Peroxynitrite inhibits enterocyte proliferation and modulates Src kinase activity in vitro.
Overproduction of nitric oxide (NO) or its toxic metabolite, peroxynitrite (ONOO-), after endotoxemia promotes gut barrier failure, in part, by inducing enterocyte apoptosis. We hypothesized that ONOO- may also inhibit enterocyte proliferation by disrupting the Src tyrosine kinase signaling pathway, thereby blunting repair of the damaged mucosa. We examined the effect of ONOO- on enterocyte proliferation and Src kinase activity. Sprague-Dawley rats were challenged with LPS or saline, whereas intestinal epithelial cell line cells were treated with ONOO- or decomposed ONOO- in vitro. Enterocyte proliferation in vivo and in vitro was measured by 5-bromo-2'-deoxyuridine (BrdU) or [3H]thymidine incorporation. Src kinase activity in cell lysates was determined at various times. LPS challenge in vivo and ONOO- treatment in vitro inhibited enterocyte proliferation. ONOO- treatment blunted the activity of Src and its downstream target, focal adhesion kinase, in a time-dependent manner. ONOO- blocked mitogen (FBS, EGF)-induced enterocyte proliferation and Src phosphorylation while increasing Src nitration. Thus ONOO- may promote gut barrier failure not only by inducing enterocyte apoptosis but also by disrupting signaling pathways involved in enterocyte proliferation.
Potoka
Douglas A
DA
Children's Hospital of Pittsburgh, 3705 Fifth Ave. Pittsburgh, PA 15213, USA.
Upperman
Jeffrey S
JS
Zhang
Xiao-Ru
XR
Kaplan
Joshua R
JR
Corey
Seth J
SJ
Grishin
Anatoly
A
Zamora
Ruben
R
Ford
Henri R
HR
D000818:Animals; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D020895:Enterocytes / Q000166:cytology* / Q000201:enzymology*; D004815:Epidermal Growth Factor / Q000494:pharmacology; D016391:Genes, src; D008070:Lipopolysaccharides / Q000494:pharmacology; D008297:Male; D008934:Mitogens / Q000494:pharmacology; D030421:Peroxynitrous Acid / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects; D051381:Rats; D017207:Rats, Sprague-Dawley; D014162:Transfection; D019061:src-Family Kinases / Q000378:metabolism*
D008070:Lipopolysaccharides; D008934:Mitogens; D030421:Peroxynitrous Acid; D004815:Epidermal Growth Factor; D019061:src-Family Kinases