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Ciliary beating amplitude controlled by intracellular Cl− and a high rate of CO2 production in ciliated human nasal epithelial cells

  • Signaling and cell physiology
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Abstract

The ciliary transport is controlled by two parameters of the ciliary beating, frequency (CBF) and amplitude. In this study, we developed a novel method to measure both CBF and ciliary bend distance (CBD, an index of ciliary beating amplitude) in ciliated human nasal epithelial cells (cHNECs) in primary culture, which are prepared from patients contracting allergic rhinitis and chronic sinusitis. An application of Cl−-free NO3− solution or bumetanide (an inhibitor of Na+/K+/2Cl− cotransport), which decreases intracellular Cl− concentration ([Cl−]i), increased CBD, not CBF, at 37 °C; however, it increased both CBD and CBF at 25 °C. Conversely, addition of Cl− channel blockers (5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB) and 4-[[4-Oxo-2-thioxo-3-[3-trifluoromethyl]phenyl]-5-thiazolidinylidene]methyl] benzoic acid (CFTR(inh)-172)), which increase [Cl−]i, decreased both CBD and CBF, suggesting that CFTR plays a crucial role for maintaining [Cl−]i in these cells. We speculate that Cl− modulates activities of the molecular motors regulating both CBD and CBF in cHNECs. Moreover, application of the CO2/HCO3−-free solution did not change intracellular pH (pHi), and addition of an inhibitor of carbonic anhydrase (acetazolamide) sustained pHi increase induced by the NH4+ pulse, which transiently increased pHi in the absence of acetazolamide. These results indicate that the cHNEC produces a large amount of CO2, which maintains a constant pHi even under the CO2/HCO3−-free condition.

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References

  1. Afzelius BA (2004) Cilia-related diseases. J Pathol 2004:470–477

    Article  CAS  Google Scholar 

  2. Angus SP, Edelmann RE, Pennock DG (2001) Targeted gene knockout of inner arm 1 in Tetrahymena thermophila. Eur J Cell Biol 80:486–497

    Article  CAS  PubMed  Google Scholar 

  3. Azbell C, Zhang S, Skinner D, Fortenberry J, Sorscher EJ, Woodwortg BA (2010) Hesperidin stimulates CFTR-mediated chloride secretion and ciliary beat frequency in sinonasal epithelium. Otolaryngol Head Neck Surg 143:397–404. https://doi.org/10.1016/j.otohns.2010.05.021

    Article  PubMed  PubMed Central  Google Scholar 

  4. Brokaw CJ (1994) Control of flagellar bending: a new agenda based on dynein diversity. Cell Motil Cytoskeleton 28:199–204

    Article  CAS  PubMed  Google Scholar 

  5. Brokaw CJ, Kamiya R (1987) Bending patterns of Chlamydomonas flagella: IV. Mutants with defects in inner and outer dynein arms indicate differences in dynein arm function. Cell Motil Cytoskeleton 8:68–75

    Article  CAS  PubMed  Google Scholar 

  6. Chen JC, Lo YF, Lin YW, Lin SH, Huang CL, Cheng CJ (2019) WNK4 kinase is a physiological intracellular chloride sensor. Proc Natl Acad Sci U S A 116:4502–4507

    Article  CAS  Google Scholar 

  7. Chilvers MA, Rutman A, O’Callaghan C (2003) Ciliary beat pattern is associated with specific ultrastructural defects in primary ciliary dyskinesia. J Allergy Clin Immunol 112:518–524

    Article  PubMed  Google Scholar 

  8. Delmotte P, Sanderson MJ (2006) Ciliary beat frequency is maintained at a maximal rate in the small airways of mouse lung slices. Am J Respir Cell Mol Biol 35:110–117

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Gibbons IR, Rowe AJ (1965) Dynein: a protein with adenosine triohosphatase activity from cilia. Science 149:424–426

    Article  CAS  PubMed  Google Scholar 

  10. Higashijima T, Ferguson KM, Sternweis PC (1987) Regulation of hormone-sensitive GTP-dependent regulatory proteins by chloride. J Biol Chem 262:3597–3602

