Skip to main content
PMC home page
British Journal of Pharmacology logoLink to British Journal of Pharmacology
. 1996 Aug;118(8):1945–1958. doi: 10.1111/j.1476-5381.1996.tb15629.x

Identification of cyclic AMP phosphodiesterases 3, 4 and 7 in human CD4+ and CD8+ T-lymphocytes: role in regulating proliferation and the biosynthesis of interleukin-2.

M A Giembycz 1, C J Corrigan 1, J Seybold 1, R Newton 1, P J Barnes 1
PMCID: PMC1909888  PMID: 8864528

Abstract

1. The cyclic AMP phosphodiesterases (PDE) expressed by CD4+ and CD8+ T-lymphocytes purified from the peripheral blood of normal adult subjects were identified and characterized, and their role in modulating proliferation and the biosynthesis of interleukin (IL)-2 and interferon (IFN)-gamma evaluated. 2. In lysates prepared from both subsets, SK&F 95654 (PDE3 inhibitor) and rolipram (PDE4 inhibitor) suppressed cyclic AMP hydrolysis indicating the presence of PDE3 and PDE4 isoenzymes in these cells. Differential centrifugation and subsequent inhibitor and kinetic studies revealed that the particulate fraction contained, predominantly, a PDE3 isoenzyme. In contrast, the soluble fraction contained a PDE4 (approximately 65% of total activity) and, in addition, a novel enzyme that had the kinetic characteristics of the recently identified PDE7. 3. Reverse transcription-polymerase chain reaction (RT-PCR) studies with primer pairs designed to recognise unique sequences in the human PDE4 and PDE7 genes amplified cDNA fragments that corresponded to the predicted sizes of HSPDE4A, HSPDE4B, HSPDE54D and HSPDE7. No message was detected for HSPDE4C after 35 cycles of amplification. 4. Functionally, rolipram inhibited phytohaemagglutinin- (PHA) and anti-CD3-induced proliferation of CD4+ and CD8+ T-lymphocytes, and the elaboration of IL-2, which was associated with a three to four fold increase in cyclic AMP mass. In all experiments, however, rolipram was approximately 60 fold more potent at suppressing IL-2 synthesis than at inhibiting mitogenesis. In contrast, SK&F 95654 failed to suppress proliferation and cytokine generation, and did not elevate the cyclic AMP content in T-cells. Although inactive alone, SK&F 95654 potentiated the ability of rolipram to suppress PHA- and anti-CD3-induced T-cell proliferation, and PHA-induced IL-2 release. 5. When a combination of phorbol myristate acetate (PMA) and ionomycin were used as a co-mitogen, rolipram did not affect proliferation but, paradoxically, suppressed IL-2 release indicating that cyclic AMP can inhibit mitogenesis by acting at, or proximal to, the level of inositol phospholipid hydrolysis. 6. Collectively, these data suggest that PDE3 and PDE4 isoenzymes regulate the cyclic AMP content, IL-2 biosynthesis and proliferation in human CD4+ and CD8+ T-lymphocytes. However, the ability of rolipram to suppress markedly mitogen-induced IL-2 generation without affecting T-cell proliferation suggests that growth and division of T-lymphocytes may be governed by mediators in addition to IL-2. Finally, T-cells have the potential to express PDE7, although elucidating the functional role of this enzyme must await the development of selective inhibitors.

