lli~:t'h#~tit a t't llioph).~icu/h't ~. I 1] "4( I qq2 ) 142-14~1 ,v, It.Jq.~2Elsevier Science Publishers B.V. All rights reserved 0J 67-488~.t/~,~2/S05.(111

BI"IAMCR 13075

Enhancement of platelet 12-HETE production in the presence of polymorphonuclear leukocytes during calcium ionophore stimulation R o s s K. M c C u l l o c h , K e v i n D . C r o f t a n d R o b e r t V a n d o n g e n lhT~aronrnr o l Medicine. Uslhe~ity o]" I¢'e.~/ern Au.~u'olia~ Royvl Pt'r:h IhJstntuL ~,rth eAuslrdli# t (Received I May ]991) (Reviccd mannseripl received 27 August 1~'~1)

Key words: P'latelet: Polymorphonuclear leukocyte; Calcium ionuphore A23187: Eicosanoid; 12 HETE

This study investigates the effect of plulelet/neutrophil interactions on eicnsanoid production. Human platelets and polymorphonuclear leukoeytes (PMNs) were stimulated alone and in combination, with calcium ionophnre A23187 and the resulting eicosanoids 12S.hydroxy-(SZ,SZ,lOE,14Z)-eicosatetraenoic acid (12-HETE), 12S-beptadecaIrienoic acid (HHT), 5S,12R.dihydroxy-(6Z,SE,lOE,14Z).eieosatelraenoie acid (LTB 4) and $.~-hydruxy(6E,8Z, I IZ,14Z)-eicosatetraenoic acid (5-HETE) were measured by HPLC. The addition of PMNs to platelet suspensions caused a 104% increase in I2-HETE, a product of lZ-lip0xygenase aclivity~ but had only a modest effect on the cyeiooxygenase product HIlT (increase of 18%). By using PMNs labelled with [t4Ciarachidonic acid it was shown thal the increases in these platelet eicusanoids could be accounted for Ir8 trauslocatlon of released araehidonie acid from PMNs to platelets and its subsequent metabolism. The observation that 12-1ipoxyAeoase was about five times more efficient than cyeloaxygenasc at utillsing exogenous arachidonic acid during the platelet/PMN interactions was confirmed in experiments in which plate[els were stimulated with A23187 in the presence of [t4Clarachidonie acid. Stimulations of platelets with thrombin in the presence of PMNs resulted in a decrease in 12.ItETE ~nd HHT levels of 40% and 26%~ respectively. The presence of platelets caused a small increase in neutrophil LTB 4 output but rc~u|te# in a decrease in S-HETE production of 43% during stimulation with A2,3187, This study demonstrates complex biochemical interactions be.tween plalelets and PMNs during elcosanoid production and provides evidence of a mechanism In explain the large enhanceme,i," .;.n 12-HETE production. Introduction During an inflammatory response polymorphonuclear leukocytes (PMNs) become activated and release various mediators such as PAF and leukotriene(LT)B 4 which recruit further PMNs and other cell types ~uch as platelets to the site of injury or attack. With these cells in close proximity and in a state of activation the potential exists for the metabolltes of one ceil type to be modified by another. Some recent studies have shown that both PMNs alld plalelets have the enzYme systems to mctabolisc products produced by the reciprocal cell type, in particular those derived from arachidonic acid, For example, it has been reported that

Correxp~ndence: R,K. McCulloch. Dcparlraent of Medicine. University pf Western Attxtta~iu. Royal Perth HospJli~], Perth. Western Attstrali;~. 60 ~~"~.

PMN-derived LTA 4 is ,ff~cientty converted to LTC# by piatel~t~ [l.2] and similarly 5-HETE, another major lipoxygcnase ~reduct of ihe aeutrophil is metabolised to 5S,t2S-dihydro.,cy4fiE,?,Z, lOE,14Z)-eieosatetra. enoic acid (5,12-diHETE) by platelets [3,4]. The reciprocal reaction also occurs in which 12-HETE released by the platelet is transformed into 5,12-diHETE by 5-1ipoxygenase [3]. Other examples of this co-operation between cells is the conversion of LTA4 by erythrocytes to LTB4[5] and the metabolism of 12-HETE by unstimulated PMNs to 12S,20-dihydroxy-(5Z,8Z, lOE, 14Z)-eieo:;aletraen0ic acid (12,20-diHETE)[6]. Another meanz by which intercellular eo-opcratlvity may occur is when the product from one cell stimulates (or inhibits) the enzyme of another and in this respect it has been shown that platelel-derived 12S~hydroperoxy(5Z,SZ, lOE,14Z )-eicosatetraenoic acid (12-HPETE) stimulales PMN 5qipoxygenase, the enzyme involved in the synthesis of LTB 4 and 5-HETE [7].

143 To fi]rlhcr investigate these cellular interactions we have examined the effect uf PMN~ on the synlhcsis of two p[atclet cicosanoids, 12-HE'rE at lipoxygcnasc product and HHT derived t'mm cychmxygcnasc, daring activatkm by calcium ionuphorc A23i87 which generates metabolites in sufficient quantity to bc readily detectable by HPLC. A proposed mechanism t . explain the observed increases in I]tcsc products is al:;o presented. The results of sprat :cciproce! ~.xpcrimc,

Enhancement of platelet 12-HETE production in the presence of polymorphonuclear leukocytes during calcium ionophore stimulation.

This study investigates the effect of platelet/neutrophil interactions on eicosanoid production. Human platelets and polymorphonuclear leukocytes (PMN...
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