Genistein and tyrphostin 47 stimulate CFTR-mediated Cl- secretion in T84 cell monolayers

C. L. Sears, F. Firoozmand, A. Mellander, F. G. Chambers, I. G. Eromar, A. G.M. Bot, B. Scholte, H. R. De Jonge, M. Donowitz

Research output: Contribution to journalArticle

Abstract

The involvement of tyrosine phosphorylation in the regulation of epithelial cell Cl- secretion is unknown. Therefore, the purpose of these studies was to determine if tyrosine kinase activation was involved in the regulation of Cl- secretion, using the tyrosine kinase inhibitors, genistein and tyrphostin 47, and human intestinal epithelial cells (T84 cells) as an intestinal Cl- secretory model. Genistein rapidly but reversibly stimulated sustained apical Cl- secretion in monolayers of T84 cells without increasing intracellular cyclic nucleotides or Ca2+ levels. Tyrphostin 47 also stimulated Cl- secretion in T84 monolayers, although it was short-lived. Transfection experiments in 3T3 fibroblasts and IEC-6 intestinal cells utilizing wild-type cystic fibrosis transmembrane conductance regulator (CFTR) showed that genistein and tyrphostin 47 stimulated 125I efflux only in CFTR-transfected cells and not in CFTR-negative cells. Thus genistein- and tyrphostin 47-stimulated Cl- secretion involved CFTR. Genistein also acted synergistically with the Ca2+- and protein kinase C-dependent acetylcholine analogue, carbachol, to stimulate Cl- secretion in T84 monolayers. However, the Cl- secretory response to saturating concentrations of the adenosine 3',5'-cyclic monophosphate (cAMP) agonist, forskolin, or the guanosine 3',5'- cyclic monophosphate (cGMP) agonist, Escherichia coli heat-stable enterotoxin, was not further enhanced by genistein. Although the mechanism of activation of Cl- secretion is unclear, these data suggest that tyrosine kinase activity limits basal Cl- secretion in T84 cells and that inhibition of T84 cell tyrosine kinase(s) stimulates apical membrane Cl- secretion, most likely through activation of the CFTR-Cl- channel. Moreover, genistein does not itself act through cAMP or cGMP elevation but appears to share a common Cl- secretory pathway with cyclic nucleotide-dependent agonists, whereas it augments the secretory responses to a Ca2+- and protein kinase C-dependent agonist.

Original languageEnglish (US)
Pages (from-to)G874-G882
JournalAmerican Journal of Physiology - Gastrointestinal and Liver Physiology
Volume269
Issue number6 32-6
DOIs
StatePublished - 1995

Keywords

  • Escherichia coli heat-stable enterotoxin
  • chloride
  • cystic fibrosis transmembrane conductance regulator
  • diarrhea
  • epithelium
  • guanylate cyclase
  • human intestinal epithelial cell
  • protein kinases
  • tyrosine kinases

ASJC Scopus subject areas

  • Physiology
  • Hepatology
  • Gastroenterology
  • Physiology (medical)

Fingerprint Dive into the research topics of 'Genistein and tyrphostin 47 stimulate CFTR-mediated Cl<sup>-</sup> secretion in T84 cell monolayers'. Together they form a unique fingerprint.

  • Cite this