Full text

PDF
1945

Images in this article

1951Figure 5
on p.1951

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Anastassiou E. D., Paliogianni F., Balow J. P., Yamada H., Boumpas D. T. Prostaglandin E2 and other cyclic AMP-elevating agents modulate IL-2 and IL-2R alpha gene expression at multiple levels. J Immunol. 1992 May 1;148(9):2845–2852. [PubMed] [Google Scholar]
  2. Averill L. E., Stein R. L., Kammer G. M. Control of human T-lymphocyte interleukin-2 production by a cAMP-dependent pathway. Cell Immunol. 1988 Aug;115(1):88–99. doi: 10.1016/0008-8749(88)90164-5. [DOI] [PubMed] [Google Scholar]
  3. Beavo J. A., Conti M., Heaslip R. J. Multiple cyclic nucleotide phosphodiesterases. Mol Pharmacol. 1994 Sep;46(3):399–405. [PubMed] [Google Scholar]
  4. Beavo J. A., Reifsnyder D. H. Primary sequence of cyclic nucleotide phosphodiesterase isozymes and the design of selective inhibitors. Trends Pharmacol Sci. 1990 Apr;11(4):150–155. doi: 10.1016/0165-6147(90)90066-H. [DOI] [PubMed] [Google Scholar]
  5. Bolger G., Michaeli T., Martins T., St John T., Steiner B., Rodgers L., Riggs M., Wigler M., Ferguson K. A family of human phosphodiesterases homologous to the dunce learning and memory gene product of Drosophila melanogaster are potential targets for antidepressant drugs. Mol Cell Biol. 1993 Oct;13(10):6558–6571. doi: 10.1128/mcb.13.10.6558. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Böyum A. Isolation of mononuclear cells and granulocytes from human blood. Isolation of monuclear cells by one centrifugation, and of granulocytes by combining centrifugation and sedimentation at 1 g. Scand J Clin Lab Invest Suppl. 1968;97:77–89. [PubMed] [Google Scholar]
  7. Chen D., Rothenberg E. V. Interleukin 2 transcription factors as molecular targets of cAMP inhibition: delayed inhibition kinetics and combinatorial transcription roles. J Exp Med. 1994 Mar 1;179(3):931–942. doi: 10.1084/jem.179.3.931. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Chomczynski P., Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987 Apr;162(1):156–159. doi: 10.1006/abio.1987.9999. [DOI] [PubMed] [Google Scholar]
  9. Conti M., Swinnen J. V., Tsikalas K. E., Jin S. L. Structure and regulation of the rat high-affinity cyclic AMP phosphodiesterases. A family of closely related enzymes. Adv Second Messenger Phosphoprotein Res. 1992;25:87–99. [PubMed] [Google Scholar]
  10. Corrigan C. J., Kay A. B. T cells and eosinophils in the pathogenesis of asthma. Immunol Today. 1992 Dec;13(12):501–507. doi: 10.1016/0167-5699(92)90026-4. [DOI] [PubMed] [Google Scholar]
  11. Davis R. L., Takayasu H., Eberwine M., Myres J. Cloning and characterization of mammalian homologs of the Drosophila dunce+ gene. Proc Natl Acad Sci U S A. 1989 May;86(10):3604–3608. doi: 10.1073/pnas.86.10.3604. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Dent G., Giembycz M. A., Evans P. M., Rabe K. F., Barnes P. J. Suppression of human eosinophil respiratory burst and cyclic AMP hydrolysis by inhibitors of type IV phosphodiesterase: interaction with the beta adrenoceptor agonist albuterol. J Pharmacol Exp Ther. 1994 Dec;271(3):1167–1174. [PubMed] [Google Scholar]
  13. Engels P., Fichtel K., Lübbert H. Expression and regulation of human and rat phosphodiesterase type IV isogenes. FEBS Lett. 1994 Aug 22;350(2-3):291–295. doi: 10.1016/0014-5793(94)00788-8. [DOI] [PubMed] [Google Scholar]
  14. Essayan D. M., Huang S. K., Undem B. J., Kagey-Sobotka A., Lichtenstein L. M. Modulation of antigen- and mitogen-induced proliferative responses of peripheral blood mononuclear cells by nonselective and isozyme selective cyclic nucleotide phosphodiesterase inhibitors. J Immunol. 1994 Oct 15;153(8):3408–3416. [PubMed] [Google Scholar]
  15. Giembycz M. A. Could isoenzyme-selective phosphodiesterase inhibitors render bronchodilator therapy redundant in the treatment of bronchial asthma? Biochem Pharmacol. 1992 May 28;43(10):2041–2051. doi: 10.1016/0006-2952(92)90160-k. [DOI] [PubMed] [Google Scholar]
