Glutamine and KGF each regulate extracellular thiol/disulfide redox and enhance proliferation in Caco-2 cells

Glutamine (Gln) and keratinocyte growth factor (KGF) each stimulate intestinal epithelial cell growth, but regulatory mechanisms are not well understood. We determined whether Gln and KGF alter intra- and extracellular thiol/disulfide redox pools in Caco-2 cells cultured in oxidizing or reducing cell medium and whether such redox variations are a determinant of proliferative responses to these agents. Cells were cultured over a physiological range of oxidizing to reducing extracellular thiol/disulfide redox (E-h) conditions, obtained by varying cysteine (Cys) and cystine (CySS) concentrations in cell medium. Cell proliferation was determined by 5-bromo-2-deoxyuridine (BrdU) incorporation. Gln (10 mmol/l) or KGF (10 mug/l) did not alter BrdU incorporation at reducing E-h (-131 to -150 mV), but significantly increased incorporation at more oxidizing E-h (Gln at 0 to -109 mV; KGF at -46 to -80 mV). Cellular glutathione/glutathione disulfide (GSH/GSSG) E-h was unaffected by Gln, KGF, or variations in extracellular Cys/CySS E-h. Control cells largely maintained extracellular Eh at initial values after 24 h (-36 to -136 mV). However, extracellular E-h shifted toward a narrow physiological range with Gln and KGF treatment (Gln -56 to -88 mV and KGF -76 to -92 mV, respectively; P<0.05 vs. control). The results indicate that thiol/disulfide redox state in the extracellular milieu is an important determinant of Caco-2 cell proliferation induced by Gln and KGF, that this control is independent of intracellular GSH redox status, and that both Gln and KGF enhance the capability of Caco-2 cells to modulate extremes of extracellular redox.