Characterization of a direct effect of phorbol myristate acetate on human neutrophil cell membrane using 31D8 monoclonal antibody

• Published in: Journal of leukocyte biology 1992-03, Vol.51 (3), p.289-295
• Main Authors: Woronick, Charles L., Maderazo, Eufronio G., Anthony, Monique N., Krause, Peter J., Sha'afi, Ramadan I.
• Format: Article
• Language: English
• Subjects: Acetamides - pharmacology; Antibodies, Monoclonal; Antigens - drug effects; Antigens - physiology; betamethasone; Betamethasone - pharmacology; Biological and medical sciences; Cell Membrane - drug effects; Cell physiology; Enzyme Inhibitors - pharmacology; Fundamental and applied biological sciences. Psychology; Humans; Indoles - pharmacology; Indomethacin - pharmacology; Leukotriene Antagonists; Lipoxygenase Inhibitors - pharmacology; membrane stabilization; Molecular and cellular biology; neutrophil oxidants; Neutrophils - cytology; Oxidants - pharmacology; Phenyl Ethers; Phorbol Esters - pharmacology; Protein Kinase C - antagonists & inhibitors; protein kinase C inhibitors; Responses to growth factors, tumor promotors, other factors; Tetradecanoylphorbol Acetate - pharmacology; α‐tocopherol
• ISSN: 0741-5400; 1938-3673
• DOI: 10.1002/jlb.51.3.289

We found that 4‐β‐phorbol 12‐myristate 13‐acetate (PMA) caused decreased expression of the polymorphonuclear neutrophil (PMN) surface antigen 31D8. In contrast to the rapid initiation of the oxidative burst caused by PMA, the effect was slow to start but increased during incubation periods up to 50 min. To study this apparent protein kinase C‐independent late effect of PMA, we measured 31D8 expression in PMNs after incubation with various concentrations of PMA. The maximum PMA‐induced inhibition was 76 ± 2%, with an ID50 of 3.9 ± 0.4 ng/ml. Oxidants and prooxidants (hydrogen peroxide, hypochlorite, taurine‐chloramine, and ferrous iron, with or without H2O2) had no direct effect on 31D8 antigen expression. The following substances were not protective against the inhibitory affect of PMA: (1) antioxidants (superoxide dismutase, catalase, azide, dimethyl sulfoxide, Desferal, and ascorbate, with the exception of α‐tocopherol), (2) inhibitors of protein kinase C (H7 and W7), (3) inhibitors of 5‐lipoxygenase (A‐63162, MK886, and high‐dose indomethacin) and (4) inhibitors of cyclooxygenase (low‐dose indomethacin). Myeloperoxidase‐deficient PMNs had normal 31D8 antigen expression and a decrease of 31D8 antigen expression by PMA, as did normal PMNs. The inactive analog of PMA, 4‐α‐phorbol didecanoate, had no effect on 31D8 antigen expression. α‐Tocopherol (50 μg/ml) and betamethasone (150 μg/ml) protected against the PMA effect by 30.5 ± 7.3 (P < .0005) and 52 ± 15 (P < 0.004) channels, respectively. These results indicate that PMA has a protein kinase C‐independent late effect on human neutrophils, which can be prevented by pretreatment with α‐tocopherol or the steroid betamethasone. These compounds probably exert their protective effect by membrane stabilization.