An in vitro system to screen for diarrheagenic chemicals

We examined an in vitro system to screen for diarrheagenic chemicals using an established intestinal cell line (T84 human colonic carcinoma). The cells were grown on Millicell-PCF (polycarbonate membrane) wells. The cells were seeded at approximately 5 x 10(6) cells/30mm well and incubated for 9-11 days in a 5% CO2 incubator saturated with water at 37 degrees C. The culture medium was a 1:1 mixture of Ham's F12 and Dulbecco's MEM with 5% fetal bovine serum and 25 micrograms/ml gentamicin sulfate. The well containing cells was removed from the incubator and mounted in a modified Ussing chamber for measurement of short-circuit current (ISC). Chemical-induced increases in ISC are usually indicative of electrogenic epithelial Cl- secretion, which is associated with diarrheagenic effects in animals and humans. T84 cells grown on Millicell-PCF membrane responded with an increase in ISC after basolateral addition of the cholinergic (muscarinic) agonist carbachol, prostaglandin E2, 16,16-dimethylprostaglandin E2, and forskolin, while non-diarrheagenic prostaglandin D2 did not affect ISC. Based on our results, this in vitro system has the potential to be adapted as a rapid screen for detecting diarrheagenic chemicals.