Phosphodiesterase profile of human B lymphocytes from normal and atopic donors and the effects of PDE inhibition on B cell proliferation.
1. CD19+ B lymphocytes were purified from the peripheral blood of normal and atopic subjects to analyse and compare the phosphodiesterase (PDE) activity profile, PDE mRNA expression and the importance of PDE activity for the regulation of B cell function. 2. The majority of cyclic AMP hydrolyzing activity of human B cells was cytosolic PDE4, followed by cytosolic PDE7-like activity; marginal PDE3 activity was found only in the particulate B cell fraction. PDE1, PDE2 and PDE5 activities were not detected. 3. By cDNA-PCR analysis mRNA of the PDE4 subtypes A, B (splice variant PDE4B2) and D were detected. In addition, a weak signal for PDE3A was found. 4. No differences in PDE activities or mRNA expression of PDE subtypes were found in B cells from either normal or atopic subjects. 5. Stimulation of B lymphocytes with the polyclonal stimulus lipopolysaccharide (LPS) induced a proliferative response in a time- and concentration-dependent manner, which was increased in the presence of interleukin-4 (IL-4). PDE4 inhibitors (rolipram, piclamilast) led to an increase in the cellular cyclic AMP concentration and to an augmentation of proliferation, whereas a PDE3 inhibitor (motapizone) was ineffective, which is in accordance with the PDE profile found. The proliferation enhancing effect of the PDE4 inhibitors was partly mimicked by the cyclic AMP analogues dibutyryl (db) cyclic AMP and 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole-3',5'-cyclic monophosphorothioate, Sp-isomer (dcl-cBIMPS), respectively. However, at concentrations exceeding 100 microM db-cyclic AMP suppressed B lymphocyte proliferation, probably as a result of cytotoxicity. Prostaglandin E2 (PGE2, 1 microM) and forskolin (10 microM) did not affect B cell proliferation, even when given in combination with rolipram. 6. Inhibition of protein kinase A (PKA) by differentially acting selective inhibitors (KT 5720, Rp-8-Br-cyclic AMPS) decreased the proliferative response of control cells and reversed the proliferation enhancing effects of rolipram. 7. Importantly, PDE4 activity in LPS/IL-4-activated B lymphocytes decreased by about 50% compared to unstimulated control values. 8. We conclude that an increase in cyclic AMP, mediated by down-regulation of PDE4 activity, is involved in the stimulation of B cell proliferation in response to LPS/IL-4. B cell proliferation in response to a mitogenic stimulus can be further enhanced by pharmacological elevation of cyclic AMP.
Gantner
F
F
Department of Biochemistry, University of Konstanz, Germany.
Götz
C
C
Gekeler
V
V
Schudt
C
C
Wendel
A
A
Hatzelmann
A
A
D018941:Antigens, CD19 / Q000032:analysis; D001402:B-Lymphocytes / Q000166:cytology / Q000201:enzymology* / Q000276:immunology; D002455:Cell Division; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D004261:DNA Replication; D003876:Dermatitis, Atopic / Q000097:blood*; D006801:Humans; D015847:Interleukin-4 / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology; D008213:Lymphocyte Activation / Q000187:drug effects; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology*; D010727:Phosphoric Diester Hydrolases / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics
D018941:Antigens, CD19; D008070:Lipopolysaccharides; D010726:Phosphodiesterase Inhibitors; D012333:RNA, Messenger; D015847:Interleukin-4; D017868:Cyclic AMP-Dependent Protein Kinases; D010727:Phosphoric Diester Hydrolases