Release of ATP during host cell killing by enteropathogenic E. coli and its role as a secretory mediator.
Enteropathogenic Escherichia coli (EPEC) causes severe, watery diarrhea in children. We investigated ATP release during EPEC-mediated killing of human cell lines and whether released adenine nucleotides function as secretory mediators. EPEC triggered a release of ATP from all human cell lines tested: HeLa, COS-7, and T84 (colon cells) as measured using a luciferase kit. Accumulation of ATP in the supernatant medium was enhanced if an inhibitor of 5'-ectonucleotidase was included and was further enhanced if an ATP-regenerating system was added. In the presence of the inhibitor/regenerator, ATP concentrations in the supernatant medium reached 1.5-2 microM 4 h after infection with wild-type EPEC strains. In the absence of the inhibitor/regenerator system, extracellular ATP was rapidly broken down to ADP, AMP, and adenosine. Conditioned medium from EPEC-infected cells triggered a brisk chloride secretory response in intestinal tissues studied in the Ussing chamber (rabbit distal colon and T84 cell monolayers), whereas conditioned medium from uninfected cells and sterile filtrates of EPEC bacteria did not. The short-circuit current response to EPEC-conditioned medium was completely reversed by adenosine receptor blockers, such as 8-(p-sulfophenyl)-theophylline and MRS1754. EPEC killing of host cells releases ATP, which is broken down to adenosine, which in turn stimulates secretion via apical adenosine A2b receptors. These findings provide new insight into how EPEC causes watery diarrhea.
Crane
John K
JK
Department of Medicine, University at Buffalo, State University of New York, Buffalo, New York 14214, USA. jcrane@acsu.buffalo.edu
Olson
Ruth A
RA
Jones
Heather M
HM
Duffey
Michael E
ME
D015720:5'-Nucleotidase / Q000502:physiology; D000241:Adenosine / Q000096:biosynthesis; D000244:Adenosine Diphosphate / Q000096:biosynthesis; D000249:Adenosine Monophosphate / Q000096:biosynthesis; D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D016923:Cell Death; D002478:Cells, Cultured; D002712:Chlorides / Q000378:metabolism; D003106:Colon / Q000382:microbiology* / Q000473:pathology / Q000503:physiopathology*; D004926:Escherichia coli / Q000502:physiology*; D004927:Escherichia coli Infections / Q000473:pathology / Q000503:physiopathology*; D006801:Humans; D008297:Male; D011817:Rabbits; D018047:Receptors, Purinergic P1 / Q000502:physiology
D002712:Chlorides; D018047:Receptors, Purinergic P1; D000249:Adenosine Monophosphate; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D015720:5'-Nucleotidase; D000241:Adenosine