Evidence for Bicarbonate Secretion by Ameloblasts in a Novel Cellular Model

E. Bori, J. Guo, R. Rácz, B. Burghardt, A. Földes, B. Kerémi, H. Harada, M. C. Steward, P. Den Besten, A. L J J Bronckers, G. Varga

Research output: Contribution to journalArticle

13 Citations (Scopus)

Abstract

Formation and growth of hydroxyapatite crystals during amelogenesis generate a large number of protons that must be neutralized, presumably by HCO3 - ions transported from ameloblasts into the developing enamel matrix. Ameloblasts express a number of transporters and channels known to be involved in HCO3 - transport in other epithelia. However, to date, there is no functional evidence for HCO3 - transport in these cells. To address questions related to HCO3 - export from ameloblasts, we have developed a polarized 2-dimensional culture system for HAT-7 cells, a rat cell line of ameloblast origin. HAT-7 cells were seeded onto Transwell permeable filters. Transepithelial resistance was measured as a function of time, and the expression of transporters and tight junction proteins was investigated by conventional and quantitative reverse transcription polymerase chain reaction. Intracellular pH regulation and HCO3 - transport were assessed by microfluorometry. HAT-7 cells formed epithelial layers with measureable transepithelial resistance on Transwell permeable supports and expressed claudin-1, claudin-4, and claudin-8 - key proteins for tight junction formation. Transport proteins previously described in maturation ameloblasts were also present in HAT-7 cells. Microfluorometry showed that the HAT-7 cells were polarized with a high apical membrane CO2 permeability and vigorous basolateral HCO3 - uptake, which was sensitive to Na+ withdrawal, to the carbonic anhydrase inhibitor acetazolamide and to H2DIDS inhibition. Measurements of transepithelial HCO3 - transport showed a marked increase in response to Ca2+- and cAMP-mobilizing stimuli. Collectively, 2-dimensional HAT-7 cell cultures on permeable supports 1) form tight junctions, 2) express typical tight junction proteins and electrolyte transporters, 3) are functionally polarized, and 4) can accumulate HCO3 - ions from the basolateral side and secrete them at the apical membrane. These studies provide evidence for a regulated, vectorial, basolateral-to-apical bicarbonate transport in polarized HAT-7 cells. We therefore propose that the HAT-7 cell line is a useful functional model for studying electrolyte transport by ameloblasts.

Original languageEnglish
Pages (from-to)588-596
Number of pages9
JournalJournal of Dental Research
Volume95
Issue number5
DOIs
Publication statusPublished - 2016

Keywords

  • cytophotometry
  • dental enamel
  • fluorescent dyes
  • in vitro techniques
  • ion transport
  • tissue engineering

ASJC Scopus subject areas

  • Dentistry(all)

Fingerprint Dive into the research topics of 'Evidence for Bicarbonate Secretion by Ameloblasts in a Novel Cellular Model'. Together they form a unique fingerprint.

  • Cite this

    Bori, E., Guo, J., Rácz, R., Burghardt, B., Földes, A., Kerémi, B., Harada, H., Steward, M. C., Den Besten, P., Bronckers, A. L. J. J., & Varga, G. (2016). Evidence for Bicarbonate Secretion by Ameloblasts in a Novel Cellular Model. Journal of Dental Research, 95(5), 588-596. https://doi.org/10.1177/0022034515625939