Interleukin-1 beta induces cytosolic phospholipase A2 and prostaglandin H synthase in rheumatoid synovial fibroblasts. Evidence for their roles in the production of prostaglandin E2.
OBJECTIVE
In order to investigate potential regulatory mechanisms for the increased production of prostaglandin E2 (PGE2) in interleukin-1 beta (IL-1 beta)-stimulated rheumatoid synovial fibroblasts (RSF), this study examined the induction of phospholipase A2 (PLA2) and prostaglandin H synthase (PGHS) enzymes and the correlation of these events with PGE2 production in IL-1 beta-stimulated RSF.


METHODS
Protein and messenger RNA (mRNA) levels of cytosolic PLA2 (cPLA2) and PGHS-2 enzymes in IL-1 beta-stimulated RSF were measured by Western and Northern blotting, respectively, using specific antisera and complementary DNA probes. Enzymatic activity of cPLA2 was determined in cell-free reaction mixtures utilizing mixed micelles of 14C-phosphatidylcholine and Triton X-100 as the substrate. PGE2 levels were quantitated using a commercial enzyme immunoassay kit.


RESULTS
Incubation of RSF with IL-1 beta increased the mRNA and protein levels for the high molecular weight cPLA2 as well as for the mitogen/growth factor-responsive PGHS (PGHS-2). The IL-1 receptor antagonist completely abolished the induction of these two enzymes and the stimulation of PGE2 production by IL-1 beta in RSF. In contrast, levels of the other known forms of these enzymes, i.e., the 14-kd secretory group II PLA2 (sPLA2) and the constitutive form of PGHS (PGHS-1), were unaffected by IL-1 beta treatment.


CONCLUSIONS
These are the first data to demonstrate the coordinate induction by IL-1 of cPLA2 and PGHS-2 in RSF. The time-course for the induction of these enzymes suggests that their increase contributes to the increased production of PGE2 in IL-1-treated RSF, and may help explain the capacity of RSF to produce large amounts of PGE2.
Hulkower
K I
KI
Hoffmann-La Roche, Inc., Nutley, New Jersey.
Wertheimer
S J
SJ
Levin
W
W
Coffey
J W
JW
Anderson
C M
CM
Chen
T
T
DeWitt
D L
DL
Crowl
R M
RM
Hope
W C
WC
Morgan
D W
DW
D001172:Arthritis, Rheumatoid / Q000276:immunology / Q000378:metabolism*; D015152:Blotting, Northern; D015153:Blotting, Western; D002478:Cells, Cultured; D003600:Cytosol / Q000201:enzymology*; D015232:Dinoprostone / Q000096:biosynthesis*; D005347:Fibroblasts / Q000201:enzymology*; D006801:Humans; D007375:Interleukin-1 / Q000276:immunology*; D010741:Phospholipases A / Q000235:genetics / Q000378:metabolism*; D054467:Phospholipases A2; D011451:Prostaglandin-Endoperoxide Synthases / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis; D013583:Synovial Membrane / Q000276:immunology / Q000378:metabolism*; D015854:Up-Regulation / Q000502:physiology
D007375:Interleukin-1; D012333:RNA, Messenger; D011451:Prostaglandin-Endoperoxide Synthases; D010741:Phospholipases A; D054467:Phospholipases A2; D015232:Dinoprostone