Effect of cyclosporin and tacrolimus (FK506) on the antigen-induced mediator release, membrane potential and 86Rb+/K+ and Ca2+ fluxes in the RBL-2H3 cell line.
The immunosuppressants cyclosporin A (CsA) and tacrolimus (FK506) inhibit the activation by antigen of T-lymphocytes as well as mast cells. The mechanism of their action on mast cells has yet to be elucidated. We, therefore, assessed their effect on antigen-induced histamine and beta-hexosaminidase release, membrane potential changes (bis-oxonol fluorescent probe), 86RB+ (marker for K+)-efflux, the intracellular free calcium concentration ([Ca2+]i in single cells) and 45Ca2+ uptake (CsA only) in RBL-2H3 cells, a mucosal-type mast cell line, passively sensitized with monoclonal mouse IgE antibody. Antigen addition induced depolarization within 1-2 min, followed by slower repolarization, reaching a steady state (approximately 90% repolarization) after 7-9 min. CsA and FK506 each dose-dependently inhibited antigen-induced histamine and beta-hexosaminidase secretion and the membrane repolarization phase, with similar IC50s for both actions, approximately 20 nM for CsA and approximately 2 nM for FK506. Antigen-induced 86Rb+-efflux was also significantly inhibited. Antigen-evoked increase in [Ca2+]i (area under the curve, AUC) was reduced by 35% and 52% in the presence of CsA or FK506 (1 microM each), respectively. However, 45Ca2+-uptake was not inhibited by CsA. These results suggest that both CsA and FK506 may inhibit mediator release from mast cells via blocking two interrelated processes, which are involved in the secretory process: 1. Membrane repolarization phase, which is essential for optimal mediator secretion and is mediated by a Ca2+-sensitive K+-efflux, yet to be further characterized, and (2) Increase in [Ca2+]i, probably via reduction of Ca(+2)-release from intracellular stores, [Ca2+]s.
Narenjkar
Jamshid
J
Department of Pharmacology, University College London, Gower Street, London WC1E 6BT, London, UK.
Assem
El-Sayed K
el-SK
Wan
Beatrice Y C
BY
Marsh
Stephen
S
Ezeamuzie
Charles I
CI
D000818:Animals; D000941:Antigens / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D045744:Cell Line, Tumor; D016572:Cyclosporine / Q000494:pharmacology*; D004140:Dinitrophenols / Q000494:pharmacology; D006241:Haptens / Q000494:pharmacology; D006636:Histamine Release / Q000187:drug effects*; D007166:Immunosuppressive Agents / Q000494:pharmacology*; D008564:Membrane Potentials / Q000187:drug effects; D051381:Rats; D012414:Rubidium Radioisotopes; D012709:Serum Albumin / Q000494:pharmacology; D016559:Tacrolimus / Q000494:pharmacology*; D001619:beta-N-Acetylhexosaminidases / Q000378:metabolism*
D000941:Antigens; D004140:Dinitrophenols; D006241:Haptens; D007166:Immunosuppressive Agents; D012414:Rubidium Radioisotopes; D012709:Serum Albumin; C058064:dinitrophenyl-human serum albumin conjugate; D016572:Cyclosporine; D001619:beta-N-Acetylhexosaminidases; D002118:Calcium; D016559:Tacrolimus