Evaluation of perinatal exposure of glyphosate and its mixture with 2,4-D and dicamba οn liver redox status in Wistar rats.

Wide-scale emergence of glyphosate-resistant weeds has led to an increase in the simultaneous application of herbicide mixtures exacerbated by the introduction of crops tolerant to glyphosate plus dicamba or glyphosate plus 2,4-D. This raises serious concerns regarding the environmental and health risks resulting from increased exposure to a mixture of herbicide active ingredients. We evaluated hepatotoxic effects following perinatal exposure to glyphosate alone or in combination with 2,4-D and dicamba from gestational day-6 until adulthood in Wistar rats. Animals were administered with glyphosate at the European Union (EU) acceptable daily intake (ADI; 0.5 mg/kg bw/day) and no-observed-adverse-effect level (NOAEL; 50 mg/kg bw/day). A mixture of glyphosate with 2,4-D (0.3 mg/kg bw/day) and dicamba (0.02 mg/kg bw/day) with each at their EU ADI was evaluated. Redox status was determined by measuring levels of reduced glutathione, decomposition rate of Η2Ο2, glutathione reductase, glutathione peroxidase, total antioxidant capacity, thiobarbituric reactive substances, and protein carbonyls. Gene expression analysis of Nr1d1, Nr1d2, Clec2g, Ier3, and Gadd45g associated with oxidative damage to DNA, was also performed. Analysis of liver samples showed that exposure to the mixture of the three herbicides induced a marked increase in the concentration of glutathione and malondialdehyde indicative of a disturbance in redox balance. Nevertheless, the effect of increased lipid peroxidation was not discernible following a 3-month recuperation period where animals were withdrawn from pesticide exposure post-weaning. Interestingly, toxic effects caused by prenatal exposure to the glyphosate NOAEL were present after the same 3-month recovery period. No statistically significant changes in the expression of genes linked with genotoxicity were observed. Our findings reinforce the importance of assessing the combined effects of chemical pollutants at doses that are asserted by regulatory agencies to be safe individually.

Comparative Toxicogenomics of Glyphosate and Roundup Herbicides by Mammalian Stem Cell-Based Genotoxicity Assays and Molecular Profiling in Sprague-Dawley Rats.

Whether glyphosate-based herbicides (GBHs) are more potent than glyphosate alone at activating cellular mechanisms, which drive carcinogenesis remain controversial. As GBHs are more cytotoxic than glyphosate, we reasoned they may also be more capable of activating carcinogenic pathways. We tested this hypothesis by comparing the effects of glyphosate with Roundup GBHs both in vitro and in vivo. First, glyphosate was compared with representative GBHs, namely MON 52276 (European Union), MON 76473 (United Kingdom), and MON 76207 (United States) using the mammalian stem cell-based ToxTracker system. Here, MON 52276 and MON 76473, but not glyphosate and MON 76207, activated oxidative stress and unfolded protein responses. Second, molecular profiling of liver was performed in female Sprague-Dawley rats exposed to glyphosate or MON 52276 (at 0.5, 50, and 175 mg/kg bw/day glyphosate) for 90 days. MON 52276 but not glyphosate increased hepatic steatosis and necrosis. MON 52276 and glyphosate altered the expression of genes in liver reflecting TP53 activation by DNA damage and circadian rhythm regulation. Genes most affected in liver were similarly altered in kidneys. Small RNA profiling in liver showed decreased amounts of miR-22 and miR-17 from MON 52276 ingestion. Glyphosate decreased miR-30, whereas miR-10 levels were increased. DNA methylation profiling of liver revealed 5727 and 4496 differentially methylated CpG sites between the control and glyphosate and MON 52276 exposed animals, respectively. Apurinic/apyrimidinic DNA damage formation in liver was increased with glyphosate exposure. Altogether, our results show that Roundup formulations cause more biological changes linked with carcinogenesis than glyphosate.

Multi-omics phenotyping of the gut-liver axis reveals metabolic perturbations from a low-dose pesticide mixture in rats.

Health effects of pesticides are not always accurately detected using the current battery of regulatory toxicity tests. We compared standard histopathology and serum biochemistry measures and multi-omics analyses in a subchronic toxicity test of a mixture of six pesticides frequently detected in foodstuffs (azoxystrobin, boscalid, chlorpyrifos, glyphosate, imidacloprid and thiabendazole) in Sprague-Dawley rats. Analysis of water and feed consumption, body weight, histopathology and serum biochemistry showed little effect. Contrastingly, serum and caecum metabolomics revealed that nicotinamide and tryptophan metabolism were affected, which suggested activation of an oxidative stress response. This was not reflected by gut microbial community composition changes evaluated by shotgun metagenomics. Transcriptomics of the liver showed that 257 genes had their expression changed. Gene functions affected included the regulation of response to steroid hormones and the activation of stress response pathways. Genome-wide DNA methylation analysis of the same liver samples showed that 4,255 CpG sites were differentially methylated. Overall, we demonstrated that in-depth molecular profiling in laboratory animals exposed to low concentrations of pesticides allows the detection of metabolic perturbations that would remain undetected by standard regulatory biochemical measures and which could thus improve the predictability of health risks from exposure to chemical pollutants.