title: Organochlorine insecticides induce NADPH oxidase-dependent reactive oxygen species in human monocytic cells via phospholipase A2/arachidonic acid.
abstract: Bioaccumulative organohalogen chemicals, such as organochlorine (OC) insecticides, have been increasingly associated with disease etiology; however, the mechanistic link between chemical exposure and diseases, such as atherosclerosis, cancer, and diabetes, is complex and poorly defined. Systemic oxidative stress stemming from OC exposure might play a vital role in the development of these pathologies. Monocytes are important surveillance cells of the innate immune system that respond to extracellular signals possessing danger-associated molecular patterns by synthesizing oxyradicals, such as superoxide, for the purpose of combating infectious pathogens. We hypothesized that OC chemicals can be toxic to monocytes because of an inappropriate elevation in superoxide-derived reactive oxygen species (ROS) capable of causing cellular oxidative damage. Reactive oxyradicals are generated in monocytes in large part by NADPH oxidase (Nox). The present study was conducted to examine the ability of two chlorinated cyclodiene compounds, trans-nonachlor and dieldrin, as well as p,p'-DDE, a chlorinated alicyclic metabolite of DDT, to stimulate Nox activity in a human monocytic cell line and to elucidate the mechanisms for this activation. Human THP-1 monocytes treated with either trans-nonachlor or dieldrin (0.1-10 μM in the culture medium) exhibited elevated levels of intracellular ROS, as evidenced by complementary methods, including flow cytometry analysis using the probe DCFH-DA and hydroethidine-based fluorometric and UPLC-MS assays. In addition, the induced reactive oxygen flux caused by trans-nonachlor was also observed in two other cell lines, murine J774 macrophages and human HL-60 cells. The central role of Nox in OC-mediated oxidative stress was demonstrated by the attenuated superoxide production in OC-exposed monocytes treated with the Nox inhibitors diphenyleneiodonium and VAS-2870. Moreover, monocytes challenged with OCs exhibited increased phospho-p47(phox) levels and enhanced p47(phox) membrane localization compared to that in vehicle-treated cells. p47(phox) is a cytosolic regulatory subunit of Nox, and its phosphorylation and translocation to the NOX2 catalytic subunit in membranes is a requisite step for Nox assembly and activation. Dieldrin and trans-nonachlor treatments of monocytes also resulted in marked increases in arachidonic acid (AA) and eicosanoid production, which could be abrogated by the phospholipase A2 (PLA2) inhibitor arachidonoyltrifluoromethyl ketone (ATK) but not by calcium-independent PLA2 inhibitor bromoenol lactone. This suggested that cytosolic PLA2 plays a crucial role in the induction of Nox activity by increasing the intracellular pool of AA that activates protein kinase C, which phosphorylates p47(phox). In addition, ATK also blocked OC-induced p47(phox) serine phosphorylation and attenuated ROS levels, which further supports the notion that the AA pool liberated by cytosolic PLA2 is responsible for Nox activation. Together, the results suggest that trans-nonachlor and dieldrin are capable of increasing intracellular superoxide levels via a Nox-dependent mechanism that relies on elevated intracellular AA levels. These findings are significant because chronic activation of monocytes by environmental toxicants might contribute to pathogenic oxidative stress and inflammation.
authors: Department of Basic Sciences, Center for Environmental Health Sciences, College of Veterinary Medicine, Mississippi State University, P.O. Box 6100, Mississippi State, Mississippi 39762, United States.
authors: Lee C
authors: LC
authors: Mangum
authors: Department of Basic Sciences, Center for Environmental Health Sciences, College of Veterinary Medicine, Mississippi State University, P.O. Box 6100, Mississippi State, Mississippi 39762, United States.
authors: Abdolsamad
authors: A
authors: Borazjani
authors: Department of Basic Sciences, Center for Environmental Health Sciences, College of Veterinary Medicine, Mississippi State University, P.O. Box 6100, Mississippi State, Mississippi 39762, United States.
authors: John V
authors: JV
authors: Stokes
authors: Department of Basic Sciences, Center for Environmental Health Sciences, College of Veterinary Medicine, Mississippi State University, P.O. Box 6100, Mississippi State, Mississippi 39762, United States.
authors: Anberitha T
authors: AT
authors: Matthews
authors: Department of Basic Sciences, Center for Environmental Health Sciences, College of Veterinary Medicine, Mississippi State University, P.O. Box 6100, Mississippi State, Mississippi 39762, United States.
authors: Jung Hwa
authors: JH
authors: Lee
authors: Department of Basic Sciences, Center for Environmental Health Sciences, College of Veterinary Medicine, Mississippi State University, P.O. Box 6100, Mississippi State, Mississippi 39762, United States.
authors: Janice E
authors: JE
authors: Chambers
authors: Department of Basic Sciences, Center for Environmental Health Sciences, College of Veterinary Medicine, Mississippi State University, P.O. Box 6100, Mississippi State, Mississippi 39762, United States.
authors: Matthew K
authors: MK
authors: Ross
mesh_terms: D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism*; D002460:Cell Line; D006801:Humans; D006843:Hydrocarbons, Chlorinated / Q000633:toxicity*; D007306:Insecticides / Q000633:toxicity*; D051379:Mice; D009000:Monocytes / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D019255:NADPH Oxidases / Q000378:metabolism*; D018384:Oxidative Stress; D054467:Phospholipases A2 / Q000378:metabolism*; D017382:Reactive Oxygen Species / Q000378:metabolism*
chemical_list: D006843:Hydrocarbons, Chlorinated; D007306:Insecticides; D017382:Reactive Oxygen Species; D016718:Arachidonic Acid; D019255:NADPH Oxidases; D054467:Phospholipases A2
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