  16. Giri J. G., Anderson D. M., Kumaki S., Park L. S., Grabstein K. H., Cosman D. IL-15, a novel T cell growth factor that shares activities and receptor components with IL-2. J Leukoc Biol. 1995 May;57(5):763–766. doi: 10.1002/jlb.57.5.763. [DOI] [PubMed] [Google Scholar]
  17. Goodwin J. S., Bankhurst A. D., Messner R. P. Suppression of human T-cell mitogenesis by prostaglandin. Existence of a prostaglandin-producing suppressor cell. J Exp Med. 1977 Dec 1;146(6):1719–1734. doi: 10.1084/jem.146.6.1719. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Goto Y., Takeshita T., Sugamura K. Adenosine 3',5'-cyclic monophosphate (cAMP) inhibits phorbol ester-induced growth of an IL-2-dependent T cell line. FEBS Lett. 1988 Nov 7;239(2):165–168. doi: 10.1016/0014-5793(88)80909-8. [DOI] [PubMed] [Google Scholar]
  19. Granja C., Lin L. L., Yunis E. J., Relias V., Dasgupta J. D. PLC gamma 1, a possible mediator of T cell receptor function. J Biol Chem. 1991 Sep 5;266(25):16277–16280. [PubMed] [Google Scholar]
  20. Ichimura M., Kase H. A new cyclic nucleotide phosphodiesterase isozyme expressed in the T-lymphocyte cell lines. Biochem Biophys Res Commun. 1993 Jun 30;193(3):985–990. doi: 10.1006/bbrc.1993.1722. [DOI] [PubMed] [Google Scholar]
  21. Johnson K. W., Davis B. H., Smith K. A. cAMP antagonizes interleukin 2-promoted T-cell cycle progression at a discrete point in early G1. Proc Natl Acad Sci U S A. 1988 Aug;85(16):6072–6076. doi: 10.1073/pnas.85.16.6072. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Klausner R. D., O'Shea J. J., Luong H., Ross P., Bluestone J. A., Samelson L. E. T cell receptor tyrosine phosphorylation. Variable coupling for different activating ligands. J Biol Chem. 1987 Sep 15;262(26):12654–12659. [PubMed] [Google Scholar]
  23. Krause D. S., Deutsch C. Cyclic AMP directly inhibits IL-2 receptor expression in human T cells: expression of both p55 and p75 subunits is affected. J Immunol. 1991 Apr 1;146(7):2285–2296. [PubMed] [Google Scholar]
  24. Lewko W. M., Smith T. L., Bowman D. J., Good R. W., Oldham R. K. Interleukin-15 and the growth of tumor derived activated T-cells. Cancer Biother. 1995 Spring;10(1):13–20. doi: 10.1089/cbr.1995.10.13. [DOI] [PubMed] [Google Scholar]
  25. Lingk D. S., Chan M. A., Gelfand E. W. Increased cyclic adenosine monophosphate levels block progression but not initiation of human T cell proliferation. J Immunol. 1990 Jul 15;145(2):449–455. [PubMed] [Google Scholar]
  26. Maier J. A., Hla T., Maciag T. Cyclooxygenase is an immediate-early gene induced by interleukin-1 in human endothelial cells. J Biol Chem. 1990 Jul 5;265(19):10805–10808. [PubMed] [Google Scholar]
  27. Marcoz P., Prigent A. F., Lagarde M., Nemoz G. Modulation of rat thymocyte proliferative response through the inhibition of different cyclic nucleotide phosphodiesterase isoforms by means of selective inhibitors and cGMP-elevating agents. Mol Pharmacol. 1993 Nov;44(5):1027–1035. [PubMed] [Google Scholar]
  28. Mary D., Aussel C., Ferrua B., Fehlmann M. Regulation of interleukin 2 synthesis by cAMP in human T cells. J Immunol. 1987 Aug 15;139(4):1179–1184. [PubMed] [Google Scholar]
  29. Maurice D. H., Haslam R. J. Molecular basis of the synergistic inhibition of platelet function by nitrovasodilators and activators of adenylate cyclase: inhibition of cyclic AMP breakdown by cyclic GMP. Mol Pharmacol. 1990 May;37(5):671–681. [PubMed] [Google Scholar]
  30. Michaeli T., Bloom T. J., Martins T., Loughney K., Ferguson K., Riggs M., Rodgers L., Beavo J. A., Wigler M. Isolation and characterization of a previously undetected human cAMP phosphodiesterase by complementation of cAMP phosphodiesterase-deficient Saccharomyces cerevisiae. J Biol Chem. 1993 Jun 15;268(17):12925–12932. [PubMed] [Google Scholar]
  31. Monaco L., Vicini E., Conti M. Structure of two rat genes coding for closely related rolipram-sensitive cAMP phosphodiesterases. Multiple mRNA variants originate from alternative splicing and multiple start sites. J Biol Chem. 1994 Jan 7;269(1):347–357. [PubMed] [Google Scholar]
