Cl- transport in an immortalized human epithelial cell line (NCM460) derived from the normal transverse colon

Cells of a newly described, immortalized, epithelial, human transverse colonic cell line, NCM460, reach similar to 90% confluence on plastic and develop transepithelial resistances of 120-250 Omega . cm(2) on porous substrates. Its utility as a model for the transverse human colon was validated by comparing second messenger-mediated Cl- transport, using the fluorescent probe 6-methoxy-quinolyl acetoethyl ester, in NCM460 cells and colonocytes isolated from human transverse crypts. Basal Cl- influx was increased (P < 0.01) by PGE(1) (1 mu M), forskolin (1 mu M), 8-bromoadenosine 3'5'-cyclic monophosphate (100 mu M), heat-stable Escherichia coli enterotoxin (STa; 1 mu M), 8-bromoguanosine 3'5'-cyclic monophosphate (100 mu M), histamine (1 mu M), and phorbol 12,13-dibutyrate (1 mu M) in both cell types. The Cl- channel blocker diphenylamine 2-carboxylic acid (50 mu M) and the Na+-K+-2Cl(-) cotransport inhibitor furosemide (1 mu M), but not the K+ channel blocker Ba2+ (3 mM), inhibited these Cl- permeabilities. These cells possess transcripts for cystic fibrosis transmembrane conductance regulator, Na+-K+-2Cl(-) cotransporter, STa receptor, and intestine-specific cGMP-dependent protein kinase II. Thus cAMP-, cGMP-, and Ca2+-dependent secretagogues act on NCM460 and primary colonocytes to stimulate Cl- transport. This validates the utility of NCM460 as a model for transverse colonic crypts and is the first demonstration of a colonic cell line whose origin is known.