Lipoxin A-induced inhibition of human natural killer cell cytotoxicity: studies on stereospecificity of inhibition and mode of action

Human leukocyte-derived lipoxin A (LXA; 5S,-6R,15S-trihydroxy-7,9,13-trans-11-cis-eicosatetraenoic acid) inhibits the cytotoxic activity of human natural killer (NK) cells. LXA and three of its isomers were prepared by total organic synthesis and assayed for activity with human NK cells. Dose-response studies showed that biologically derived LXA and synthetic LXA were equally effective in inhibiting NK cell cytotoxicity. 6S-LXA, with its 6S-OH group in an (S) configuration, proved to be approximately half as potent as LXA. In contrast, 6S-11-trans-LXA and 11-trans-LXA displayed virtually no inhibitory activities. The methyl esters of both LXA and 6S-LXA proved to be more potent than their corresponding free acids. Thus, LXA inhibition of NK cells displays clear-cut stereochemistry. In the absence of putative inhibitors, NK cells bind to their targets to form conjugates. This event is followed by polarization of the NK Golgi apparatus, which moves towards the plasma membrane that is in contact with the target cell. However, in the presence of either the methyl ester or free acid of LXA, the Golgi apparati of NK cells bound to their targets were randomly oriented. In contrast, neither 6S-11-trans-LXA nor the potent NK inhibitor prostaglandin E2 affected the polarization. Furthermore, although prostaglandin E2 resulted in a decrease in NK-target cell binding efficiency, LXA and its isomers failed to affect conjugate formation. Together these results indicate that LXA-induced inhibition of NK cytotoxicity does not act on NK cell binding but may block cytotoxicity by disrupting "signals" involved in the specific orientation of the Golgi. Thus, this latter event may appear to be important in cytotoxicity.     

The action of lipoxin-A on glomerular microcirculatory dynamics in the rat

Intrarenal administration of 750 ng/kg/min of LX-A in euvolemic rats resulted in significant increases in single nephron GFR (38.4 +/- 1.7 to 45.5 +/- 3.0 nl/min) and plasma flow rate (95 +/- 6 to 127 +/- 9 nl/min). The latter was due to a dramatic fall in afferent arteriolar resistance. Mean transcapillary hydraulic pressure difference increased from 33 +/- 1 to 43 +/- 3 mmHg (p less than 0.05) and the glomerular capillary ultrafiltration coefficient fell from 0.060 +/- 0.013 to 0.033 +/- 0.005 nl/(s X mmHg) (p less than 0.05). These responses to LXA in the renal microcirculation are in sharp contrast to those previously observed for the leukotrienes, and thus may represent the first example of counterregulatory (constrictor/dilator) vascular interactions within the lipoxygenase pathways.     

Identification of a novel 7-cis-11-trans-lipoxin A4 generated by human neutrophils: total synthesis, spasmogenic activities and comparison with other geometric isomers of lipoxins A4 and B4

Addition of (15S)-hydroxy-5,8,11-cis-13-trans-eicosatetraenoic acid (15-HETE) and the ionophore A23187 (2.5 microM) to human neutrophils led to the formation of both lipoxin A4 and lipoxin B4 as well as a novel 5,6,15-trihydroxyeicosatetraenoic acid. The new compound was identified using an improved isolation and detection system and its basic structure was determined by physical methods. On the basis of biosynthetic considerations, geometric isomers of lipoxin A4 and lipoxin B4 were prepared by total synthesis. Comparison of these synthetic materials with the neutrophil-derived product showed that the new compound is (5S,6R,15S)-trihydroxy-9,11,13-trans-7-cis-eicosatetraenoic acid or the 7-cis-11-trans-isomer of LXA4 (7-cis-11-trans-LXA4). LXA4, 11-trans-LXA4, 7-cis-LXA4 and 7-cis-11-trans-LXA4 all evoked dose-dependent (0.1-10 microM) contractions of the guinea pig lung strip, whereas 6-cis-LXB4 and 6-cis-8-trans-LXB4 relaxed this preparation. LXA4 and 7-cis-LXA4 were approx. 10-times more potent than the compounds with 11-trans geometry. However, all four double-bond isomers of LXA4 caused contractions which, based upon pharmacological evidence, appeared to involve specific activation of the same site as cysteinyl-containing leukotrienes. In conclusion, 7-cis-11-trans-LXA4 was isolated and identified as a novel biologically active eicosanoid formed by human neutrophils.     