    CAS  PubMed  Google Scholar 

  11. Hirosue S, Senn K, Clement N, Nonnenmacher M, Gigout L, Linden RM, Weber T (2007) Effect of inhibition of dynein function and microtubule-altering drugs on AAV2 transduction. Virology 367:10–18

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Hosogi S, Miyazaki H, Nakajima K, Ashihara E, Niisato N, Kusuzaki K, Marunaka Y (2012) An inhibitor of Na+/H+ exchanger (NHE), ethyl-isopropyl amiloride (EIPA), diminishes proliferation of MKN28 human gastric cancer cells by decreasing the cytosolic Cl− concentration via DIDS-sensitive pathways. Cell Physiol Biochem 30:1241–1253

    Article  CAS  PubMed  Google Scholar 

  13. Ikeuchi Y, Kogiso H, Tanaka S, Hosogi S, Nakahari T, Marunaka Y (2017) Carbocistein-activated bronchiolar ciliary beating via Cl− and pH-mediated pathways in mice. J Physiol Sci 67(Suppl):S137

    Google Scholar 

  14. Ikeuchi Y, Kogiso H, Hosogi S, Tanaka S, Shimamoto C, Inui T, Nakahari T, Marunaka Y (2018) Measurement of [Cl−]i unaffected by the cell volume change using MQAE-based two-photon microscopy in airway ciliary cells of mice. J Physiol Sci 68:191–199

    Article  CAS  PubMed  Google Scholar 

  15. Ikeuchi Y, Kogiso H, Hosogi S, Tanaka S, Shimamoto C, Matsumura H, Inui T, Nakahari T, Marunaka Y (2018) Carbocisteine stimulated an increase in ciliary bend angle via a decrease in [Cl−]i in mouse airway cilia. Pflugers Arch - Eur J Physiol 471:365–380. https://doi.org/10.1007/s00424-018-2212-2

    Article  CAS  Google Scholar 

  16. Inui T, Yasuda M, Hirano S, Ikeuchi Y, Kogiso H, Inui T, Marunaka Y, Nakahari T (2018) Daizein-stimulated increase in the ciliary beating amplitude via [Cl−]i decrease in ciliated human nasal epithelia; cells. Int J Mol Sci 19:3757. https://doi.org/10.3390/ijms19123754

    Article  CAS  Google Scholar 

  17. Kogiso H, Hosogi S, Ikeuchi Y, Tanaka S, Shimamoto C, Matsumura H, Nakano T, Sano K, Inui T, Marunaka Y, Nakahari T (2017) A low [Ca2+]i-induced enhancement of cAMP-activated ciliary beating by PDE1A inhibition in mouse airway cilia. Pflugers Arch - Eur J Physiol 469:1215–1227

    Article  CAS  Google Scholar 

  18. Kogiso H, Hosogi S, Ikeuchi Y, Tanaka S, Inui T, Marunaka Y, Nakahari T (2018a) [Ca2+]i modulation of cAMP-stimulated ciliary beat frequency via PDE1 in airway ciliary cells of mice. Exp Physiol 103:381–390

    Article  CAS  PubMed  Google Scholar 

  19. Kogiso H, Ikeuchi Y, Sumiya M, Hosogi S, Tanaka S, Shimamoto C, Inui T, Marunaka Y, Nakahari T (2018b) Seihai-to (TJ-90)-induced activation of airway ciliary beatings of mice: Ca2+ modulation of cAMP-stimulated ciliary beatings via PDE1. Int J Mol Sci 19(3):658. https://doi.org/10.3390/ijms19030658

    Article  CAS  PubMed Central  Google Scholar 

  20. Komatani-Tamiya, N, Daikoku E, Takemura Y, Shimamoto C, Nakano T, Iwasaki Y, Kohda Y, Matsumura H, Marunaka Y, Nakahari T (2012) Procaterol-stimulated increases in ciliary bend amplitude and ciliary beat frequency in mouse bronchioles. Cell Physiol Biochem 2012 29: 511–522

