Intracellular regulation of intestinal folate uptake

Studies with cultured IEC-6 epithelial cells

Hamid M. Said, Thomas Ma, Alvaro Ortiz, Alicia Tapia, Claudia K. Valerio

Research output: Contribution to journalArticle

32 Citations (Scopus)

Abstract

Although the mechanism of folate uptake in the small intestine has been well characterized, very little is known about the intracellular regulation of the uptake process. Using mature confluent monolayers of the intestinal epithelial cell line IEC-6 as an in vitro intestinal epithelial cell model, we have found the uptake of folic acid to be similar to that of the native small intestine in that it is 1) temperature, energy, and pH dependent, 2) Na+ independent, 3) inhibited by structural analogs and anion transport inhibitors, and 4) saturable as a function of substrate concentration [apparent Michaelis constant (K(m)) = 0.45 ± 0.06 μM; maximal velocity (V(max)) = 3.08 ± 0.14 pmol-mg protein-1·5 min-1]. Furthermore, IEC-6 cells were found by Northern blot analysis to lack the expression of the membrane folate-binding protein. Pretreatment of IEC-6 monolayers with specific protein tyrosine kinase (PTK) inhibitors genistein and tyrphostin A25 caused a significant inhibition in folic acid uptake. On the other hand, their negative controls, genistin and tyrphostin A1, respectively, had no effect. The inhibitory effect of genistein was mediated through inhibition in the V(max) of the folate uptake process with no change in the apparent K(m). Pretreatment of IEC-6 monolayers with compounds that increase intracellular adenosine 3',5'-cyclic monophosphate (cAMP) level (e.g., dibutyryl cAMP) also resulted in a significant (though modest) inhibition in folic acid uptake; however, specific inhibitors of protein kinase A did not affect the uptake process. Specific modulators of protein kinase C and Ca2+/calmodulin- mediated pathways did not significantly affect folic acid uptake. These results demonstrate the suitability of IEC-6 monolayers as an intestinal epithelial model to study folate transport and demonstrate for the first time that uptake of folic acid is regulated by a PTK- and a cAMP-mediated pathway.

Original languageEnglish (US)
JournalAmerican Journal of Physiology - Cell Physiology
Volume272
Issue number2 41-2
StatePublished - Feb 1 1997

Fingerprint

Folic Acid
Epithelial Cells
Genistein
Small Intestine
Calmodulin
Protein Kinase Inhibitors
Cyclic AMP-Dependent Protein Kinases
Northern Blotting
Cyclic AMP
Protein-Tyrosine Kinases
Protein Kinase C
Anions
Tyrosine
Carrier Proteins
Cell Line
Temperature
Membranes

All Science Journal Classification (ASJC) codes

  • Physiology
  • Cell Biology

Cite this

Said, Hamid M. ; Ma, Thomas ; Ortiz, Alvaro ; Tapia, Alicia ; Valerio, Claudia K. / Intracellular regulation of intestinal folate uptake : Studies with cultured IEC-6 epithelial cells. In: American Journal of Physiology - Cell Physiology. 1997 ; Vol. 272, No. 2 41-2.
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Intracellular regulation of intestinal folate uptake : Studies with cultured IEC-6 epithelial cells. / Said, Hamid M.; Ma, Thomas; Ortiz, Alvaro; Tapia, Alicia; Valerio, Claudia K.

In: American Journal of Physiology - Cell Physiology, Vol. 272, No. 2 41-2, 01.02.1997.

Research output: Contribution to journalArticle

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AU - Said, Hamid M.

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