, Volume 167, Issue 1, pp 277–284 | Cite as

Inhibition of ciliary activity by phorbol esters in rabbit tracheal epithelial cells

  • Kenji Kobayashi
  • Jun Tamaoki
  • Noritaka Sakai
  • Atsushi Chiyotani
  • Takao Takizawa


To study the effect of protein kinase C activation on respiratory ciliary activity, we measured ciliary beat frequency (CBF) by a photoelectric technique in response to phorbol esters and cell-permeable diglyceride in cultured tracheal epithelial cells from rabbits. Phorbol 12-myristate 13-acetate (PMA) resulted in a concentration- and time-dependent inhibition of CBF (half maximum inhibitory concentration (IC50)=3×10−10 M) with the maximal decrease being 21.0±1.4% (mean±SE,p<0.001) observed at 10−6 M. L-α-dioctanoylglycerol (DiC8), another known activator of protein kinase C, likewise reduced CBF in a dose-dependent fashion. In contrast, phorbol 12,13-didecanoate, a non-tumor-promoting phorbol ester that does not stimulate protein kinase C, produced no significant changes in CBF. The decrease in CBF induced by PMA was not affected by blockade of arachidonic acid metabolism with indomethacin and nordihydroguaiaretic acid, but was antagonized by pretreatment with H-7, a specific inhibitor of protein kinase C (p<0.01). Maximal ciliary inhibition with either PMA or DiC8 was not accompanied by a decrease in intracellular concentration of cyclic AMP. These results indicate that activation of protein kinase C has a significant depressive effect on ciliary activity, and hence the airway mucociliary transport function, presumably through a regulatory pathway that is not dependent on cyclic AMP or arachidonic acid metabolites.

