Fig. 1.

Digitonin-mediated release of cytosolic and mitochondrial enzymes from isolated rat hepatocytes. (A) Hepatocytes (2 × 10⁶ cells/ml) were incubated at room temperature in ICB and treated with different concentrations of digitonin for 10 min. A portion of digitonin-treated cells was taken to assess trypan blue labeling (TB), and the rest was sedimented by centrifugation at 14,000 rpm for 60 s for enzymatic analysis of the supernatants. Data are means from 6 experiments. (B) Hepatocytes (2 × 10⁶ cells/ml) treated with 0, 8 or 80 μM of digitonin for 10 min in ICB were centrifuged, and supernatants were subjected to Western blotting for cytochrome c, as described in Materials and methods. Lanes are: first lane (far left), molecular weight markers (MWM); second, third and fourth lanes, supernatants after 0, 8 and 80 μM digitonin; last lane (far right), 100 ng cytochrome c, (Cyt c). Representative Western blots are shown from three or more independent hepatocyte incubations yielding similar results.