Regulation of expression of matrix metalloproteinase-9 in early human T cells of the HSB.2 cultured line by the EP(3) subtype of prostaglandin E(2) receptor

The expression by T lymphocytes (T cells) of more than one of the functionally distinct subtypes of prostaglandin E(2) (PGE(2)) receptors (Rs), designated EP(1), EP(2), EP(3), and EP(4) RS, is a principal determinant of specificity and diversity of the immune effects of PGE(2). The cultured line of human leukemic T cells, termed HSB.2, co-expresses a total of 7282 +/- 1805 EP(3), EP(4), and EP(2) Rs per cell with a K-d of 3.7 +/- 1.4 nM (mean +/- S.E., n = 9), The EP(3)/EP(1) R-selective agonist sulprostone, EP(3)/EP(2)/EP(4) R-selective agonists M&B 28767 and misoprostol, and EP(2) R-selective agonist butaprost but not the EP(1) R-selective antagonist SC-19220 competitively inhibited the binding of [H-3]PGE(2) to HSB.2 cells, Stimulation of increases in the intracellular concentration of cyclic AMP ([cAMP](i)) by PGE(2), misoprostol, and butaprost and of increases in the intracellular concentration of calcium ([Ca2+](i)) by PGE(2) and sulprostone demonstrated the respective involvement of EP(2)/EP(4) Rs and EP(3) Rs in transduction of biochemical signals, Matrix metalloproteinase (MMP)-9 was identified by zymography and Western blots as the principal MMP secreted by HSB.2 cells, The cytosolic level and secretion of MMP-9 were increased maximally after 24 h of incubation of HSB.2 cells with 10(-8)-10(-6) M PGE(2), sulprostone, M&B 28767, and misoprostol but not with 10(-6) M PGF(2 alpha), PGD(2), PGI(2), or butaprost, suggesting a principal dependence on EP(3) Rs. That stimulation of MMP-9 secretion by PGE(2) was not diminished in Ca2+-free medium but was suppressed significantly and dose-dependently by thapsigargin, an inhibitor of endomembrane Ca2+-ATPase, suggested that MMP-9 expression by HSB.2 cells is mediated by increases in [Ca2+](i) attributable to release of Ca2+ from intracellular stores, The lack of effect of dibutyryl cAMP, forskolin, and SQ 22536, an adenylyl cyclase inhibitor, on MMP-9 secretion by HSB.2 cells argued against any role for cAMP-dependent mechanisms linked to EP(2)/EP(4) Rs, Cycloheximide and actinomycin D, which respectively inhibited protein and RNA synthesis, suppressed basal and PGE(2) induction of MMP-9 production by HSB.2 cells, Northern analysis indicated that PGE(2) and sulprostone time-dependently increased expression of MMP-9 mRNA, Thus, stimulation of MMP-9 in HSB.2 T cells by PGE(2) is attributable to [Ca2+](i)-dependent EP(3) R-mediation of increases in message transcription.