| Abstrakt: |
The proteolytic digestion of lung tissue is a necessary prerequisite for the isolation of relatively pure populations of type II alveolar cells. The choice of digestion conditions can have a profound effect on the yield and metabolic integrity of such cells. The specific effects of various proteases on isolation of type II cells from adult rabbits were investigated in a systematic way; maximal numbers of cells were obtained using a mixture of purified elastase (0.3 mg/ml), purified trypsin (0.025 mg/ml) and inflation of intact lung with the protease mixture. Under such conditions, the average yield of cells from an adult rabbit was 10±2.1×107, of which 80–90% were type II cells. Lower yields were obtained by exposing lung tissue to protease after mincing, even if protease concentrations were increased 2–4 fold. By virtue of the lowered protease concentrations, cellular damage as measured by effects on cellular enzyme activities was minimal. NADPH cytochromecreductase activity was not reduced by isolation conditions. In addition, no significant enhanced release of activity into the 150,000 supernatant was observed. The isolated cells were enriched 2–3 fold for the enzymes of phospholipid synthesis compared to whole lung and alveolar macrophages. |
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