Phorbol myristate acetate stimulates formation of diphosphoinositide in rat mast cell granules.
Rat mast cell granules were obtained by homogenization of highly purified rat mast cells and isolated in a Percoll gradient. Diphosphoinositide (DPI) synthesis in rat mast cell granules was assayed by measuring the incorporation of 32P from [gamma 32P]ATP into DPI in the absence of exogenous phosphatidylinositol (PI). Lipids were isolated with methanol/chloroform/HCl and were separated by thin-layer chromatography on oxalic acid impregnated silica gel plates. DPI areas were identified by staining with iodine, scraped and measured for 32P radioactivity. The addition of phorbol myristate acetate (PMA) to the granules caused an increase of 32P incorporation from [gamma 32P]ATP in the DPI fraction, which can be catalyzed by PI kinase. This effect of PMA in the DPI synthesis was dose dependent and maximal effects were observed at 10 ng/ml.
Kurosawa
M
M
First Department of Internal Medicine, Gunma University School of Medicine, Maebashi, Japan.
Okayama
Y
Y
Kobayashi
S
S
D000818:Animals; D003594:Cytoplasmic Granules / Q000378:metabolism; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D066298:In Vitro Techniques; D008297:Male; D008407:Mast Cells / Q000378:metabolism*; D010703:Phorbol Esters / Q000494:pharmacology; D010716:Phosphatidylinositols / Q000378:metabolism*; D010766:Phosphorylation; D051381:Rats; D011919:Rats, Inbred Strains; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D010703:Phorbol Esters; D010716:Phosphatidylinositols; D013755:Tetradecanoylphorbol Acetate; D004121:Dimethyl Sulfoxide