Ethanol dependent interaction between prostaglandins and lipoxygenase products in human peripheral lymphocytes.

The effect of lipoxygenase products, 12-L-HETE and 15-L-HPETE, on cyclic AMP levels in human peripheral lymphocytes was examined in the absence and in the presence of a prostaglandin of the E-type (0.6-3.0 microM) or isoprenaline (33 microM). The studies were performed either in the absence or in the presence of 6 per cent ethanol. For comparison the effect of arachidonic acid and linolenic acid were studied. In the absence of ethanol 12-L-HETE and 15-L-HPETE had no significant effect on cyclic AMP accumulation. However, in the presence of ethanol 12-L-HETE (above 1 microM) inhibited prostaglandin E1 but not isoprenaline induced cyclic AMP accumulation. 15-L-HPETE had a biphasic effect on prostaglandin E2 induced cyclic AMP accumulation. Concentrations below 1 microM stimulated, those above inhibited. Virtually complete inhibition was seen at 15 microM. The two other fatty acids inhibited both prostaglandin E2 and isoprenaline induced cyclic AMP accumulation in the presence, but not in the absence of ethanol. The results show that lipoxygense products have little or no effect on cyclic AMP accumulation in human peripheral lymphocytes unless ethanol is present. In the presence of ethanol both 12-L-HETE and 15-L-HPETE appeared to selectively affect the cyclic AMP accumulation stimulated by PGE.

Effect of 5'-methylthioadenosine, 3-deazaadenosine, and related compounds on human natural killer cell activity. Relation to cyclic AMP and methylation potential.

The effect of 5'-methylthioadenosine (MTA) on human natural killer (NK) cell activity was examined and compared with the effect of 3-deazaadenosine (c3-ado) and periodate-oxidized adenosine (ado-ox). MTA inhibited NK cell activity in concentrations above 30 microM, but in concentrations below 10 microM a slight enhancing effect was often observed. C3-ado and ado-ox were 10 and 3 times more potent, respectively, as inhibitory agents and did not increase NK cell activity in low concentrations. The inhibitory effect of c3-ado was unaffected by preincubation of the cells but was enhanced by the addition of L-homocysteine. In concentrations that caused inhibition of NK cell activity all three agents caused a fall in the methylation index (AdoMet/AdoHcy) but no or an inconsistent effect on the level of cyclic AMP. An increase in the level of AdoHcy was observed already after 1 h of incubation but was more pronounced after 4 h of preincubation with the adenosine derivatives. The inhibition of cytotoxicity was mainly on their initiation of lysis, with a smaller effect on target cell binding. Antibody-dependent cellular cytotoxicity and lectin-dependent cellular cytotoxicity appeared to be less sensitive to inhibition by c3-ado. Our results show that several adenosine analogues inhibit NK-cell-mediated cytotoxicity in parallel with a decreased methylation index. The results suggest that a methylation step is critical in lymphocyte-mediated cytotoxicity and that NK cell activity is more sensitive to inhibition of this step than antibody- or lectin-dependent cytoxicity.

Inhibition of human NK cell cytotoxicity by induction of cyclic AMP depends on impaired target cell recognition.

Induction of cyclic AMP (cAMP) depresses natural killer (NK) cell activity. The present results demonstrate that this is dependent on a decreased capacity of the effector cells to conjugate to target cells. This was found either if dibutyryl-cAMP was used or if cAMP was induced by adenylate cyclase stimulation with prostaglandin E1 (PGE1) or by inhibition of phosphodiesterase activity with the inhibitor ZK 62711. The sites of action for cAMP-induced NK suppression and interferon (IFN)-induced NK enhancement are demonstrated to be distinct, since IFN acts by increasing the lytic efficiency and the recycling capacity without influencing target binding. Sequential treatment with cAMP/IFN and IFN/cAMP shows that IFN can neither restore target binding when added after cAMP nor protect against the cAMP-induced target binding inhibition when added before cAMP. The results are discussed in view of earlier data on cAMP in relation to cell membrane functions and cellular recognition, the mechanism underlying the cAMP-induced target binding inhibition, and the potential of the NK system as an indicator for immunosuppression. The present work also demonstrates the particular subpopulation in peripheral blood which mediates most NK activity, to respond strongly to PGE1 stimulation with regard to cAMP induction.

