Autonomous regulation of inducible nitric oxide synthase and cytochrome P450 2E1-mediated oxidative stress in maneb- and paraquat-treated rat polymorphs.

Maneb (MB)- and paraquat (PQ)-induced oxidative stress in rat polymorphonuclear leukocytes (PMNs) is regulated in parallel by cytochrome P450 2E1 (CYP2E1) and inducible nitric oxide synthase (iNOS). However, mechanism underlying their regulation is not yet understood. The study investigated the role of nuclear factor- kappa B (NF-κB) and mitogen-activated protein kinase/extracellular signal regulated kinase/protein kinase C (MEK/ERK/PKC) pathway in the regulation of iNOS- and CYP2E1-induced oxidative stress in PMNs. MB + PQ-induced changes in nitrite content, lipid peroxidation (LPO), iNOS expression/activity and inflammatory mediators were alleviated by aminoguanidine (AG), an iNOS inhibitor, without any change in CYP2E1. Alternatively, diallyl sulphide (DAS), a CYP2E1 inhibitor, rescued from MB + PQ-induced changes in CYP2E1 activity/expression, free radical generation, superoxide dismutase (SOD) activity, LPO and pro-inflammatory cytokines without any alterations in nitrite content and iNOS activity/expression. Pyrrolidine dithiocarbamate (PDTC), NF-κB inhibitor, did not alter CYP2E1 but mitigated free radical generation, SOD activity, LPO, nitrite content, iNOS activity/expression and levels of pro-inflammatory cytokines (tumor necrosis factor-α, interleukine-1β and interleukine-4). Ex-vivo treatment with MEK inhibitor (PD98059), ERK1/2 inhibitor (AG126) or PKC inhibitor (rottlerin) ameliorated MB + PQ-induced increase in free radical generation and CYP2E1 activity/expression in PMNs. While PD98059 and AG126 abated MB + PQ-induced increase in ERK1/2, PKC-α/δ and CYP2E1 levels, rottlerin restored PKC-α/δ and CYP2E1 towards normalcy without affecting ERK1/2 level in MB + PQ-treated group. The results suggest that iNOS and CYP2E1 contributing to MB + PQ-induced oxidative stress in rat PMNs exhibit differential regulatory mechanisms. The inflammatory mediators regulate iNOS expression while CYP2E1 expression is triggered via MEK-ERK1/2-PKC pathway.