PULMONARY PROINFLAMMATORY GENE INDUCTION FOLLOWING ACUTE EXPOSURE TO RESIDUAL OIL FLY ASH: ROLES OF PARTICLE-ASSOCIATED METALS

Abstract

Residual oil fly ash (ROFA), an emission source particulate, has been shown to induce acute lung injury and fibrosis in the rat. However, the mechanism(s) and the identities of various inflammatory mediators induced by ROFA are not known. Also the extent to which ROFA-associated metals contribute to proinflammatory gene induction is yet to be determined. To examine the mechanism of ROFA-induced lung injury, male SpragueDawley rats (60 days old) were intratracheally instilled with 0.3 ml of either acidified saline (brought to pH 2.5 using H2 SO4, similar to the pH of the ROFA suspension in saline), ROFA (2.5 mg/rat in saline, yields pH of 2.5), or predominant ROFA-associated metals such as Fe2 (SO4)3 (Fe, 0.54 mol/rat), VSO4 (V, 1.7 mol/rat), and NiSO4 (Ni, 1.0 mol/rat), individually or as a mixture (Fe + V + Ni). The quantity of metals instilled reflected the amount present in the leachable material of ROFA. Histopathological findings indicated marked and progressive acute focal lung injury characterized by inflammation, edema, alveolar cell hyperplasia, thickening of the alveolar and airway walls, and bronchiolar secretory cell hypertrophy following ROFA exposure. The metal mixture induced similar pathology; however, the severity of lesions appeared less pronounced than with ROFA. Ni by itself caused the most severe damage, hemorrhage and inflammation, while V and Fe alone at a concentration present in ROFA produced less severe pathology. ROFAinstilled rats showed denudation of airway epithelium at 3 h that was less severe with individual metals and the metal mixture. Focal alveolar fibrosis was found in the case of ROFA and the metal mixture, and alveolar wall hyperplasia in the case of Ni, which predominated at 96 h postinstillation. Results obtained from semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) analyses indicated that interleukin 1 (IL-1) and IL-5 were induced as early as 3 h and returned to control levels by 24 h following ROFA exposure. The metal mixture as well as Fe and V produced similar induction of IL1 and IL-5 at 3 h. Although the extent of IL-1 and IL-5 induction by Ni was similar to ROFA at 3 h, unlike ROFA or the other metals, this induction persisted for up to 96 h postexposure. Macrophage inflammatory protein 2 (MIP-2) and IL-6 mRNA were induced by ROFA, the metal mixture, and individual metals, as early as 3 h after instillation. Both cytokines remained elevated especially in ROFA-, metal mixture-, and Ni-instilled rats at 24 h. MIP-2 expression remained elevated through 96 h in Ni-instilled rats. Surprisingly, tumor necrosis factor (TNF-) mRNA was not affected by ROFA or metals at any time points examined. Vascular cell adhesion molecule-1 (VCAM-1) expression was increased slightly by ROFA, the metal mixture, and Ni, but not by Fe or V, at 3 h and returned to control levels by 24 h postexposure. E-selectin mRNA increased following ROFA, metal mix, V, and Ni instillation at 3 h, again returning to control levels by 24 h postexposure. These studies suggest that ROFA could induce a number of proinflammatory cytokine genes very early and that much of this induction was due to ROFA-associated metals. Overall, the persistency and the extent of cytokine induction over 96 h by ROFA and metals were in the order of Ni > ROFA metal mix V Fe. The persistency of cytokine induction rather than degree was more closely associated with the histopathology induced by ROFA and associated metals.