Prostaglandin E2 is a potent regulator of interleukin-12- and interleukin-18-induced natural killer cell interferon-gamma synthesis.

Synthesis of interferon (IFN)-gamma by natural killer (NK) cells is an important pro-inflammatory event with interleukin (IL)-12 and IL-18 playing major inductive roles. However, other temporal events are likely to regulate such processes and as prostaglandin E2 (PGE2) is ubiquitous during inflammation this study tested the hypothesis that PGE2 was capable of directly modulating cytokine-induced NK cell IFN-gamma synthesis in the absence of other immune cells. Using homogeneous NK cell lines to establish direct effects, PGE2 (0.1-1 micro m) was found to suppress NK cell IFN-gamma synthesis and antagonized the potent synergistic IFN-gamma-inducing effects of IL-12 and IL-18. The actions of PGE2 were mimicked by synthetic PGE2 analogues including misoprostol and butaprost. The selective EP2 receptor agonist butaprost, but not the EP1/EP3 agonist sulprostone, suppressed IFN-gamma synthesis and exclusively competed with PGE2 for receptor binding on NK cells. Further analysis showed that PGE2 did not modulate IL-12 receptor mRNA expression and the effects of PGE2 could be mimicked by the phosphodiesterase inhibitor 3-iosobutyl-1-methylxanthine. The absence of demonstrable receptor modulation coupled with the observed suppression of IFN-gamma synthesis by both EP2 receptor-selective agonists and IBMX suggest that PGE2 acts directly on NK cells via EP2 receptors with its downstream effects on cAMP metabolism. This conclusion is further supported by findings that PGE2 and its analogues consistently elevated levels of cAMP in NK cells. The ability of PGE2 to antagonize the potent inductive signal provided by the combination of IL-12 and IL-18 supports the concept that PGE2 may play an important role in limiting innate inflammatory processes in vivo through direct suppression of NK cell IFN-gamma synthesis.

An ELISA for PGE2 utilizing monoclonal antibody.

An ELISA for PGE2 has been developed which is sensitive to concentrations of 0.5 to 20.0 ng PGE2/ml. Mouse monoclonal anti-PGE2 ascites is utilized in a binding competition between the test sample and an adsorbed conjugate of PGE2-BSA. The antibody which remains bound to the solid phase is quantitated colorimetrically by incubation with alkaline phosphatase-conjugated goat anti-mouse IgG followed by incubation with p-nitro-phenylphosphate. PGE1, PGA1, PGA2, PGB2, 6-keto-PGF1 alpha, PGF2 alpha, 13,14-dihydro-15-keto-PGE2, thromboxane B2 and arachidonic acid showed minimal cross-reactivity with the anti-PGE2. The PGE2 ELISA permits the quantitative analysis of large numbers of samples at a fraction of the cost and time required to process a commercial RIA kit. When linked to the appropriate computer software, data collection and analysis can be performed in less than 10 minutes per 96-well plate. Furthermore, the use of an ELISA system eliminates the radioactive and toxic chemical waste generated by RIA methods.

Regulation of human natural killer (NK) cell function: induction of killing of an NK-resistant renal carcinoma cell line.

Natural killer (NK)-like activity against a renal carcinoma cell line, Cur, was assessed. There was no spontaneous killing of Cur cells by human peripheral blood mononuclear cells in 4-hr assays. Cur killing was observed in 18-hr assays, but the magnitude of killing was variable and always markedly less than that against K562. Cur killing was mediated by a nonadherent, nonphagocytic lymphocyte, the activity of which could be modulated both positively and negatively by monocytes or their products. Preincubation of effectors with monocyte supernatant, interleukin 1 (IL-1), alpha-interferon (alpha IFN), or interleukin 2 (IL-2) greatly increased the magnitude of Cur killing and accelerated the kinetics of lysis. The addition of prostaglandin E2 (PGE2) during in vitro activation of NK by IL-2 profoundly inhibited subsequent Cur lysis, whereas only minimal inhibition of K562 lysis was noted. However, following activation with IL-2, lysis of Cur targets was less sensitive to the inhibitory effects of PGE2. Removal of Leu 11b(+), OKM1(+), or L-leucyl-leucine methyl ester-sensitive cells markedly decreased both Cur and K562 lysis. Moreover, CD16(+) cells purified with the fluorescence-activated cell sorter were found to mediate Cur killing. Whereas Cur and K562 lysis is mediated by phenotypically similar effector cells, the present studies demonstrate that the cytotoxic functions defined by the ability to lyse these two targets differ in response to a variety of immunoregulatory stimuli.

Macrophage function in the Schistosoma mansoni egg-induced pulmonary granuloma. Role of arachidonic acid metabolites in macrophage Ia antigen expression.

The ability of arachidonic acid (AA) metabolites to regulate I-region-associated (Ia) antigen expression on macrophages from schistosome-egg-induced pulmonary granulomas was examined. The prostaglandin (PG) analog 15-S-15-CH3-PGE1 (M-PGE1) and PGF2 alpha were found to modulate the kinetics of Ia expression when administered in vivo. Methyl-PGE1 significantly suppressed Ia antigen expression by hypersensitivity granuloma macrophages, while PGF2 alpha appeared to potentiate the expression. Lymphokine-induced Ia antigen expression by cultured granuloma macrophages was likewise dramatically inhibited by M-PGE1. Further analysis using systemically administered inhibitors of AA metabolism demonstrated that the cyclooxygenase inhibitor indomethacin caused augmentation of Ia expression. In contrast, lipoxygenase inhibitors significantly reduced both Ia expression and granuloma size. The role of AA metabolites in modulating chronic inflammation is discussed.