Calcitonin receptor-mediated CFTR activation in human intestinal epithelial cells.
High levels of calcitonin (CT) observed in medullary thyroid carcinoma and other CT-secreting tumours cause severe diarrhoea. Previous studies have suggested that CT induces active chloride secretion. However, the involvement of CT receptor (CTR) and the molecular mechanisms underlying the modulation of intestinal electrolyte secreting intestinal epithelial cells have not been investigated. Therefore, current studies were undertaken to investigate the direct effects of CT on ion transport in intestinal epithelial cells. Real time quantitative RT-PCR and Western blot analysis demonstrated the expression of CTR in intestinal epithelial T84 cells. Exposure of T84 cells to CT from the basolateral but not from apical side significantly increased short circuit current (I(SC) ) in a dose-dependent manner that was blocked by 1 μM of CTR antagonist, CT8-32. CT-induced I(SC) was blocked by replacing chloride in the bath solutions with equimolar gluconate and was significantly inhibited by the specific cystic fibrosis transmembrane conductance regulator (CFTR) inhibitor, CFTR(127inh). Further, biotinylation studies showed that CT increased CFTR levels on the apical membrane. The presence of either the Ca(2+) chelator, bis(2-aminophenoxy)ethane tetraacetic acid-acetoxymethyl (BAPTA-AM) ester or the protein kinase A (PKA) inhibitor, H89, significantly inhibited I(SC) induced by CT (∼32-58% reduction). Response to CT was retained after permeabilization of the basolateral or the apical membranes of T84 cells with nystatin. In conclusion, the activation of CTR by CT induced chloride secretion across T84 monolayers via CFTR channel and the involvement of PKA- and Ca(2+) -dependent signalling pathways. These data elucidate the molecular mechanisms underlying CT-induced diarrhoea.
Section of Digestive Diseases and Nutrition, Department of Medicine, University of Illinois at Chicago, Chicago, IL, USA.
Hongguang
H
Liu
Amika
A
Singla
Mei
M
Ao
Ravinder K
RK
Gill
Jayashree
J
Venkatasubramanian
Mrinalini C
MC
Rao
Waddah A
WA
Alrefai
Pradeep K
PK
Dudeja
D019921:Biotinylation; D015153:Blotting, Western; D002116:Calcitonin / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D049109:Cell Proliferation; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D000242:Cyclic AMP / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics / Q000378:metabolism*; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism*; D007422:Intestines / Q000166:cytology / Q000187:drug effects; D017136:Ion Transport; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D018003:Receptors, Calcitonin / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction
C505032:CFTR protein, human; D002614:Chelating Agents; D012333:RNA, Messenger; D018003:Receptors, Calcitonin; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D004533:Egtazic Acid; D002116:Calcitonin; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases
Am J Physiol. 1984 Feb;246(2 Pt 1):G204-8
6141741
Am J Physiol Gastrointest Liver Physiol. 2007 Jan;292(1):G369-76
17008555
Am J Physiol Cell Physiol. 2007 Jan;292(1):C452-9
16899552
Crit Rev Neurobiol. 1997;11(2-3):167-239
9209829
FEBS Lett. 1999 Sep 24;458(3):409-14
10570950
Trends Pharmacol Sci. 2005 Apr;26(4):172-5
15808339
Metabolism. 1971 Dec;20(12):1129-37
5001933
J Biol Chem. 1995 Dec 8;270(49):29194-200
7493947
Dig Dis Sci. 2005 Dec;50(12):2203-6
16416161
Gut. 1988 Apr;29(4):537-43
3371722
Am J Physiol Gastrointest Liver Physiol. 2004 Aug;287(2):G370-8
15075254
Dig Surg. 2004;21(4):321-4
15375334
J Cell Biol. 1980 Jun;85(3):682-94
7391137
J Endocrinol. 2007 Dec;195(3):415-27
18000304
Peptides. 2004 Oct;25(10):1633-42
15476930
Am J Physiol Cell Physiol. 2003 Feb;284(2):C339-48
12388102
J Nutr. 2008 Jul;138(7):1355-9
18567760
Proc Soc Exp Biol Med. 1975 Oct;150(1):151-4
1187688
Am J Respir Crit Care Med. 1994 Jan;149(1):183-90
8111580
J Bone Miner Metab. 2005;23(5):359-65
16133685
Gastroenterology. 1977 Mar;72(3):462-8
13025
Gastroenterology. 2009 Sep;137(3):976-85
19454284
J Physiol. 1996 Jun 15;493 ( Pt 3):885-94
8799908
Anticancer Res. 1997 Nov-Dec;17(6D):4667-70
9494586
Am J Physiol Gastrointest Liver Physiol. 2009 Feb;296(2):G275-83
19033536
Cancer Lett. 1994 Jun 15;81(1):19-25
8019984
J Biol Chem. 2004 Feb 20;279(8):7264-74
14615490
Mol Cell Probes. 2005 Apr;19(2):119-26
15680213
Nat Rev Drug Discov. 2009 Feb;8(2):153-71
19153558
Gastroenterology. 1977 Jul;73(1):89-94
193761
Recept Channels. 2002;8(3-4):243-55
12529940
J Pathol. 1982 Jan;136(1):27-39
7057295
Recent Prog Horm Res. 1972;28:399-470
4347303
Gastroenterology. 1976 Sep;71(3):392-8
950088
J Biol Chem. 2004 Sep 17;279(38):39310-6
15234961