Key words

Cilia Phorbol ester Protein kinase C Airway, mucociliary transport 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    Bell JD, Brunton LL (1986) Enhancement of adenylate cyclase activity in S49 lymphoma cells by phorbol esters: withdrawal of GTP-dependent inhibition. J Biol Chem 261:12036–12041PubMedGoogle Scholar
  2. 2.
    Castagna M, Takai Y, Kaibuchi K, Sano K, Kikkawa U, Nishizuka Y (1982) Direct activation of calcium-activated, phospholipid-dependent protein kinase by tumor-promoting phorbol esters. J Biol Chem 257:7847–7851PubMedGoogle Scholar
  3. 3.
    Chang EB, Wang NS, Rao MC (1985) Phorbol ester stimulation of active anion secretion in intestine. Am J Physiol 249:C356-C361PubMedGoogle Scholar
  4. 4.
    Dixon BS, Breckon R, Burke C, Anderson RJ (1988) Phorbol esters inhibit adenylate cyclase activity in cultured collecting tubular cells. Am J Physiol 254:C183-C191PubMedGoogle Scholar
  5. 5.
    Ebeling JG, Vandenbark GR, Kuhn LJ, Ganong BR, Bell RM, Niedel JE (1985) Diacylglycerols mimic phorbol diester induction of leukemic cell differentiation. Proc Natl Acad Sci USA 82:815–819PubMedCrossRefGoogle Scholar
  6. 6.
    George BZ, Stephen PH, Ramadan IS, Maurice BF (1985) Phorbol myristate acetate inhibits thrombin-stimulated Ca2+ mobilization and phosphatidylinositol 4,5-bisphosphate hydrolysis in human platelets. Proc Natl Acad Sci USA 82:3859–3862CrossRefGoogle Scholar
  7. 7.
    Hidaka J, Inagaki M, Kawamoto S, Sasaki Y (1984) Isoquinoline sulfonamides, novel and potent inhibitors of cyclic nucleotide dependent protein kinase and protein kinase C. Biochemistry 23:5036–5041PubMedCrossRefGoogle Scholar
  8. 8.
    John RH, Gregory DB, Fred LR Jr, Beulah H, James GW, John ER (1978) Comparison of the metabolism of alveolar macrophages from humans, rats, and rabbits: response to heat-killed bacteria or phorbol myristate acetate. J Lab Clin Med 92:787–794Google Scholar
  9. 9.
    Kotlikoff MI, Murray RK, Reynolds EE (1987) Histamine-induced calcium release and phorbol antagonism in cultured airway smooth muscle cells. Am J Physiol 253:C561-C566PubMedGoogle Scholar
  10. 10.
    Lawrence RD, Pamela SS, Richard BJ Jr (1976) Effect of phorbol myristate acetate on the oxidative metabolism of human polymorphonuclear leukocytes. Blood 47:545–554Google Scholar
  11. 11.
    Leatherman GF, Kim D, Smith TW (1987) Effect of phorbol esters on contractile state and calcium flux in cultured chick heart cells. Am J Physiol 253:H205-H209PubMedGoogle Scholar
  12. 12.
    Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with the Folin phenol reagent. J Biol Chem 193:265–275PubMedGoogle Scholar
  13. 13.
    Nishizuka Y (1988) The molecular heterogeneity of protein kinase C and its implications for cellular regulation. Nature 334:661–665PubMedCrossRefGoogle Scholar
  14. 14.
    Rink TJ, Sanchez A, Hallum J (1983) Diacylglycerol and phorbol ester stimulate secretion without raising cytoplasmic free calcium in human platelets. Nature 305:317–319PubMedCrossRefGoogle Scholar
  15. 15.
    Robert IL, Lenore C (1981) Receptor-mediated regulation of superoxide production in human neutrophils stimulated by phorbol myristate. J Clin Invest 68:1314–1320CrossRefGoogle Scholar
  16. 16.
    Ronit SE, -Harry L, Israel P (1985) Protein kinase C regulation of the receptor-coupled calcium signal in histamine-secreting rat basophilic leukaemia cells. Nature 313:59–60CrossRefGoogle Scholar
  17. 17.
    Ross SM, Corrsin S (1974) Results of an analytical model of mucociliary pumping. J Appl Physiol 37:333–340PubMedGoogle Scholar
  18. 18.
    Seymour H (1984) 12-O-tetradecanoyl-phorbol-13-acetate-induced ACTH secretion in pituitary tumor cells. Eur J Pharmacol 98:177–183CrossRefGoogle Scholar
  19. 19.
    Stephen PW, Eduardo GL (1985) 1,2-Diacylglycerol and phorbol ester inhibit agonist-induced formation of inositol phosphates in human platelets: possible implications for negative feedback regulation of inositol phospholipid hydrolysis. Proc Natl Acad Sci USA 82:2623–2626CrossRefGoogle Scholar
  20. 20.
    Tamaoki J, Kondo M, Takizawa T (1988) Stimulation of ciliary activity by indomethacin in rabbit tracheal epithelial organ culture. Respiration 54:127–131PubMedCrossRefGoogle Scholar
  21. 21.
    Tamaoki J, Kondo M, Takizawa T. Effect of cyclic AMP on ciliary function in rabbit tracheal epithelial cells. J Appl Physiol, in pressGoogle Scholar
  22. 22.
    Usuki I (1959) Effect of adenosine triphosphate on the ciliary activity and its histochemical demonstration in the oyster gill. Sci Rep Res Inst Tohoku Univ 25:65–72Google Scholar
  23. 23.
    Wanner A (1977) State of the art: clinical aspects of mucociliary transport. Am Rev Respir Dis 116:73–125PubMedGoogle Scholar
  24. 24.
    Wanner A, Maurer D, Abraham WM, Szepfalusi Z, Sielczak M (1983) Effects of chemical mediators of anaphylaxis on ciliary function. J Allergy Clin Immunol 72:663–667PubMedCrossRefGoogle Scholar
  25. 25.
    Wanner A (1985) Effects of methylxanthines on airway mucociliary function. Am J Med 79:16–21PubMedCrossRefGoogle Scholar
  26. 26.
    Welsh MJ (1987) Effects of phorbol ester and calcium ionophore on chloride secretion in canine tracheal epithelium. Am J Physiol 253:C828-C834PubMedGoogle Scholar
  27. 27.
    Yager J, Chen T-M, Dulfano MJ (1978) Measurement of frequency of ciliary beats of human respiratory epithelium. Chest 73:627–633PubMedGoogle Scholar

Copyright information

© Springer-Verlag New York, Inc 1989

Authors and Affiliations

  • Kenji Kobayashi
    • 1
  • Jun Tamaoki
    • 1
  • Noritaka Sakai
    • 1
  • Atsushi Chiyotani
    • 1
  • Takao Takizawa
    • 1
  1. 1.The First Department of MedicineTokyo Women’s Medical CollegeTokyoJapan

Personalised recommendations