PGE1 and prostacyclin suppression of NK-cell mediated cytotoxicity and its relation to cyclic AMP.

The capacity of three prostanoids (PGE1, 6-beta-PGI1, PGI2 or prostacyclin) and a phosphodiesterase inhibitor (rolipram) to inhibit NK ("natural killer") cell cytotoxicity and to raise cyclic AMP levels in purified NK cells was compared. PGE1 was about 200 times more potent than prostacyclin both in its ability to raise cyclic AMP and to inhibit NK cell cytotoxicity. The stable prostacyclin analogue, 6-beta-PGI1, had an intermediate potency. A 50% inhibition of cytotoxicity was obtained at approximately 10(-8) M for PGE1, 10(-7) M for 6-beta-PGI1, and 10(-6) M for both prostacyclin and rolipram. These doses raised the level of cyclic AMP by approximately 100%. These results suggest that PGE1 is likely to be more important as an endogenous regulator of lymphocyte cytotoxicity than prostacyclin. The results also provide further evidence that cyclic AMP is the mediator of prostanoid-induced reduction in NK cell activity.

Comparison of the effects of different arachidonic acid metabolites on cyclic nucleotide accumulation in human peripheral lymphocytes.

The effect of PGE1, PGE2, PGD2, PGF2 alpha, PGI2, PGG2, PGA1, 12L-HETE, arachidonic acid, 15- HPETEa and linolenic acid on the accumulation of cyclic AMP in human peripheral lymphocytes was studied. PGE1, PGE2 and PGD2 were essentially equipotent as stimulators of cyclic AMP accumulation (threshold at about 10(-8)M and EC50 about 0.15 microM), PGF2 alpha was about 20 times less potent, while PGG2, 12L-HETE, 15-HPETE, PGA1 and linolenic acid were inactive. PGI2 caused a weak stimulation between 5 and 600 nM and a secondary stimulation above 3 microM. Arachidonic acid had no effect on cyclic AMP levels up to 100 microM. PGE1, PGD2, PGI2 and PGF2 alpha increased cyclic GMP in the concentrations that produced a rise in cyclic AMP, but the cyclic GMP increase was of smaller magnitude. Exogenous arachidonic acid was converted mainly to 12L-HETE, HHT and thromboxane B2 by lymphocyte suspensions. This conversion could be accounted for by contamination with blood platelets. The results show that the degree of cyclic AMP accumulation in human lymphocytes following stimulation of arachidonic acid metabolism will be critically dependent upon which prostaglandins are in fact formed by cells surrounding the lymphocytes.

Effects of ethanol on human lymphocyte levels of cyclic AMP. In vitro: Potentiation of the response to isoproterenol, prostaglandin E2 or adenosine stimulation.

The effects of ethanol, acetone and dimethylsulfoxide (DMSO) were tested on the accumulation of cyclic AMP in human peripheral blood lymphocytes in vitro. Isoproterenol (1.0X10(-8) - 1.0X10(-4)M), PGE2 (3X10(-8)-3X10(-6)M), adenosine (10(-7) - 10(-4)M) and phenylisopropyladenosine (10(-8) - 10(-4)M) caused a dose dependent increase in cyclic AMP accumulation. Over the entire range of concentration of stimulating drugs, ethanol caused an enhanced accumulation of cyclic AMP. At temperatures between 15 degrees and 30 degrees the effect of ethanol rose with increasing concentration from 0.2-6%. At 37 degrees and 40 degrees, 6% ethanol had less stimulatory effect than 2% ethanol. The effect of ethanol was shared by acetone and to a minor extent by DMSO, and was present also when phosphodiesterase was inhibited by isobutylmethylxanthine. It is suggested that ethanol enhances adenylate cyclase activity as a consequence of altered cell membrane fluidity. Since the effects on cyclic AMP accumulation can be observed already at rather low concentration of the solvents they may be of toxicological significance.