132815 | GEO | 2020/01/13

REPOSITORIES: GEO

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Project description:Purpose: The International Commission on Radiological Protection (ICRP) recently recommended reducing the occupational equivalent dose limit for the lens of the eye. Based primarily on a review of epidemiological data, the absorbed dose threshold is now considered to be 0.5 Gy for induction of acute opacities, reduced from the previous threshold of 2 Gy. However, direct mechanistic evidence to support an understanding of the underlying molecular mechanisms of damage is still lacking. To this end, we explored the effects of a broad dose-range of ionizing radiation exposure on gene expression changes in a human lens epithelial (HLE) cell-line in order to better understand the shape of the dose-response relationship and identify transcriptional thresholds of effects. Methods: HLE cells were exposed to doses of 0, 0.01, 0.05, 0.25, 0.5, 2, and 5 Gy of X-ray radiation at two dose rates (1.62 cGy/min and 38.2 cGy/min). Cell culture lysates were collected 20 h post-exposure and analysed using whole-genome RNA-sequencing. Pathways and dose-thresholds of biological effects were identified using benchmark dose (BMD) modeling. Results: Transcriptional responses were minimal at doses less than 2 Gy. At higher doses there were a significant number of differentially expressed genes (DEGs) (p < 0.05, fold change > |1.5|) at both dose rates, with 1308 DEGs for the low dose rate (LDR) and 840 DEGs for the high dose rate (HDR) exposure. Dose-response modeling showed that a number of genes exhibited non-linear bi-phasic responses, which was verified by digital droplet PCR. BMD analysis showed the majority of the pathways responded at BMD median values in the dose range of 1.5-2.5 Gy, with the lowest BMD median value being 0.6 Gy for the HDR exposure. The minimum pathway BMD median value for LDR exposure, however, was 2.5 Gy. Although the LDR and HDR exposures shared pathways involved in extracellular matrix reorganization and collagen production with BMD median value of 2.9 Gy, HDR exposures were more effective in activating pathways associated with DNA damage response, apoptosis, and cell cycling relative to LDR exposure. Conclusion: Overall, the results suggest that radiation induces complex non-linear transcriptional dose-response relationships that are dose-rate dependent. Pathways shared between the two dose rates may be important contributors to radiation-induced cataractogenesis. BMD analysis suggests that the majority of pathways are activated above 0.6 Gy, which supports current ICRP identified dose thresholds for deterministic effects to the lens of the eye of 0.5 Gy.

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Application of toxicogenomics for the risk assessment of the food contaminant acetamide

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