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PMID: 1864964 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Phosphatidic acid as a second messenger in human polymorphonuclear leukocytes. Effects on activation of NADPH oxidase.

The Journal of clinical investigation ·Vol. 88 ·No. 2 ·1991-08-00 ·Pages 531-9

Agwu DE, McPhail LC, Sozzani S, Bass DA, McCall CE

Abstract

Receptor-mediated agonists, such as FMLP, induce an early, phospholipase D (PLD)-mediated accumulation of phosphatidic acid (PA) which may play a role in the activation of NADPH oxidase in human PMN. We have determined the effect of changes in PA production on O2 consumption in intact PMN and the level of NADPH oxidase activity measured in a cell-free assay. Pretreatment of cells with various concentrations of propranolol enhanced (less than or equal to 200 microM) or inhibited (greater than 300 microM) PLD-induced production of PA (mass and radiolabel) in a manner that correlated with enhancement or inhibition of O2 consumption in PMN stimulated with 1 microM FMLP in the absence of cytochalasin B. The concentration-dependent effects of propranolol on FMLP-induced NADPH oxidase activation was confirmed by direct assay of the enzyme in subcellular fractions. In PA extracted from cells pretreated with 200 microM propranolol before stimulation with 1 microM FMLP, phospholipase A1 (PLA1)-digestion for 90 min, followed by quantitation of residual PA, showed that a minimum of 44% of PA in control (undigested) sample was diacyl-PA; alkylacyl-PA remained undigested by PLA1. Propranolol was also observed to have a concentration-dependent enhancement of mass of 1,2-DG formed in PMN stimulated with FMLP. DG levels reached a maximum at 300 microM propranolol and remained unchanged up to 500 microM propranolol. However, in contrast to PA levels, the level of DG produced did not correlate with NADPH oxidase activation. Exogenously added didecanoyl-PA activated NADPH oxidase in a concentration-dependent manner (1-300 microM) in a reconstitution assay using membrane and cytosolic fractions from unstimulated PMN. In addition, PA synergized with SDS for oxidase activation. Taken together, these results indicate that PA plays a second messenger role in the activation of NADPH oxidase in human PMN and that regulation of phospholipase D is a key step in the activation pathway.

MeSH Terms
Diglycerides/metabolism Enzyme Activation/drug effects Humans N-Formylmethionine Leucyl-Phenylalanine/pharmacology NADH, NADPH Oxidoreductases/metabolism NADPH Oxidases Neutrophils/enzymology Phosphatidic Acids/physiology Phospholipase D/metabolism Propranolol/pharmacology Second Messenger Systems/physiology
Chemicals
Diglycerides Phosphatidic Acids N-Formylmethionine Leucyl-Phenylalanine Propranolol NADH, NADPH Oxidoreductases NADPH Oxidases Phospholipase D
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Agwu D E
Department of Medicine, Wake Forest University Medical Center, Winston-Salem, North Carolina 27103.
McPhail L C
Sozzani S
Bass D A
McCall C E
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Article Info
Journal
The Journal of clinical investigation
Abbr.
J Clin Invest
ISSN
0021-9738
DOI
10.1172/JCI115336
Published
1991-08-00
Pages
531-9
Language
English
Region
United States
NLM ID
7802877
PMCID
PMC295380
Subset
IM
Grants
NIAID NIH HHS · R01 AI022564 · United States
NIAID NIH HHS · AI-09169 · United States
NIAID NIH HHS · AI-14929 · United States
NIAID NIH HHS · AI-22564 · United States
External Links
PubMed source DOI: 10.1172/JCI115336 PMC Full Text View journal details
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