Oxidative stress causes a protein kinase C-independent increase of paracellular permeability in an in vitro epithelial model.
To evaluate the response of an epithelial barrier to a moderate but sustained oxidative stress, we cultured monolayers of Madin Darby canine kidney cells on microporous filters and exposed them to the hypoxanthine-xanthine oxidase (HX-XO) reaction. The transepithelial permeability coefficient for mannitol (Pm) was assessed as a marker of paracellular permeability. When the oxidative stress was limited in intensity and duration (production of 10 nmol/ml/min O2- with generation of 467 +/- 30 nmol/ml H2O2 over 1 h), we observed an increase of Pm with a delay of several hours (324 +/- 65% of baseline by 6 h, P < 0.005). There was complete return to control values by 24 h. The increase of Pm did not appear to be related to a depletion of cellular ATP. Protein kinase C (PKC) activity did not increase, and the rise in Pm was not prevented by CGP 41,251, a specific inhibitor of PKC. By contrast, CGP 41,251 inhibited the Pm increase that was elicited by PDBU, a phorbol ester that activates PKC. In our model, we conclude that a reversible increase of paracellular permeability occurs after oxidative stress independently of ATP depletion or PKC activation. Other, as yet unknown mechanisms have to be involved in this process.
Rochat
T
T
Department of Medicine, University of Geneva, Switzerland.
Burkhard
C
C
Finci-Cerkez
V
V
Meda
P
P
D000255:Adenosine Triphosphate / Q000378:metabolism; D000470:Alkaloids / Q000494:pharmacology; D000818:Animals; D002460:Cell Line; D004285:Dogs; D004594:Electrophysiology; D004848:Epithelium / Q000502:physiology*; D006861:Hydrogen Peroxide / Q000633:toxicity*; D066298:In Vitro Techniques; D007668:Kidney; D008353:Mannitol / Q000378:metabolism; D010084:Oxidation-Reduction; D010539:Permeability; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000502:physiology*; D019311:Staurosporine / Q000031:analogs & derivatives*; D013481:Superoxides / Q000633:toxicity*; D013997:Time Factors
D000470:Alkaloids; D013481:Superoxides; D015240:Phorbol 12,13-Dibutyrate; D008353:Mannitol; D000255:Adenosine Triphosphate; D006861:Hydrogen Peroxide; D011493:Protein Kinase C; D019311:Staurosporine; C059539:midostaurin