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Suppression of human synovial fibroblast 85 kDa phospholipase A2 by antisense reduces interleukin-1 beta induced
A Roshak1, E Mochan, L A Marshall
1Department of Inflammation Pharmacology, SmithKline Beecham Pharmaceuticals, King of Prussia, PA, USA.
Objective:
To evaluate the relative roles of rheumatoid synovial fibroblast phospholipases A2 (PLA2) in interleukin- 1beta (IL-1 beta) stimulated prostaglandin E2 (PGE2) production.
Methods:
The role of the cytosolic 85 kDa PLA2 in IL-1beta induced human rheumatoid synovial fibroblast PGE2 formation was directly evaluated using an antisense phosphorothioate oligonucleotide to the initiation site of the 85 kDa PLA2 mRNA. Contribution of the 14 kDa PLA2 was assessed using selective inhibitors or a neutralizing monoclonal antibody (Mab).
Results:
Antisense, but not sense, decreased IL-1beta upregulation of 85 kDa PLA2 activity and protein levels. The antisense effect was specific, since it did not affect 14 kDa PLA2 activity released into the media or induced cyclooxygenase II protein levels over 24 h. Antisense, but not sense, reduced PGE2 formation in a concentration dependent manner. IL-1 beta significantly upregulated cell associated 14 kDa PLA2 and its subsequent release. Specific inhibition of this enzyme by a neutralizing Mab or selective inhibitors of 14 kDA PLA2 activity did not alter IL-1beta induced PGE2 levels.
Conclusion:
These data directly support a role for the 85 kDa PLA2, but not the 14 kDa PLA2, in IL- 1beta stimulated PGE2 production from human rheumatoid synovial fibroblast.