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J. Biol. Chem., Vol. 275, Issue 30, 23281-23286, July 28, 2000
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From We have previously shown that hypertonicity
stimulates cyclooxygenase-2 (COX-2) expression in cultured medullary
epithelial cells. The aims of the present study were (i) to examine the
role of cytoplasmic signaling through MAPK pathways in tonicity
regulation of COX-2 expression in collecting duct cells and (ii) to
assess the possible contribution of COX-2 to the survival of inner
medullary collecting duct (IMCD) cells under hypertonic conditions. In
mIMCD-K2 cells, a cell line derived from mouse IMCDs, hypertonicity
induced a marked increase in COX-2 protein expression. The stimulation was reduced significantly by inhibition of MEK1 (PD-98059, 5-50 µM) and p38 (SB-203580, 5-100 µM)
and was almost abolished by the combination of the two compounds. To
study the role of JNK in tonicity-stimulated COX-2 expression, IMCD-3
cell lines stably transfected with dominant-negative mutants of three
JNKs (JNK-1, -2, and -3) were used. Hypertonicity-stimulated COX-2
protein expression was significantly reduced in dominant-negative
JNK-2-expressing cells and was unchanged in dominant-negative JNK-1-
and JNK-3-expressing cells compared with controls. The reduction of
COX-2 expression was associated with greatly reduced viability of
dominant-negative JNK-2-expressing cells during hypertonicity
treatment.
4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine (PP2) (2-8 µM), an inhibitor of Src kinases, reduced the
tonicity-stimulated COX-2 expression in a dose-dependent
manner, whereas PP3, an inactive analog of PP2, had no effect.
Inhibition of COX-2 activity by NS-398 (30-90 µM) and
SC-58236 (10-20 µM) significantly reduced viability of
mIMCD-K2 cells subjected to prolonged hypertonic treatment. We conclude
that 1) all three members of the MAPK family (ERK, JNK-2, and p38) as
well as Src kinases are required for tonicity-stimulated COX-2
expression in mouse collecting duct cells and that 2) COX-2 may play a
role in cell survival of medullary cells under hypertonic conditions.
MAPK Mediation of Hypertonicity-stimulated Cyclooxygenase-2
Expression in Renal Medullary Collecting Duct Cells*
,,, and¶
NIDDK, National Institutes of Health, Bethesda,
Maryland 20892 and the § Department of Internal Medicine,
University of Colorado, Denver, Colorado 80262
*
This work was supported by NIDDK Grants DK-37448, DK-39255,
and DK-40042 from the National Institutes of Health and in part by the
General Clinical Research Center at the University of Michigan, funded
by United States Public Health Service Grant M01RR00042 from the
National Center for Research Resources, National Institutes of Health.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.
Copyright © 2000 by The American Society for Biochemistry and Molecular Biology, Inc.
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