Signaling of short- and long-term regulation of intestinal epithelial type 1 Na+/H+ exchanger by interferon-gamma.
The present study evaluated the effect of interferon-gamma (IFN-gamma) on intestinal Na+/H+ exchange (NHE) activity and the intracellular signaling pathways set into motion after IFN-gamma receptor activation. Caco-2 cells express endogenous NHE1, NHE2 and NHE3 proteins, as detected by immunoblotting. Short- (0.5 h) and long- (24 h) term exposure of Caco-2 cells to IFN-gamma resulted in a concentration-dependent decrease in NHE activity. Inhibition of NHE activity by IFN-gamma was absent in cariporide-treated cells, but not in cells treated with S-3226. The long-term exposure to IFN-gamma was accompanied by a 20% increase in surface NHE1 abundance and no changes in total NHE1 abundance. Inhibition of Raf1, mitogen-activated protein kinase kinase (MAPKK/MEK) and p38 MAPK with, respectively, GW 5074, PD 98059 and SB 203580 and downregulation of protein kinase C (PKC) with phorbol-12,13-dibutyrate (100 nM for 24 h) prevented inhibition of NHE activity by IFN-gamma (0.5 and 24 h exposure). The signal transducer and activator transcription factor 1 (STAT1) inhibitor epigallocatechin-3-gallate (EGCG) prevented inhibition of NHE activity by long- but not the short-term treatment with IFN-gamma. Treatment with IFN-gamma activated phospho-p38 MAPK, this effect being detected as early as 1 h, persisting over 3 h and decreasing after 24 h. IFN-gamma produced a sustained action of phospho-STAT1 that was prevented by EGCG and partially attenuated by SB 203580 and insensitive to downregulation of PKC. In conclusion, short- and long-term inhibition of NHE1 activity by IFN-gamma involves a complex signaling pathway that includes PKC activation and STAT1 phosphorylation, respectively, but is not accompanied by downregulation of NHE1.
Magro
Fernando
F
Faculty of Medicine, Institute of Pharmacology and Therapeutics, 4200-319 Porto, Portugal.
Fraga
Sónia
S
Soares-da-Silva
Patrício
P
D018938:Caco-2 Cells; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D007371:Interferon-gamma / Q000494:pharmacology*; D007413:Intestinal Mucosa / Q000187:drug effects* / Q000378:metabolism*; D017471:Receptors, Interferon / Q000378:metabolism; D015398:Signal Transduction; D017923:Sodium-Hydrogen Exchangers / Q000037:antagonists & inhibitors* / Q000378:metabolism; D013997:Time Factors; D000096743:Interferon gamma Receptor
D007371:Interferon-gamma; D017471:Receptors, Interferon; D017923:Sodium-Hydrogen Exchangers; D000096743:Interferon gamma Receptor
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