In vitro metabolic activation of the pulmonary toxin, 4-ipomeanol, in nonciliated bronchiolar epithelial (Clara) and alveolar type II cells isolated from rabbit lung

Abstract

The metabolism and covalent binding of 4-ipomeanol (IPO), a pulmonary toxin, were investigated in pulmonary cells isolated from rabbit. 3H-labeled IPO was incubated with freshly, isolated, intact alveolar type II cells (83% purity), nonciliated bronchiolar epithelial (Clara) cells (77% purity) and alveolar macrophages (greater than 90% purity). Covalent binding of radioactive material to type II and Clara cells was observed by autoradiography and by a biochemical method. IPO binding to cells was almost totally prevented by 1 mM piperonyl butoxide, an inhibitor of the cytochrome P-450-dependent metabolism of IPO. No covalent binding was observed with alveolar macrophages in the presence or absence of piperonyl butoxide. The maximal rates of enzyme-mediated covalent binding of IPO to protein were greater in the Clara cells (135 pmol/10(6) cells/min) than in the type II cells (13 pmol/10(6) cells/min). Incubation of either sonicated Clara or type II cell fractions with [3H]IPO, glutathione and NADPH (20 min, 37 degrees C) resulted in the formation of two distinct radiolabeled glutathione conjugates.