1. Volume 137, Issue 3, Pages 976-985 (September 2009)
Activation of Intestinal Cl− Secretion by Lubiprostone Requires the Cystic Fibrosis Transmembrane Conductance Regulator 
Marcel J.C. Bijvelds, Alice G.M. Bot, Johanna C. Escher, Hugo R. de Jonge 
Gastroenterology 
Volume 137, Issue 3, Pages (September 2009)
DOI: /j.gastro
Copyright © 2009 AGA Institute Terms and Conditions

2. Figure 1
Lubiprostone evokes a CFTR-dependent Isc response in T84 monolayers. The lubiprostone-induced Isc response was inhibited by the CFTR blocker CFTRinh172 (C172) and by bumetanide (Bu), but not by CdCl2. The traces depict Isc recordings of a representative experiment. Aggregate data are summarized in the bar diagram. Lubiprostone and CFTRinh172 were added bilaterally. CdCl2 was added to the apical bath, whereas bumetanide was added to the subapical bath. ***P < .001.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2009 AGA Institute Terms and Conditions

3. Figure 2
Lubiprostone and forskolin activate the same pool of apical Cl− channels. Isc recordings were made on nystatin-treated T84 monolayers, in the presence of a transepithelial Cl− gradient. (A) Prior stimulation with forskolin greatly attenuated the response to lubiprostone. (B, C) Vice versa, lubiprostone treatment greatly attenuated the response to forskolin. Cftrinh172, but not CdCl2, inhibited the combined lubiprostone/forskolin response. The traces depict Isc recordings of a representative experiment. Aggregate data are summarized in the bar diagram. ***P < .001.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2009 AGA Institute Terms and Conditions

4. Figure 3
Inhibition of PKA greatly attenuates the lubiprostone-evoked Isc response in T84 monolayers. H89 was added bilaterally, 30 minutes before stimulation with lubiprostone. The traces depict Isc recordings of a representative experiment. Aggregate data are summarized in the bar diagram. ***P < .001.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2009 AGA Institute Terms and Conditions

5. Figure 4
Lubiprostone evokes a CFTR-dependent Isc response in mouse ileum. (A) Lubiprostone, added bilaterally, induced a dose-dependent elevation in Isc. This response attenuated a subsequent response to forskolin, but not to carbachol. Vice versa, forskolin pretreatment virtually abolished the response to lubiprostone. (B) CdCl2, added to the apical bath, either after or before lubiprostone, did not affect the lubiprostone-induced Isc response. (C) Tissue from Cftr-null mice did not respond to lubiprostone or forskolin, whereas carbachol elicited a small, transient Isc response. Addition of glucose to the apical bath triggered Na+-coupled glucose uptake. (D) Summary of data, showing the effect of forskolin, lubiprostone, and carbachol on the ileal Isc response of control mice, Cftr-null mice, and homozygous F508del animals. **P < .01; ***P < .001.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2009 AGA Institute Terms and Conditions

6. Figure 5
The lubiprostone response is mediated via EP4-type prostanoid receptor stimulation and cAMP signaling. (A) T84 monolayers, and mouse ileum and colon were stimulated with lubiprostone at a dose shown to evoke near maximal stimulation of the Isc. Subsequent addition of the EP4 receptor antagonist L-161,982 inhibited the lubiprostone response dose dependently, but did not block the forskolin response. (B) Both PGE2 and lubiprostone induced a rise in cellular cAMP levels that is sensitive to L-161,982. In combination, they were equally effective in increasing cAMP levels as was PGE2 on its own accord. IBMX further potentiated the lubiprostone-induced increase in cellular cAMP levels. L-161,982 did not affect a forskolin-induced rise in cAMP levels. n = 3. ***P < .001; #P < .05 vs control; $P < .001 vs control. ns, no statistically significant difference between means.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2009 AGA Institute Terms and Conditions

7. Figure 6
The lubiprostone response in human intestinal epithelium requires CFTR and is inhibited by EP4 receptor blockage. The traces depict Isc recordings of a representative experiment. In rectal biopsies, amiloride was used to inhibit Na+ uptake through apically located cation channels (ENaC), and helped to ascertain the proper orientation of the tissue. (A) Lubiprostone induced a secretory response across rectal and ileal tissue from healthy subjects and non-CF patients, respectively. (B) In tissue from homozygous F508del patients (CFTR Δ/Δ), neither lubiprostone nor forskolin induced a secretory response. Hyperpolarization of epithelial cells by carbachol-induced opening of Ca2+-dependent K+ channels further stimulated Cl− secretion in tissue from healthy controls (A, C), but, in the absence of an apical Cl− conductance, K+ efflux predominates, causing a “reversed” Isc response in CF patients. (C) Preincubation of rectal tissue with L-161,982 blocked responsiveness to lubiprostone, but not to forskolin. (D) Peak responses induced by 1 μmol/L lubiprostone (added bilaterally) in rectal epithelium of healthy controls and homozygous F508del patients (CFTR Δ/Δ). *P < .05; **P < .01.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2009 AGA Institute Terms and Conditions