  32. Murray K. J., Eden R. J., Dolan J. S., Grimsditch D. C., Stutchbury C. A., Patel B., Knowles A., Worby A., Lynham J. A., Coates W. J. The effect of SK&F 95654, a novel phosphodiesterase inhibitor, on cardiovascular, respiratory and platelet function. Br J Pharmacol. 1992 Oct;107(2):463–470. doi: 10.1111/j.1476-5381.1992.tb12768.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Novak T. J., Rothenberg E. V. cAMP inhibits induction of interleukin 2 but not of interleukin 4 in T cells. Proc Natl Acad Sci U S A. 1990 Dec;87(23):9353–9357. doi: 10.1073/pnas.87.23.9353. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Park D. J., Min H. K., Rhee S. G. Inhibition of CD3-linked phospholipase C by phorbol ester and by cAMP is associated with decreased phosphotyrosine and increased phosphoserine contents of PLC-gamma 1. J Biol Chem. 1992 Jan 25;267(3):1496–1501. [PubMed] [Google Scholar]
  35. Patel M. D., Samelson L. E., Klausner R. D. Multiple kinases and signal transduction. Phosphorylation of the T cell antigen receptor complex. J Biol Chem. 1987 Apr 25;262(12):5831–5838. [PubMed] [Google Scholar]
  36. Rincón M., Tugores A., López-Botet M. Cyclic AMP and calcium regulate at a transcriptional level the expression of the CD7 leukocyte differentiation antigen. J Biol Chem. 1992 Sep 5;267(25):18026–18031. [PubMed] [Google Scholar]
  37. Rincón M., Tugores A., López-Rivas A., Silva A., Alonso M., De Landázuri M. O., López-Botet M. Prostaglandin E2 and the increase of intracellular cAMP inhibit the expression of interleukin 2 receptors in human T cells. Eur J Immunol. 1988 Nov;18(11):1791–1796. doi: 10.1002/eji.1830181121. [DOI] [PubMed] [Google Scholar]
  38. Robicsek S. A., Blanchard D. K., Djeu J. Y., Krzanowski J. J., Szentivanyi A., Polson J. B. Multiple high-affinity cAMP-phosphodiesterases in human T-lymphocytes. Biochem Pharmacol. 1991 Jul 25;42(4):869–877. doi: 10.1016/0006-2952(91)90047-9. [DOI] [PubMed] [Google Scholar]
  39. Robicsek S. A., Krzanowski J. J., Szentivanyi A., Polson J. B. High pressure liquid chromatography of cyclic nucleotide phosphodiesterase from purified human T-lymphocytes. Biochem Biophys Res Commun. 1989 Aug 30;163(1):554–560. doi: 10.1016/0006-291x(89)92173-6. [DOI] [PubMed] [Google Scholar]
  40. Rott O., Cash E., Fleischer B. Phosphodiesterase inhibitor pentoxifylline, a selective suppressor of T helper type 1- but not type 2-associated lymphokine production, prevents induction of experimental autoimmune encephalomyelitis in Lewis rats. Eur J Immunol. 1993 Aug;23(8):1745–1751. doi: 10.1002/eji.1830230802. [DOI] [PubMed] [Google Scholar]
  41. Schwartz J. P., Passonneau J. V. Cyclic AMP-mediated induction of the cyclic AMP phosphodiesterase of C-6 glioma cells. Proc Natl Acad Sci U S A. 1974 Oct;71(10):3844–3848. doi: 10.1073/pnas.71.10.3844. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Seder R. A., Grabstein K. H., Berzofsky J. A., McDyer J. F. Cytokine interactions in human immunodeficiency virus-infected individuals: roles of interleukin (IL)-2, IL-12, and IL-15. J Exp Med. 1995 Oct 1;182(4):1067–1077. doi: 10.1084/jem.182.4.1067. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Seldon P. M., Barnes P. J., Meja K., Giembycz M. A. Suppression of lipopolysaccharide-induced tumor necrosis factor-alpha generation from human peripheral blood monocytes by inhibitors of phosphodiesterase 4: interaction with stimulants of adenylyl cyclase. Mol Pharmacol. 1995 Oct;48(4):747–757. [PubMed] [Google Scholar]
  44. Snijdewint F. G., Kaliński P., Wierenga E. A., Bos J. D., Kapsenberg M. L. Prostaglandin E2 differentially modulates cytokine secretion profiles of human T helper lymphocytes. J Immunol. 1993 Jun 15;150(12):5321–5329. [PubMed] [Google Scholar]
