Glycerol-releasing activity of histamine-sensitizing factor of Bordetella pertussis for rat adipocytes in vitro.

Histamine-sensitizing factor (HSF) purified from Bordetella pertussis induced specifically the release of glycerol from rat epididymal adipocytes in vitro. The most sensitive and reproducible results were obtained by using 1 to 2 x 10(5) adipocytes/tube from rats weighing 150 to 200 g, and by incubation at 37 C for 180 min. After a lag period of about 60 min, HSF-treated adipocytes released glycerol in increasing amounts between 60 and 240 min, depending on the dose of HSF. A close correlation between the glycerol-releasing (GR) activity of HSF for adipocytes and histamine-sensitizing or leukocytosis-promoting activity in mice was observed. The GR activity was inactivated by heating at 56 C for 60 min, 63 C for 30 min or 96 C for 10 min. The adipocytes washed out with a Krebs-Ringer bicarbonate buffer immediately after being exposed to HSF for 1 to 3 min manifested about 75% of the total GR activity induced by HSF, and those washed out after being exposed for 30 min or longer had full activity. Anti-HSF serum neutralized the activity when it was added to adipocytes simultaneously with HSF, but did not when it was added 30 min after being exposed to HSF. By using both native and 125 I-labeled HSF, the ratio of binding of HSF to adipocytes was estimated to be 10 to 15% of the total HSF per 2 x 10(5) cells/tube, and to be about 1,000 molecules of HSF per cell to induce the release of glycerol. The GR activity induced with 10 ng of HSF was inhibited by addition of insulin at a dose of over 1 micro IU/tube, but not by concanavalin A.