Cyclic Adenosine Monophosphate and Alteration of Chinese Hamster Ovary Cell Morphology: a Rapid, Sensitive In Vitro Assay for the Enterotoxins of Vibrio cholerae and Escherichia coli

Abstract

The major limitation to our understanding of the clinical importance of enterotoxigenic Escherichia coli in diarrheal illness has been the lack of a simple rapid assay for the enterotoxin produced by certain E. coli. On the basis of the activation of adenylate cyclase by heat-labile enterotoxin of E. coli (LT) and by cholera toxin (CT) in intestinal and other tissues, cultured Chinese hamster ovary (CHO) cells with known morphological responses to dibutyryl cyclic adenosine 5′-monophosphate (AMP) were exposed to these enterotoxins. Crude culture filtrates of LT-producing E. coli and CT stimulated cyclic AMP accumulation and cell elongation in CHO cells. The similarity of time course, concentration dependence, and potentiation by phosphodiesterase inhibitors suggested cyclic AMP mediation of the morphological change. Heat inactivated CT and LT in this system. Choleragenoid inhibited CT; antiserum against CT inhibited both enterotoxin effects. In contrast to culture filtrates of 16 strains of E. coli known to produce LT, culture filtrates from 13 E. coli that do not produce LT did not alter CHO cell morphology. The morphological change is a simple, specific assay for these enterotoxins and detect 3 × 10 −17 mol of CT or a 1:250 dilution of crude culture filtrate of LT-producing E. coli 334.