Fig. 12

Effect of the addition of trypsin/salt extract prepared from detergent-insoluble complexes on the stimulation of glucose transport by PIG-P in reconstituted and untreated adipocytes.

Isolated rat adipocytes, which had been treated with 100 µg/ml trypsin and 0.5 M NaCl (squares, triangles, exes) or left untreated (circles, diamonds, asterisks) were supplemented with the indicated amounts of normal or 10-fold concentrated trypsin/salt extract (squares, circles) or NEM-treated trypsin/salt extract (triangles, diamonds) or trypsin extract (exes, asterisks), which had been prepared from detergent-insoluble complexes as described in Materials and Methods. The pretreated adipocytes, which had been reconstituted and washed twice by flotation (squares, triangles, exes), as well as the untreated cells (circles, diamonds, asterisks) were incubated with 2.5 µm PIG-P (20 min, 37^°C) and subsequently assayed for glucose transport. Each point represents the mean ± SD of four different reconstitutions or incubations using different adipocyte preparations, with extract prepared in triplicate and transport measurements in quadruplicate. The stimulation of glucose uptake was calculated as the difference between the presence and absence of 2.5 µm PIG-P in normal adipocytes and set at 100%, which corresponded to 58 ± 11% of the maximal PIG-P action (at 5 µm) and 43 ± 8% of the maximal insulin action (at 5 nM). Basal glucose uptake in normal adipocytes was set at 0%.