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J. Biol. Chem., Vol. 276, Issue 25, 22048-22055, June 22, 2001
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From the Oxidative stress compromises the tight
junction, but the mechanisms underlying its recovery remain unclear. We
developed a model in which oxidative stress reversibly disrupts
the tight junction. Exposure of Madin-Darby canine
kidney cells to hydrogen peroxide markedly reduced
transepithelial resistance and disrupted the staining patterns of the
tight junction proteins ZO-1 and occludin. These changes were reversed
by catalase. The short-term reassembly of tight junctions was not
dependent on new protein synthesis, suggesting that recovery occurs
through re-utilization of existing proteins. Although ATP levels were
reduced, the reduction was insufficient to explain the observed
changes, since a comparable reduction of ATP levels (with
2-deoxy-D-glucose) did not induce these changes. The
intracellular hydrogen peroxide scavenger pyruvate protected
Madin-Darby canine kidney cells from loss of transepithelial resistance
as did the heavy metal scavenger
N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine. Of a wide variety of agents examined, only tyrosine kinase inhibitors and protein kinase C inhibitors markedly inhibited tight junction reassembly. During reassembly, tyrosine phosphorylation in or near the
lateral membrane, was detected by immunofluorescence. The tyrosine
kinase inhibitors genistein and PP-2 inhibited the recovery of
transepithelial resistance and perturbed the relocalization of ZO-1 and
occludin to the tight junction, indicating that tyrosine kinases,
possibly members of the Src family, are critical for reassembly after
oxidative stress.
Reassembly of the Tight Junction after Oxidative Stress Depends
on Tyrosine Kinase Activity*
§¶,
,,**, and
Renal Division, Department of Medicine,
Brigham and Women's Hospital and Harvard Medical School, Boston,
Massachusetts 02115, the Department of Surgery, Children's
Hospital, Boston, Massachusetts 02115, and the § Department
of Pediatrics and Medicine, Division of Nephrology and Hypertension,
University of California in San Diego,
La Jolla, California 92093
*
The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
Supported by National Institutes of Health Grant DK53507. To
whom correspondence should be addressed: University of California in
San Diego, Depts. of Pediatrics and Medicine, Div. of Nephrology and
Hypertension, 9500 Gilman Dr., La Jolla, CA 92093-0693. Tel.: 858-822-3482; Fax: 858-822-3483; E-mail: snigam@ucsd.edu.
Copyright © 2001 by The American Society for Biochemistry and Molecular Biology, Inc.
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