Evaluation of acute hepatic toxicity and inflammation induced by nitrosamines in Mus musculus and Danio rerio

Abstract

Nitrosamines are compounds classified as emerging contaminants, carcinogens, and oxidants. They can alter the properties of cell membranes and inducing an increase in biologically active lipids, such as arachidonic acid and PAF, a potent pro-inflammatory agent capable of inducing the onset of liver diseases. In this work, we will use two nitrosamines (N-Nitrosodiethylamine and N-Nitrosodimethylamine), chosen because they are the most found commercially, to perform hepatotoxic and acute inflammation on Mus musculus specimens will be exposed to different concentrations of each of the nitrosamines, to perform subsequent biochemical, histological and hematological analyses that show possible liver damage induced by exposure to this contaminant. Blood from Mus musculus specimens will be collected, and subsequently, the liver will be subjected to histological evaluation. An aliquot of the blood pool collected from the groups exposed and not exposed to nitrosamines will be submitted to hematological analysis, using smear techniques and subsequent cell counting in a Neubauer chamber. Another aliquot will be used for biochemical monitoring of lactate dehydrogenase, oxalacetic transaminase (GOT), pyruvic transaminase (GPT), gamma-glutamyl transferase (GGT), and C Reactive Protein, to monitor hepatic alterations. In addition, we will evaluate the levels of interleukin 6 (IL-6), tumor necrosis factor-alpha (TNF alpha), malondialdehyde (MDA) levels, and the enzyme lipoprotein-associated phospholipase A2 (Lp-PLA2), also known as platelet-activating factor acetylhydrolase (PAF-AH), because it is crucial in the process of controlling inflammation, being considered an anti-inflammatory protein since PAF-AH acts by regulating the plasma levels of platelet-activating factor (PAF), a lipid factor extremely important in the progression and development of the inflammatory process. Thus, our project aims to show that nitrosamines are agents that can induce hepatocyte dysfunction, that can affect energy metabolism and that platelet aggregation factor (PAF) would be an important agent causing hepatotoxicity. In addition to these in vivo and biochemical investigations, we will use molecular docking techniques that will be done in our laboratory and molecular dynamics, these will be done at BioSIM (University of Porto) to investigate the ability of nitrosamines as a potential non-selective inhibitor of LPCAT and PAF-AH enzymes. Since nitrosamine is a common compound produced in water environments, we also performed CL50% of both compound and monitoring of hematological and hepatic histology modifications induced by both compound in Danio rerio.