Efeitos da exposição ao malathion em Colossoma macropomum: aspectos bioquímicos, histopatológicos e genéticos

Abstract

Malathion is an organophosphate insecticide used to control agricultural pests and combat disease vectors. Studies with fish demonstrate that insecticides are able to promote the differential expression of genes involved in tumorigenesis events, including the tumour suppressor tp53. In mammals, the degree and duration of the stressful condition is crucial to define which tp53-dependent pathway will be activated. In fish, however, it is unclear whether the regulation of tp53 target anti and pro-oxidant genes follows the same pattern described for mammals. In addition, fish responses after exposure to insecticides may vary depending on exposure time and mechanisms activated by fish to minimize contaminant impacts. Therefore, the general goal of the study was to understand the effects of malathion on tambaqui, through the analysis of multiple biomarkers, and how fish defense mechanisms are modulated to deal with these effects. In the first chapter, the lethal concentration capable of killing 50% of the individual was determined (LC50-96 h) and, later, the fish were sublethally exposed to malathion (7.30 mg L -1 ) for 96 h. the insecticide promoted an increase in the activity of biotransformation and antioxidant enzymes and a reduction in the production of reactive oxygen species. Mitochondrial respiration and acetylcholinesterase enzyme activity remained constant in both groups. However, fish exposed to the insecticide showed severe histopathological gill damage and increased ras proto-oncogene expression. The second chapter evaluated whether the duration of exposure to malathion differentially affected the biochemical responses of tambaqui. The fish were exposed for 6, 12, 24, 36 and 48 h to the insecticide (6.54 mg L - 1 ). Even in the first hours of exposure, acetylcholinesterase activity was kept similar in both, control and exposed to malathion groups, indicating absence of neurotoxicity. Oxidative damages occurred early within the first six hours of exposure and was accompanied by activation of antioxidant defenses. Mitochondrial respiration was differentially affected by the length of exposure to the insecticide, indicating the tambaqui seeks to maintain homeostasis and ATP demand. In the third chapter, the expression of target genes that participate in the regulation pathway of the tumour suppressor tp53 and tumorigenesis processes in tambaqui after exposure to malathion was evaluated. The fish were exposed to the insecticide for 6 and 48 h and the expression of 10 genes of interest was evaluated in the liver. The insecticide induced an increase in tp53 transcripts and differential expression of genes related to the tp53 regulatory pathway. Fish exposed to malathion showed an increase in transcripts of DNA damage response genes (atm/atr), pro-apoptotic gene bax, proto-oncogene mdm2 and antioxidant sesn1, as well as down regulation of anti-apoptotic bcl2. The malathion promoted an increase in hif-1 transcripts at 6 and 48 h, but the expression of the proto-oncogene ras was similar in both groups. Taken together, the results demonstrate that tambaqui has some resistance to malathion, possibly due to the activation of defense mechanisms that minimize the impacts of the contaminant, but the adverse effects of the insecticide are evident. At the transcriptional level, under severe stress conditions the activation of tp53 favors the apoptotic response. For the conditions tested in this study, the differential regulation of tumour suppressor genes and proto-oncogenes is an indication that malathion may predispose fish to the risk of tumorigenesis.