Involvement of NADPH Oxidase and iNOS in Rodent Pulmonary Cytokine Responses to Urban Air and Mineral Particles

Abstract

We have investigated the potential of two complex mineral particles (feldspar and mylonite), quartz (Min-U-Sil), and suspended particulate matter (SRM-1648) (SPM) from urban air to induce inflammatory cytokine responses in primary rat alveolar type 2 cells and alveolar macrophages, and the involvement of cellular formation of free radicals in these responses. All particle types induced an increased release of interleukin (IL)-6 and macrophage inflammatory protein (MIP)-2 from type 2 cells. Diphenyleneiodonium chloride (DPI), a selective inhibitor of NADPH-oxidase, reduced the IL-6 and MIP-2 responses to quartz, SPM and mylonite. N-(3-[Aminomethyl] benzyl) acetamidine (1400W), a selective inhibitor of inducible nitric oxide synthase (iNOS), significantly reduced the Il-6 response to SPM and feldspar in the type 2 cells. The macrophages displayed significantly increased TNF-α and MIP-2 release upon exposure to quartz or SPM. Here, DPI significantly reduced the tumor necrosis factor (TNF)-α and MIP-2 responses to quartz, and the MIP-2 response to SPM. No significant effect of 1400 W was detected in the alveolar macrophages. The role of particle-induced cellular generation of free radicals in lung cytokine responses was further elucidated in mice that lacked either NADPH-oxidase or iNOS as well as in wild-type (wt) mice. All particles were able to elicit increased cytokine levels in the bronchoalveolar lavage (BAL) fluid of the mice, although the levels depended on particle type. The NADPH-oxidase knockout (KO) mice demonstrated a significantly lower IL-6 and MIP-2 responses to SPM compared to their respective wt mice. The iNOS KO mice displayed significantly reduced IL-6, TNF-α, and MIP-2 responses to SPM. The overall results indicate the involvement of cellular free-radical formation in the pulmonary cytokine responses to particles of varying composition.