Dichlorodiphenylchloroethylene elevates cytosolic calcium concentrations and oscillations in human granulosa cells

Background and Significance: 1,1-Dichloro-2,2-bis(p-chlorophenyl)ethylene (DDE), a metabolite of DDT (1,1-dichlorodiphenyltrichloroethane) is a persistent hormonally active environmental toxicant which has been found in human serum and follicular fluid. It is present in over 90% of the North American population and could have adverse reproductive effects.Objective: The objective of this study was to determine whether DDE can alter free calcium ion concentrations [Ca2+]cyt in the cytosol of human granulosa cells.Materials and Methods: Changes of [Ca2+]cyt concentration in Fura-2 loaded single cells under various experimental conditions were studied using a dynamic digital Ca2+ imaging system.Results: At a concentration of 100 ng/mL, DDE stimulated small elevations of [Ca2+]cyt accompanied by Ca2+ oscillations. At 1000 ng DDE/mL there was a biphasic Ca2+ response with marked elevations of [Ca2+ ]cyt over time. In Ca2+ free medium, cells still showed an initial small elevation of [Ca2+ ]cyt which was magnified on addition of Ca2+ to the medium. Washing the cells after DDE treatment failed to remove the elevated [Ca2+]cyt and oscillations both of which were eliminated by addition of EGTA. Adenosine triphosphate (ATP) also induced [Ca2+]cyt elevations and oscillations and these effects were potentiated on the addition of DDE. Follicle stimulating hormone (FSH) induced transient [Ca2+]cyt elevations in 3 of 7 experiments.Conclusions: These results suggest that DDE at concentrations normally found in human tissues is an environmental toxicant and can induce elevations in [Ca2+]cyt. Our data therefore highlight a novel mechanism through which DDE can alter endocrine homeostasis and act as an endocrine toxicant. Background and Significance: 1,1-Dichloro-2,2-bis(p-chlorophenyl)ethylene (DDE), a metabolite of DDT (1,1-dichlorodiphenyltrichloroethane) is a persistent hormonally active environmental toxicant which has been found in human serum and follicular fluid. It is present in over 90% of the North American population and could have adverse reproductive effects. Objective: The objective of this study was to determine whether DDE can alter free calcium ion concentrations [Ca2+]cyt in the cytosol of human granulosa cells. Materials and Methods: Changes of [Ca2+]cyt concentration in Fura-2 loaded single cells under various experimental conditions were studied using a dynamic digital Ca2+ imaging system. Results: At a concentration of 100 ng/mL, DDE stimulated small elevations of [Ca2+]cyt accompanied by Ca2+ oscillations. At 1000 ng DDE/mL there was a biphasic Ca2+ response with marked elevations of [Ca2+ ]cyt over time. In Ca2+ free medium, cells still showed an initial small elevation of [Ca2+ ]cyt which was magnified on addition of Ca2+ to the medium. Washing the cells after DDE treatment failed to remove the elevated [Ca2+]cyt and oscillations both of which were eliminated by addition of EGTA. Adenosine triphosphate (ATP) also induced [Ca2+]cyt elevations and oscillations and these effects were potentiated on the addition of DDE. Follicle stimulating hormone (FSH) induced transient [Ca2+]cyt elevations in 3 of 7 experiments. Conclusions: These results suggest that DDE at concentrations normally found in human tissues is an environmental toxicant and can induce elevations in [Ca2+]cyt. Our data therefore highlight a novel mechanism through which DDE can alter endocrine homeostasis and act as an endocrine toxicant.