INTERLEUKIN-1 RECEPTOR ANTAGONIST BLOCKS INTERLEUKIN-1-INDUCED EXPRESSION OF CYCLOOXYGENASE-2 IN ENDOMETRIUM

OBJECTIVE: Our purpose was to test the hypothesis that the interleukin -1 receptor antagonist can inhibit interleukin-1-induced prostaglandin production and de novo expression of the inducible cyclooxygenase-2 i soform in a human endometrial epithelial cell line. STUDY DESIGN: A co ntinuous line of human endometrial epithelial cells was established fr om a hysterectomy specimen from a nonmalignant uterus. Cells were main tained as a monolayer culture in medium 199 supplemented with 10% feta l bovine serum and 50 mu g/ml gentamicin. Cultures were treated with c ytokines (interleukin-1 alpha or interleukin-1 beta, interleukin-1 rec eptor antagonist, or tumor necrosis factor-alpha), and media were coll ected for analysis of prostaglandin E-2 and prostaglandin F-2 alpha) b y radioimmunoassay, whereas cells were harvested for ribonucleic acid and protein extractions and subsequent Northern blot or Western blot a nalyses, respectively. RESULTS: When endometrial cells were incubated with interleukin-1 alpha or interleukin-1 beta, each cytokine was show n to stimulate the production of prostaglandin E-2 and prostaglandin F -2 alpha in a time-and dose-dependent fashion, with interleukin-1 alph a being far more potent than interleukin-1 beta. Interleukin-1 recepto r antagonist inhibited interleukin-1 alpha- and interleukin-1 beta-ind uced prostaglandin formation, with 50% inhibitory concentration Values of 30 ng/ml for prostaglandin E-2 and 90 ng/ml for prostaglandin F-2 alpha. When Northern blots of interleukin-1 alpha-treated cells were p robed with a complementary deoxyribonucleic acid fragment specific for either cyclooxygenase-1 or cyclooxygenase-2, rapid de novo induction of cyclooxygenase-2 messenger ribonucleic acid was observed; however, cyclooxygenase-1 expression was constant regardless of interleukin-1 a lpha concentration or incubation time. Coincubation of cells with inte rleukin-1 alpha (10 ng/ml) and cycloheximide caused superinduction of cyclooxygenase-2 messenger ribonucleic acid but had no effect on the e xpression of cyclooxygenase-1 messenger ribonucleic acid. Actinomycin D completely abolished interleukin-1 alpha-induced cyclooxygenase-1 me ssenger ribonucleic acid expression, suggesting that the cytokine caus ed transcriptional activation of the cyclooxygenase-2 gene. Experiment s were conducted to examine whether interleukin-1 receptor antagonist could suppress interleukin-1-induced cyclooxygenase-2 expression. Cell s were preincubated for 30 minutes with interleukin-1 receptor antagon ist and then challenged with interleukin-1 alpha. Northern and Western analyses revealed that interleukin-1 receptor antagonist blocked inte rleukin-1 alpha-induced expression of cyclooxygenase-2 messenger ribon ucleic acid transcripts and the subsequent appearance of cyclooxygenas e-2 protein. Interleukin-1 receptor antagonist had no effect on the co nstitutive expression of cyclooxygenase-1 messenger ribonucleic acid a nd protein. Interleukin-1 receptor antagonist railed to alter prostagl andin E-2 formation in response to tumor necrosis factor-alpha, indica ting that the antagonist is specific for interleukin-1 family cytokine s. Finally, interleukin-1 receptor antagonist acted as a partial agoni st in some experiments in that relatively high concentrations (>100 ng /ml) caused a modest increase in prostaglandin E-2 and F-2 alpha produ ction. CONCLUSIONS: These data indicate that interleukin-1 receptor an tagonist is a potent inhibitor of interleukin-1-induced arachidonic ac id metabolism and could possibly serve as an endogenous or exogenous m odulator of interleukin-1 action in the endometrial epithelium.