  21. Kuremoto T, Kogiso H, Yasuda M, Inui T, Murakami K, Hirano S, Ikeuchi Y, Hosogi S, Inui T, Marunaka Y, Nakahari T (2018) Spontaneous oscillation of the ciliary beat frequency regulated by release of Ca2+ from intracellular stores in mouse nasal epithelia. Biochem Biophys Res Commun 507:211–216

    Article  CAS  PubMed  Google Scholar 

  22. Lorenzo IM, Liedtke W, Sanderson MJ, Valverde MA (2008) TRPV4 channel participates in receptor-operated calcium entry and ciliary beat frequency regulation in mouse airway epithelial cells. Pro Natl Acad Sci USA 105:12611–12616

    Article  Google Scholar 

  23. Miertzschke M, Koerner C, Spoerner M, Wittinghofer A (2014) Structural insights into the small G-protein Arl13B and implications for Joubert syndrome. Biochem J 457:301–311

    Article  CAS  PubMed  Google Scholar 

  24. Müller L, Brighton LE, Carson JL, Fischer WA, Jaspers I (2013) Culturing of human nasal epithelial cells at the air liquid interface. J Vis Exp 80:e50646. https://doi.org/10.3791/50646

    Article  CAS  Google Scholar 

  25. Neesen J, Kirschner R, Ochs M, Schmiedl A, Habermann B, Muellwe C, Holstein AF, Nuesslein T, Adham I, Engel W (2001) Disruption on an inner arm dynein heavy chain gene results in asthenozoospermia and reduced ciliary beat frequency. Hum Mol Genet 10:1117–1128

    Article  CAS  PubMed  Google Scholar 

  26. Nakajima K, Niisato N, Marunaka Y (2012) Enhancement of tubulin polymerization by Cl−-induced blockade of intrinsic GTPase. Biochem Biophys Res Commun 425:225–229

    Article  CAS  PubMed  Google Scholar 

  27. Nakajima K, Marunaka Y (2016) Intracellular chloride ion concentration in differentiating neuronal cell and its role in growing neurite. Biochem Biophys Res Commun 479:338–342

    Article  CAS  PubMed  Google Scholar 

  28. Salathe M (2007) Regulation of mammalian ciliary beating. Annu Rev Physiol 69:401–422

    Article  CAS  PubMed  Google Scholar 

  29. Shpetner HS, Paschal BM, Vallee RB (1988) Characterization of the microtubule-activated ATPase of brain cytoplasmic dynein (MAP 1C). J Cell Biol 107:1001–1009

    Article  CAS  PubMed  Google Scholar 

  30. Shiima-Kinoshita C, Min K-Y, Hanafusa T, Mori H, Nakahari T (2004) β2-adrenergic regulation of ciliary beat frequency in rat bronchiolar epithelium: potentiation by isosmotic cell shrinkage. J Physiol 554:403–416

    Article  CAS  PubMed  Google Scholar 

  31. Shimamoto C, Umegaki E, Katsu K, Kato M, Fijiwara S, Kubota T, Nakahari T (2007) [Cl−]i modulation of Ca2+-regulated exocytosis in ACh-stimulated antral mucous cells of Guinea pig. Am J Physiol Gastrointest Liver Physiol 293:G824–G837

    Article  CAS  PubMed  Google Scholar 

  32. Sutto Z, Conner GE, Salathe M (2004) Regulation of human airway ciliary beat frequency by intracellular pH. J Physiol 560:519–532

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Terker AS, Zhang C, Erspamer KJ, Gamba G, Yang CL, Ellison DH (2016) Unique chloride-sensing properties of WNK4 permit the distal nephron to modulate potassium homeostasis. Kidney Int 89:127–134

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Tohda H, Foskett JK, O'Brodovich H, Marunaka Y (1994) Cl− regulation of a Ca2+-activated nonselective cation channel in beta-agonist-treated fetal distal lung epithelium. Am J Phys Cell Phys 266:C104–C109

    Article  CAS  Google Scholar 

  35. Tokuda S, Shimamoto C, Yoshida H, Murao H, Kishima G, Ito S, Kubota T, Hanafusa T, Sugimoto T, Niisato N, Marunaka Y, Nakahari T (2007) HCO3 −-dependent pHi recovery and overacidification induced by NH4 + pulse in rat lung alveolar type II cells: HCO3 −-dependent NH3 excretion from lungs? Pflugers Arch - Eur J Physiol 455:223–239

