Morphologic changes and covalent binding of 1,1-dichloroethylene in Clara and alveolar type II cells isolated from lungs of mice following in vivo administration.

We have investigated the metabolism and covalent binding of 1,1-dichloroethylene (1,1-DCE) in isolated unseparated lung cells and in enriched fractions of Clara and alveolar type II cells from mice. Lung cells from control mice separated by centrifugal elutriation were viable and metabolically active as assessed by measurements of 7-ethoxycoumarin deethylase, NADPH cytochrome c reductase, and glutathione S-transferase activities, which were highest in fractions enriched in Clara cells. Mice were treated with [14C]1,1-DCE (125 mg/kg; 20 microCi/kg) in vivo and, 1 hr later, lung cells were isolated and binding of [14C]1,1-DCE determined. Covalent binding was highest in the Clara cell fraction (480 +/- 205 pmol/10(6) cells; 41% Clara cell purity) when compared to the levels present in the fractions containing type II cells (126 +/- 63 pmol/10(6) cells; 51% type II cell purity) and mixed cells from whole lung (29 +/- 13 pmol/10(6) cells). Ultrastructurally, alveolar type II cells from lungs of control and 1,1-DCE-treated mice exhibited normal morphology with well-preserved lamellar bodies. Whereas Clara cells isolated from lungs of control mice appeared structurally unimpaired, those from the lungs of 1,1-DCE-treated mice displayed severe damage and disruption of cellular organelles. The results of these experiments demonstrate the highest binding of [14C]1,1-DCE-metabolite(s) in Clara cells, whereas significantly lower binding was found in both alveolar type II and unseparated lung cells. The substantial binding of [14C]1,1-DCE in Clara cells correlated positively with the high monooxygenase capacity and the preferential damage sustained by this cell population.

Biotransformation activities in Clara and alveolar type II cells isolated from hamster lungs.

A method is described for isolation of enriched preparations of nonciliated bronchiolar epithelial (Clara) and alveolar type II cells from hamster lungs. Clara cells were enriched to 48% (1.7 +/- 0.7 x 10(5) cells per hamster), and type II cells to 86% (3.4 +/- 1.7 x 10(6) cells per hamster). Analysis of biotransformation activities indicated concentration of 7-ethoxycoumarin deethylase activity in Clara cells, whereas glutathione S-transferase was more widely distributed among cell types, with highest activity in type II cells. The significance of differential cellular localization of activation and detoxification activities to susceptibility to pulmonary toxicants, is discussed.

Isolation of nonciliated bronchiolar epithelial (Clara) cells and alveolar type II cells from mouse lungs.

A method is described for the isolation of alveolar type II cells and nonciliated bronchiolar epithelial (Clara) cells from mouse lungs. Following digestion of lung tissue with Sigma type I protease, viable cells were isolated to 65% purity for type II cells (6.4 +/- 1.5 X 10(5) cells/mouse) and 55-60% purity for Clara cells (2.6 +/- 0.9 X 10(5) cells/mouse). Viability, as assessed by trypan blue exclusion, was routinely greater than 90% in all enriched cell fractions. Although minor mitochondrial changes occurred during isolation, the morphology of the cells showed good preservation, as revealed by electron microscopy. The isolated cells were found to be metabolically active, as indicated by the presence of 7-ethoxycoumarin deethylase (a cytochrome P-450-mediated activity). The highest activity of this enzyme (278 +/- 116 pmol.min-1.mg protein-1) was found in the fraction enriched in Clara cells. The results indicate that this method produces viable cell populations that can be of value in investigations of the cellular distribution of lung metabolism activities.