Environmental Research 94 (2004) 38–46 DDT induces apoptosis in human mononuclear cells in vitro and is associated with increased apoptosis in exposed children Iva´n N. Pe´rez-Maldonado, a Fernando Dı´az-Barriga, a Hortensia de la Fuente, a Roberto Gonza´lez-Amaro, a Jaqueline Caldero´n, a and Leticia Ya´ n˜ ez b, a Facultad de Medicina, Universidad Auto´noma de San Luis Potosı´ , San Luis Potosı´ , S.L.P., Mexico b Laboratorio de Toxicologia Ambiental, Universidad Auto´noma de San Luis Potosı´ , Avenida Venustiano Carranza 2405, San Luis Potosı´ , S.L.P. 78210, Mexico Received 16 October 2002; received in revised form 20 May 2003; accepted 23 May 2003 Abstract The aim of the present work was to investigate whether DDT and its metabolites are able to induce apoptosis of human peripheral blood mononuclear cells (PBMC) both in vitro and in vivo. Cells isolated from healthy individuals were incubated in the presence of increasing concentrations of p 0 p-DDT, p 0 p-DDE, or p 0 p-DDD (0–150 mg/mL) for different intervals. Apoptosis was then determined by flow cytometry (DNA cell content analysis) and fluorescence microscopy (Hoechst staining). A significant level of apoptosis was induced by DDT, DDD, and DDE at 80 mg/mL compared to controls, reaching a maximum effect at 100 mg/mL. We began to detect apoptosis at 12 h, with a maximum effect at 24 h of incubation. These results were confirmed using the TUNEL assay in cells treated with the three compounds tested as well as with o 0 p-DDT at 100 mg/mL and 24 h of incubation. Our data demonstrate that DDT and its metabolites are able to induce apoptosis of human PBMC in vitro. Therefore, we performed a preliminary study in children exposed to this insecticide. When compared to a control population, the exposed children had higher levels of DDT, DDD, and DDE in blood and also had a higher frequency of apoptosis. In the exposed children, a weak positive association was found between the frequency of apoptosis and the exposure to DDT and DDE. Our results showed that more studies are needed in people exposed to DDT, as apoptosis may cause serious public health effects such as immunosuppression. r 2003 Elsevier Inc. All rights reserved. Keywords: DDT; DDE; Malaria; Apoptosis; Children 1. Introduction DDT [1,1,1-trichloro-2,2-bis(p-chlorophenyl)ethane] was a widely used insecticide in the malaria control program in Mexico (Stapleton, 1998). Thus, in malar- ious areas, due to the amount that was applied and as a result of its environmental persistence, many individuals are heavily exposed to DDT and its metabolites, DDD and DDE (Dı´az-Barriga et al., 2002; Waliszewski et al., 1999a,b; Yan˜ez et al., 2002). In these areas, human exposure to DDT could be considered a major public health issue. However, DDT-induced health effects are still debated, as human data of DDT toxicity are somewhat limited. The central nervous system seems to be a major DDT target organ in humans and animals (ATSDR, 2002). Nevertheless, it has also been reported that DDT induces immunodeficiency in rodents (Banerjee, 1987, 1999). Although the mechanism of immunotoxicity of DDT has not been determined, it has been found that p 0 p-DDT is able to induce apoptosis of rat thymocytes at a concentration of 7 mg/mL (Teoburi et al., 1998). Furthermore, in vivo experiments in rats have demon- strated that p 0 p-DDT treatment for 10 days causes a dose-dependent thymic atrophy reaching 31% at 100 mg/kg body weight (Teoburi et al., 1998). Apoptosis has also been found in murine embryos exposed in vitro to o 0 p-DDT at 0.1 mg/mL; however, in this case the embryos were unresponsive to similar concentrations of p 0 p-DDT (Greenlee et al., 1999). Taking into account that in several countries people are still exposed to DDT, the apoptotic properties of ARTICLE IN PRESS  Corresponding author. Fax: +52-48-262-354. E-mail address: lyanez@uaslp.mx (L. Ya´n˜ez). 0013-9351/03/$-see front matter r 2003 Elsevier Inc. All rights reserved. doi:10.1016/S0013-9351(03)00112-9