Oxidative stress induced by organophophorus pesticides depends on membrane lipid composition and domain organization

BELVER GOTTAU MB ; OUSSET J; VENTURINO A; JAUREGUIBERRY MS

Salta

Congreso; LV Annual SAIB Meeting and XIV PABMB Congress; 2019

Sociedad Argentina de Investigación Bioquímica y Biología Molecular (SAIB)

Chlorpyrifos (CP) is an organophosphorus pesticide (OP) widely used in the Alto Valle region since the fruit growing market is a major economic activity. Over the last decades, urban areas have overlapped with producing lands, leaving periurban inhabitants chronically exposed to environmental hazard. Although acetilcholinesterase (AChE) is the primary target of OPs, secondary targets such as oxidative stress enzymes become relevant at sublethal concentrations. Plasma membrane (PM) cholesterol (Chol) content and domain organization influence cell function. OPs have been suggested to compete with Chol for partition domains at the PM and to cause structure perturbations. We aimed to analyze the effect of PM lipid composition on the expression of enzymes involved in oxidative stress response, upon CP exposure in vitro. Our cell line model shows three different PM lipid composition achieved by stable transfection of CHO-K1 cells with rat fatty acid desaturases. CHO-K1 cells transfected with stearoil Co-A desaturase (SCD cells) show higher Chol content while cells co-transfected with fatty acid desaturases 1 and 2 (FADS cells), have lower Chol level at the PM due to higher polyunsaturated fatty acid content. First, we analyzed AChE expression by RT-PCR and found no constitutive expression compared to two housekeeping genes (beta-actin and GAPDH), validating this cell model as ideal for studying CP secondary targets. Then we analyzed catalase (CAT) and glutathione S-transferase (GST) expression after 1uM or 500 uM CP chronic (24h) exposure, relative to three housekeeping genes (beta-actin, GAPDH and Eif3-i). Regarding GST, expression level was not significantly altered at 1 uM CP for either cell line. However, CHO-K1 and SCD showed moderate increase at 500 uM CP, while expression in FADS cells rised over 2 fold after exposure to the highest concentration. Additionally, CAT results exhibited higher expression only at 500 uM CP both in SCD and FADS cells, whereas CHO-K1 CAT expression was not significantly altered by CP exposure. We also analyzed enzymatic activity by spectrophotometry (for CAT and GST) and found differential response upon CP exposure, depending to membrane lipid composition. These preliminary results suggest that lipid composition and domain organization at the PM affect not only CP entry into target cells but also their response to oxidative stress induced by this pesticide, probably through toxicant absorption kinetics.