A Drosophila hsp70 gene contains long,antiparallel, coupled open reading frames (LAC ORFs) conserved in homologous loci

A clone isolated from a Drosophila auraria heat-shock cDNA library presents two long, antiparallel, coupled (LAC) open reading frames (ORFs). One strand ORF is 1,929 nucleotides long and exhibits great identity (87.5% at the nucleotide level and 94% at the amino acid level) with the hsp70 gene copies of D. melanogaster, while the second strand ORF, in antiparallel in-frame register arrangement, is 1,839 nucleotides long and exhibits 32% identity with a putative, recently identified, NAD⁺-dependent glutamate dehydrogenase (NAD⁺-GDH). The overlap of the two ORFs is 1,824 nucleotides long. Computational analysis shows that this LAC ORF arrangement is conserved in other hsp70 loci in a wide range of organisms, raising questions about possible evolutionary benefits of such a peculiar genomic organization.Correspondence to: Z.G. Scouras     

Synthesis and biological properties of 12-fluoroprostacyclins

The synthesis of the sodium salts of enantiomerically pure 12-fluoroPGI₂ (9), (±)-12-fluoroPGI₂ (9), (±)-15-epi-12-fluoroPGI₂ (10), (±)-12-fluoro-13,14-dihydroPGI₂ (11), (±)-12-fluoro-4(E)-isoPGI₂ (12), and (±)-5,6-dihydro-12-fluoroPGI₂ (13) is detailed starting from the corresponding derivatives of 12-fluoroPGF_2α methyl ester. Prostacyclins 9, (±)-9, (±)-10, (±)-11, (±)-12, and (±)-13 have been evaluated for their ability to inhibit human platelet aggregation and their effect on smooth muscle (isolated cat coronary artery).     

Circulatory and platelet actions of 6,9-thiaprostacyclin (PGI2-S) in the cat.

A new analog of prostacyclin, 6,9-Thiaprostacyclin was infused intravenously in pentobarbital anesthetized cats in order to determine its hemodynamic and anti-platelet aggregating properties. At an infusion rate of 0.01 μmoles/kg/min, PGI₂-S moderately decreased arterial blood pressure without altering heart rate of superior mesenteric artery flow or platelet aggregation responses to ADP. However, at 0.05 μmoles/kg/min, PGI₂-S significantly reduced arterial blood pressure and significantly increased heart rate, and superior mesenteric artery flow. Moreover, at 0.05 μmoles/kg/min, PGI₂-S inhibited ADP platelet aggregation by 80%. PGI₂-S may be a useful agent in circulatory shock.     

Lipidomic analysis reveals prostanoid profiles in human term pregnant myometrium

Prostanoids modulate the activity of human pregnant myometrium and their functional role can be appreciated through characterisation of prostanoid receptors and tissue concentration of prostanoids. We have applied a lipidomic approach to elucidate the profile of prostanoids in human non-labouring and labouring myometrium. We have identified a total of nineteen prostanoids including prostacyclin, thromboxanes, prostaglandins and dihydro-prostaglandins. Prostacyclin was the predominant prostanoid in both non-labouring and labouring myometria, with PGD₂ and PGF_2α being the second most abundant. Although the total amount of prostanoids was increased in the labouring tissue, PGE₂ and 13,14-dihydro-15-keto-PGE₂ were the only prostanoids to increase significantly at early and late labour (p≤0.001). Our data suggest that PGF_2α plays an important role in parturition, whilst the increase in PGE₂ could occur to facilitate cervical dilation and relaxation of the lower myometrium during labour. Although the elevation in TXA₂ was less marked than expected, in terms of translation to function even a relatively small increase in the level of this potent spasmogen may have significant effects.     

Rainfall manipulation experiments as simulated by terrestrial biosphere models: Where do we stand?

Changes in rainfall amounts and patterns have been observed and are expected to continue in the near future with potentially significant ecological and societal consequences. Modelling vegetation responses to changes in rainfall is thus crucial to project water and carbon cycles in the future. In this study, we present the results of a new model‐data intercomparison project, where we tested the ability of 10 terrestrial biosphere models to reproduce the observed sensitivity of ecosystem productivity to rainfall changes at 10 sites across the globe, in nine of which, rainfall exclusion and/or irrigation experiments had been performed. The key results are as follows: (a) Inter‐model variation is generally large and model agreement varies with timescales. In severely water‐limited sites, models only agree on the interannual variability of evapotranspiration and to a smaller extent on gross primary productivity. In more mesic sites, model agreement for both water and carbon fluxes is typically higher on fine (daily–monthly) timescales and reduces on longer (seasonal–annual) scales. (b) Models on average overestimate the relationship between ecosystem productivity and mean rainfall amounts across sites (in space) and have a low capacity in reproducing the temporal (interannual) sensitivity of vegetation productivity to annual rainfall at a given site, even though observation uncertainty is comparable to inter‐model variability. (c) Most models reproduced the sign of the observed patterns in productivity changes in rainfall manipulation experiments but had a low capacity in reproducing the observed magnitude of productivity changes. Models better reproduced the observed productivity responses due to rainfall exclusion than addition. (d) All models attribute ecosystem productivity changes to the intensity of vegetation stress and peak leaf area, whereas the impact of the change in growing season length is negligible. The relative contribution of the peak leaf area and vegetation stress intensity was highly variable among models.