Changes of phospholipase D activity of rat peritoneal mast cells in degranulation

To study the changes of phospholipase D (PLD) activity of actively sensitized rat peritoneal mast cells (RPMC) in degranulation.

Degranulation of RPMC was determined by measurement of beta-hexosaminidase release. PLD activity assay was carried out by measurement of PLD product, choline, with chemiluminescent oxidation of luminol.

Actively sensitized RPMC challenged with ovalbumin (0.5-8 mg/L for 120 s, 4 mg/L for 15-120 s) resulted in significant activation of PLD accompanied with the increment of beta-hexosaminidase release. PLD activity of sensitized RPMC was increased by more than 2-fold compared with that of unsensitized RPMC which contained low levels of PLD activity [(35+/-13) pmol choline/min in 10(6)cells], but beta-hexosaminidase releases of the sensitized cells were as low as spontaneous releases. After challenge with ovalbumin 4 mg/L for 120 s, PLD activity of sensitized RPMC was increased to (155+/-43) pmol choline/min in 10(6)cells and beta-hexosaminidase release was also elevated significantly (4.5-fold of spontaneous release, n=6, P<0.05). When unsensitized RPMC were stimulated with antigen, PLD activity and beta-hexosaminidase release of the cells were hardly changed. Sensitized RPMC were treated with 1 % 1-butanol or 2,3- disphosphoglycerate 10 mmol/L before challenge with ovalbumin, these drugs induced an inhibition of PLD activity and a reduction of beta-hexosaminidase release to basal level. 1-Butanol 0.1 % also worked.

Phospholipase D plays an important role in the regulation of beta-hexosaminidase release in actively sensitized rat peritoneal mast cells.