J. Biochem, 1994, Vol. 115, No. 2 238-244
© 1994 Japanese Biochemical Society
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Phosphatidic Acid Induces the Release of ß-Glucuronidase but Not Lactoferrin from Electropermeabilized Human Neutrophils1
*Departments of Biochemistry, Kyushu University School of Medicine, 3-1-1 Maidashi Higashi-ku, Fukuoka 812
**Research Institute for Diseases of the Chest, Kyushu University School of Medicine, 3-1-1 Maidashi Higashi-ku, Fukuoka 812
***Pharmacology, Kyushu University School of Medicine, 3-1-1 Maidashi Higashi-ku, Fukuoka 812
3 To whom correspondence should be addressed
We studied the degranulation reaction of electropermeabilized human neutrophils induced by 1,2-didecanoyl-3-sn-phosphatidic acid (PA10). PA10 dose-dependently induced the release of ß-glucuronidase, an enzyme of azurophil granules, but did not induce the release of lactoferrin, a protein of specific granules. The enzyme release by PA10 absolutely required Ca2+, ATP, and Mg2+ and the concentrations for the half-maximal response were 2.5 µM, 60 µM, and 0.25 mM, respectively. Although Ca2+ alone at concentrations higher than 10 µM induced the release of both ß-glucuronidase and lactoferrin, the extents of the release were far less than that of the ß-glucuronidase release by PA10. Phorbol myristate acetate (PMA) and 1-oleoyl-2-acetyl-sn-glycerol induced the release of lactoferrin alone at concentrations of Ca2+ below 0.5 µM while they induced the release of both ß-glucuronidase and lactoferrin at higher Ca2+ concentrations, indicating that the degranulation induced by PA10 is not mediated by diacylglycerol which might be formed from PA. The degranulation reactions induced by PA10 and PMA were dose-dependently inhibited by staurosporine and calphostin C, protein kinase C inhibitors, although no direct activation of protein kinase C by PA10 was observed. The extent of the ß-glucuronidase release by PA10 was not enhanced by the addition of PMA. Propranolol, which inhibits protein kinase C as well as phosphatidic acid phosphohydrolase, strongly inhibited the degranulation reactions induced by PA10 and PMA. Ethanol, a metabolic modulator of phospholipase D, and cyclic AMP did not affect the degranulation reactions by PMA and PA10. These observations suggest that selective enzyme release from azurophil granules of the permeabilized neutrophils might be induced through the activation of protein kinase C or some other protein kinase sensitive to the protein kinase C inhibitors and that phospholipase D may not be involved in the PMA-induced release.
1This study was supported in part by a Grant-in-Aid for Scientific Research from the Ministry of Education, Science and Culture of Japan.
2Present address Graduate School of Health and Nutrition Sciences, Nakamura Gakuen College, 5-7-1 Befu, Jonan-ku, Fukuoka 814-01.
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