Fig. 1.

H₂O₂ increases AMP-activated protein kinase (AMPK) phosphorylation in intestinal epithelial cells. T84 and HT-29.cl19a cells grown as monolayers on semipermeable supports were treated bilaterally with H₂O₂ (500 μM) for 2–30 min or the AMPK activator 5-aminoimidazole-4-carboxamide-1-β-d-ribofuranoside (AICAR, 2.5 mM) for 30 min. A: Western blot analysis of phosphorylation of the activating Thr¹⁷² residue on the catalytic α-subunit of AMPK. Molecular mass (kDa) of each protein is shown at right. B: densitometric quantitation of Western blot results in A and normalization to total AMPKα levels. C: CCh-induced change in ion transport reported as change in short-circuit current (ΔI_sc). T84 monolayers mounted in Ussing chambers were not treated (control) or preincubated with compound C (1–100 μM, bilaterally) for 30 min before administration of H₂O₂ (500 μM) for 30 min followed by stimulation of electrogenic Cl⁻ secretion with carbachol (CCh, 100 μM, basolaterally, n = 4). D: change in ion transport reported as change in short-circuit current (ΔI_sc). HT-29.cl19a monolayers mounted in Ussing chambers were untreated (control) or pretreated with compound C (50 μM, bilaterally) for 30 min before administration of H₂O₂ (500 μM, bilaterally) for 30 min and subsequent CCh stimulation of ion transport (n = 3). Values are means ± SE. *P < 0.05, **P < 0.01 vs. control (B) and vs. H₂O₂ + CCh (C and D); #P < 0.05 vs. compound C (1 μM).