    Article  CAS  Google Scholar 

  36. Treharne KJ, Marshall LJ, Mehta A. A novel chloride-dependent GTP-utilizing protein kinase in plasma membranes from human respiratory epithelium. Am J Physiol 267 (Lung Cell Mol Physiol 11) 267: L592-L601, 1994

  37. Wanner A, Salathe M, O'Riordan TG (1996) Mucociliary clearance in the airways. Am J Respir Crit Care Med 154:1868–1902

    Article  CAS  PubMed  Google Scholar 

  38. Wood CR, Hard R, Hennessey TM (2007) Targeted gene disruption of dynein heavy chain 7 of Tetrahymena thermophila results in altred ciliary waveform and reduced swim speed. J Cell Sci 120:3075–3085

    Article  CAS  PubMed  Google Scholar 

  39. Yaghi A, Dolovich MB (2016) Airway epithelial cell cilia and obstructive lung disease. Cells 5(4):40. https://doi.org/10.3390/cells5040040

    Article  CAS  PubMed Central  Google Scholar 

  40. Yasuda M, Niisato N, Miyazaki H, Iwasaki Y, Hama T, Dejima K, Hisa Y, Marunaka Y (2007) Epithelial Na+ channel and ion transport in human nasal polyp and paranasal sinus mucosa. Biochem Biophys Res Commun 362:753–758

    Article  CAS  PubMed  Google Scholar 

  41. Yasuda M, Niisato N, Miyazaki H, Hama T, Dejima K, Hisa Y, Marunaka Y (2007) Epithelial ion transport of human nasal polyp and paranasal sinus mucosa. Am J Respir Cell Mol Biol 36:466–472

    Article  CAS  PubMed  Google Scholar 

  42. Zhang L, Sanderson MJ (2003) The role of cGMP in regulation of rabbit airway ciliary beat frequency. J Physiol 551:765–776

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

The authors thank Osaka medical College for renting out the video-microscope equipped with a high speed camera. Experiments were carried out in Kyoto Prefectural University of Medicine (2018–2019) and in Ritsumeikan University (2019).

Funding

This work was supported by JSPS KAKENHI to YM (No. JP18H03182), JSPS KAKENHI to MY (No. JP18K09325), and research funding from Saisei Mirai.

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Authors and Affiliations

  1. Department of Molecular Cell Physiology, Graduate School of Medical Sciences, Kyoto Prefectural University of Medine, Kyoto, 602-8566, Japan

    Taka-aki Inui, Kentaro Murakami, Yukiko Ikeuchi, Haruka Kogiso, Shigekuni Hosogi & Yoshinori Marunaka

  2. Department of Otolaryngology-Head and Neck Surgery, Graduate School of Medical Science, Kyoto Prefectural University of Medicine, Kyoto, 602-8566, Japan

    Taka-aki Inui, Kentaro Murakami, Makoto Yasuda & Shigeru Hirano

  3. Research Laboratory for Epithelial Physiology, Research Organization of Science and Technology BKC, Ritsumeikan University, Kusatsu, 525-8577, Japan

    Yukiko Ikeuchi, Haruka Kogiso, Toshio Inui, Yoshinori Marunaka & Takashi Nakahari

  4. Saisei Mirai Clinics, Moriguchi, 570-0012, Japan

    Toshio Inui

  5. Research Institute for Clinical Physiology, Kyoto Industrial Health Association, Kyoto, 604-8472, Japan

    Yoshinori Marunaka

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  1. Taka-aki Inui
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Correspondence to Makoto Yasuda or Takashi Nakahari.

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Inui, Ta., Murakami, K., Yasuda, M. et al. Ciliary beating amplitude controlled by intracellular Cl− and a high rate of CO2 production in ciliated human nasal epithelial cells. Pflugers Arch - Eur J Physiol 471, 1127–1142 (2019). https://doi.org/10.1007/s00424-019-02280-5

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