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

Residual oil fly ash (ROFA), an emission source particulate, has been shown to induce acute icing injury and fibrosis in the rat. However, t he mechanism(s) and the identities of various inflammatory mediators i nduced by ROFA are not known. Also the extent to which ROFA-associated metals contribute to proinflammatory gene induction is yet to be dete rmined. To examine the mechanism of ROFA-induced lung injury, male Spr ague-Dawley rats (60 days old) were intratracheally instilled with 0.3 ml of either acidified saline (brought to pH 2.5 using H2SO4, similar to the pH of the ROFA suspension in saline), ROFA (2.5 mg/rat in sali ne, yields pH of 2.5), or predominant ROFA-associated metals such as F e-2(SO4)(3) (Fe, 0.54 mu mol/rat), VSO4 (V, 1.7 mu mol/rat), and NiSO4 (Ni, 1.0 mu mol/rat), individually or as a mixture (Fe + V + Ni). The quantity of metals instilled reflected the amount present in the leac hable material of ROFA. Histopathological findings indicated marked an d progressive acute focal lung injury characterized by inflammation, e dema, alveolar cell hyperplasia, thickening of the alveolar and airway walls, and bronchiolar secretory cell hypertrophy following ROFA expo sure. The metal mixture induced similar pathology; however, the severi ty of lesions appeared less pronounced than with ROFA. Ni by itself ca used the most severe damage, hemorrhage and inflammation, while V and Fe alone at a concentration present in ROFA produced less severe patho logy. ROFA-instilled rats showed denudation of airway epithelium at 3 h that was less severe with individual metals and the metal mixture. F ocal alveolar fibrosis was round in the case of ROFA and the metal mix ture, and alveolar wall hyperplasia in the case of Ni, which pre domin ated at 96 h postinstillation. Results obtained from semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) analyses indi cated that interleukin 1 beta (IL-1 beta) and IL-5 were induced as ear ly as 3 h and returned to control levels by 24 h following ROFA exposu re. The metal mixture as well as Fe and V produced similar induction o f IL-1 beta and IL-5 at 3 h. Although the extent of IL-1 beta 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. Macropha ge 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 instil lation. Both cytokines remained elevated especially in ROFA-, metal mi xture-, and Ni-instilled rats at 24 h. MIP-2 expression remained eleva ted through 96 h in Ni-instilled rats. Surprisingly, tumor necrosis fa ctor alpha (TNF-alpha) mRNA was not affected by ROFA or metals at any time points examined. Vascular cell adhesion molecule-1 (VCAM-1) expre ssion was increased slightly by ROFA, the metal mixture, and Ni, but n ot by Fe or V, at 3 h and returned to control levels by 24 h postexpos ure, E-selectin mRNA increased following ROFA, metal mix, V, and Ni in stillation at 3 h, again returning to control levels by 24 h postexpos ure. These studies suggest that ROFA could induce a number of proinfla mmatory 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 greater than or equal to metal mix greater than or equal to V greater than or equal to Fe. The persistency cytokine induction r ather than degree was more closely associated with the histopathology induced by ROFA and associated metals.