  45. Spears G., Sneyd J. G., Loten E. G. A method for deriving kinetic constants for two enzymes acting on the same substrate. Biochem J. 1971 Dec;125(4):1149–1151. doi: 10.1042/bj1251149. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Sun L., Ganea D. Vasoactive intestinal peptide inhibits interleukin (IL)-2 and IL-4 production through different molecular mechanisms in T cells activated via the T cell receptor/CD3 complex. J Neuroimmunol. 1993 Oct;48(1):59–69. doi: 10.1016/0165-5728(93)90059-8. [DOI] [PubMed] [Google Scholar]
  47. Takayama H., Trenn G., Sitkovsky M. V. Locus of inhibitory action of cAMP-dependent protein kinase in the antigen receptor-triggered cytotoxic T lymphocyte activation pathway. J Biol Chem. 1988 Feb 15;263(5):2330–2336. [PubMed] [Google Scholar]
  48. Takemoto D. J., Lee W. N., Kaplan S. A., Appleman M. M. Cyclic AMP phosphodiesterase in human lymphocytes and lymphoblasts. J Cyclic Nucleotide Res. 1978 Apr;4(2):123–132. [PubMed] [Google Scholar]
  49. Tamir A., Isakov N. Cyclic AMP inhibits phosphatidylinositol-coupled and -uncoupled mitogenic signals in T lymphocytes. Evidence that cAMP alters PKC-induced transcription regulation of members of the jun and fos family of genes. J Immunol. 1994 Apr 1;152(7):3391–3399. [PubMed] [Google Scholar]
  50. Tenor H., Staniciu L., Schudt C., Hatzelmann A., Wendel A., Djukanović R., Church M. K., Shute J. K. Cyclic nucleotide phosphodiesterases from purified human CD4+ and CD8+ T lymphocytes. Clin Exp Allergy. 1995 Jul;25(7):616–624. doi: 10.1111/j.1365-2222.1995.tb01109.x. [DOI] [PubMed] [Google Scholar]
  51. Thompson W. J., Appleman M. M. Multiple cyclic nucleotide phosphodiesterase activities from rat brain. Biochemistry. 1971 Jan 19;10(2):311–316. [PubMed] [Google Scholar]
  52. Thompson W. J., Ross C. P., Pledger W. J., Strada S. J., Banner R. L., Hersh E. M. Cyclic adenosine 3':5'-monophosphate phosphodiesterase. Distinct forms in human lymphocytes and monocytes. J Biol Chem. 1976 Aug 25;251(16):4922–4929. [PubMed] [Google Scholar]
  53. Torphy T. J., Stadel J. M., Burman M., Cieslinski L. B., McLaughlin M. M., White J. R., Livi G. P. Coexpression of human cAMP-specific phosphodiesterase activity and high affinity rolipram binding in yeast. J Biol Chem. 1992 Jan 25;267(3):1798–1804. [PubMed] [Google Scholar]
  54. Torphy T. J., Undem B. J. Phosphodiesterase inhibitors: new opportunities for the treatment of asthma. Thorax. 1991 Jul;46(7):512–523. doi: 10.1136/thx.46.7.512. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Tsuruta L., Lee H. J., Masuda E. S., Koyano-Nakagawa N., Arai N., Arai K., Yokota T. Cyclic AMP inhibits expression of the IL-2 gene through the nuclear factor of activated T cells (NF-AT) site, and transfection of NF-AT cDNAs abrogates the sensitivity of EL-4 cells to cyclic AMP. J Immunol. 1995 May 15;154(10):5255–5264. [PubMed] [Google Scholar]
  56. Wacholtz M. C., Minakuchi R., Lipsky P. E. Characterization of the 3',5'-cyclic adenosine monophosphate-mediated regulation of IL2 production by T cells and Jurkat cells. Cell Immunol. 1991 Jul;135(2):285–298. doi: 10.1016/0008-8749(91)90274-f. [DOI] [PubMed] [Google Scholar]
  57. Welch J. E., Swinnen J. V., O'Brien D. A., Eddy E. M., Conti M. Unique adenosine 3',5' cyclic monophosphate phosphodiesterase messenger ribonucleic acids in rat spermatogenic cells: evidence for differential gene expression during spermatogenesis. Biol Reprod. 1992 Jun;46(6):1027–1033. doi: 10.1095/biolreprod46.6.1027. [DOI] [PubMed] [Google Scholar]
  58. Wilkinson P. C., Liew F. Y. Chemoattraction of human blood T lymphocytes by interleukin-15. J Exp Med. 1995 Mar 1;181(3):1255–1259. doi: 10.1084/jem.181.3.1255. [DOI] [PMC free article] [PubMed] [Google Scholar]
  59. van Tits L. J., Michel M. C., Motulsky H. J., Maisel A. S., Brodde O. E. Cyclic AMP counteracts mitogen-induced inositol phosphate generation and increases in intracellular Ca2+ concentrations in human lymphocytes. Br J Pharmacol. 1991 Jun;103(2):1288–1294. doi: 10.1111/j.1476-5381.1991.tb09782.x. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from British Journal of Pharmacology are provided here courtesy of The British Pharmacological Society

RESOURCES