RF22 | PMON10 Exposure to a Real Life Environmental Chemical Mixture Alters the Metabolite Profile in the Plasma of Sheep: An Untargeted Metabolomics Study.
Authors: Evans Neil , Lea Richard , Padmanabhan Vasantha , Thangaraj Soundara Viveka , Maureen Kanchan , Bellingham Michelle , Sinclair Kevin 
Published: November 2022

Abstract
Humans are exposed to a mixture of endocrine disrupting chemicals (EDCs) in everyday life, believed to contribute to a range of non-communicable diseases. While the adverse health impacts of exposure to individual EDCs have been extensively investigated, studies on effects of real-world exposures characterized by chronic low doses of a mixture of chemicals, are only recently emerging. Biosolids derived from human wastes, exemplify both the array and dosage of chemicals humans are exposed to in real life and represents a novel model to investigate the risk posed by the EDCs. Previous studies in sheep have demonstrated aberrant reproductive and metabolic phenotypic effects in offsprings, as a result of maternal biosolid exposure. We hypothesize that EDC-mediated disruption in the maternal metabolic milieu is a potential mediator of the aberrant phenotypic outcomes in offsprings in the biosolid-exposure model. Recent untargeted metabolomic studies provide evidence of the technique's ability to unravel biomarkers of both exposure and effect by identifying the disrupted endogenous metabolite profile and xenobiotic mixture, respectively. In this study, we sought to identify biosolid exposure-induced maternal metabolic fingerprints that could influence fetal programming.
Ewes were grazed on either organic fertilizer or biosolid-treated pastures from 1 month before mating until close to parturition. Plasma from blood samples collected on day 90 of the 147-day gestation period, from ewes carrying singleton pregnancies (Control n=15, Treatment n=15) were subjected to untargeted metabolomics using liquid chromatography-quadrupole time-of-flight mass spectrometry. Metabolites were identified by matching the retention time, mass and isotope profile to authentic standards or metabolome databases using Agilent's MassHunter Find by Molecular Feature workflow (v7.0).
Untargeted metabolomics identified 1006 and 1295 annotated features from the negative and positive mode respectively. Principal component analysis and orthogonal signal correction partial least-squares discriminant analysis discriminated the metabolomes of control sheep and sheep grazed in biosolid-treated pasture. The key differential metabolites identified by variable importance in projection score primarily belonged to the super-class organic acids, fatty acids, benzenoids, glycerophospholipids and organoheterocyclic compounds. These differentially expressed metabolites are implicated in metabolic pathways including carnitine synthesis, alanine metabolism, glucose alanine cycle, phosphatidyl choline biosynthesis and glutathione metabolism. Some of the top differential metabolites- oleamide, pyroglutamic acid and 15-HETrE have been implicated in fetal development. Oleamide, an endocannabinoid, has growth inhibitory properties. 15-HETrE modulates arachidonic acid metabolism and upregulate PPARγ expression, which plays a pivotal role in fetal development and metabolic outcomes.
These findings indicate that biosolids exposure results in alterations in the maternal metabolome which likely contributes to the aberrant phenotypic outcomes reported in offspring of biosolid-exposed mothers. This is the first study to apply metabolomics to validate the effect of real-life EDC exposure in sheep, a translationally relevant precocial model with a developmental trajectory similar to humans.
Presentation: Sunday, June 12, 2022 1:00 p.m. - 1:05 p.m., Monday, June 13, 2022 12:30 p.m. - 2:30 p.m.