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Chemotactic activity of thromboxane B2, prostaglandins and their metabolites for polymorphonuclear leucocytes
PROSTAGiANDINS
ME!l'ABOLITESF0RPOLYMXPBCNDCLEMR
E. Ann Kitchen, J. R. Boot, W. Dawson
Lilly F+asearch Centre Limited ErlwoodManor
Abstract
(1)
The cheemtacticactivitiesof thrcmbaxaneB2 (m2)r =2, PGF2c(,the 15-oxo, 15-oxo-13,14-dihydro and 13,14-dihm metabolitesof PSE2, PGF2ar and a mataboliteof TxB2 for polymrphonuclearleumcytes (PMV)havebeen investigated.
(2)
ThrmaxmaneB2 increasedthe directicmalmigrationofrat peritonealPI@Iataamcentrationof 2.Ow/ml audofhman peripheralneutrophilsat a concentrationof 0.5 rJ,s/ml.
(3) Neither PCZ2, PGF2a nor their mtabolites shm.ed chemtactic activity for rat peritonealm. (4) PGF2a and 15-oxc~13,14-dihydro-t chemtactic activity forhmanperipheral
B2 showed no M.
(5) The possible role of thxnboxaneB2ininflamnaticmisdiscussed. Introduction Prostaglantis (PGs) havebeen iqlicatedinmanyoftheevents whichoccurduringanacute inflamnatoryreaction. chleof themany roles attributedto a minority of the prostaglandinsis that of cherlotaxis. Mst of the studies carried out on the chexrotactic prcpwties of theprostaglandins have involvedtheuseofhman periphe.ralbloodPFNandresultshave generally shcm that the par&z PGs of the A B D E and F series are not cheaotacticfor either mononuclearcells or PMV (1,2,3). PGEl,hmever, ischemtactic for rabbit peritmeal M (4,5)but not for rabbit peripheralblood Pm nor for rat PMV derived f_m either peripheralblood or peritoneum (6). HE!l?E(12-Lhydmxy-5,8,10,14 eicosatetraenoicacid) hasbeenshawntobecherrotacticforhua~nblood~s(7).
AUGUST
1978 VOL. 16 NO. 2
239
PROSTAGLANDINS
ThrmboxaneB2has previously been shmn tobe chemYzacti.c for museperitonealm in vitroand tc causethe accum lationof rat PIN in vim after I.D. injection(8). 'Iheaimof thispresentstudywas toextendthese findingsand assessthe chemtacticactivityof TxB2 in vitro for rat peritoneal PkN and forhmanperiphe-calbloodPMJinordertoestablishwhether TxB2ismre generallychemtacticthan,forinstance,FCElwhich appearsto be specificfor rabbitP&N. Inaddition,we have investigated the chemtacticactivities of PG2 and PGF2cr for both rat andhumanbloodPM\Iand15-oxo-13,14-dihydro-TxB2 forhman P&N. Method.5
thrmboxaneB2 and the Thranboxane B2, 15-oxo-13,14-dihydroE-oxo, E-oxo-13,14-dihydx-o and 13,14-dihyd~o mtabolites of m2 and PGF2awerep~paredbiosyntheticallyand characterisedby GLCmass spectrcmatqas describedpreviously(9). PGS2 andPGF2,were obtai.nedfrmCambrianChemicals.Glycogen(exoyster)wasobtained frm Koch Light. Preparation of cell suspensionsA mixedpopulationof cellgcontaining 60-70%PkN,was harvestedfmantheperitinealcavityof rats (Lilly Lodge&&Wistar, male, 150-170g bodyweight) 24 h-afteran intraperitoneal injectionof2 mls 2% glycogenin saline. H~P~wereo$tainedfrantheperipheralbloodofhealthyfiale volunteersand the red blood cellssedimentid with 0.5% methyl cellulosefor 45/60mins. Chemtaxis Assay Chemtaxis was studiedin vitrousing a modification of theE!oydenchanbertechniqueas describedby Wilkinson (10). Polycarbonate filterswith a pore size of 5.0 p (Nuclepore Corporation) wereused. CellswerewashedandsuspendedinGeysbalancedsalt solution4BS.S) , pH 7.3, to give a finalconcentration of 5.0 x 106 cellsml- andO. mls of this cellsuspensionwereplacedin the upperchanber. Agentsundertestwerepreparedin Geys BSS and of TxB2 were used placedin the lmer chanber. Threepreparations rat serm randanlythroughouttheseexperin-mts.Zymsan-activated (11)(ZAS)at a concentration of 5% was used in all testsas the positive chesotactic amtmland Geys BSS as thenegativecontrol.In experiments todistinguishbetween chataxis and&emkinesis (stimulabsdrandm novement)agents~~rtestwereaddedtobothupperandl~~~s. At the end of this Thechanberswereincubatedat3~ for 2h. time,the filtersweresirmltaneouslydetachedandfixed,byimrr?rsion in 95% ethanolfor a minimm periodof 60 min and stainedwith and absolutecountswere carriedout on haematoxylin.Differential cellswhichhadmicratedto thelmer surfaceof'chefilter. Between 4-6 filterswere u&d for each sampleand a minimm of 10 fieldsper filterwerecomtedatamagnificationof 630X.
240
AUGUST 1978 VOL. 16 NO. 2
PROSTAGLANDINS
*suits
The effectof !I%32 cm the migrationof rat peritonealPM+ is sumwisedin table1. TxB2 inueasedthemigrationofrat PMNat 2.0 &ml but failedto show any effectat loweromcentrationsof 1.0 and 0.5 w/ml. These findingswaresimLlarineachofthree expsriments c!axried out. Arandcanmigraticn-~studysha&that this effectwasdue to an jmxease inthedirectionalmigraticm (dmmtaxis) in additiontoa slight increase in c!henokinesis. Neither-2 at 1.0 &ml nor PGF2u at 2.0 and 1.0 w/ml nor the metaboliteof !CxB2at 5.0 and 10.0 pg/ml shmed chemtacticactivity for PIW. Similarlynoeffectwasseenwiththe oxreqxmding matabolites of thesePGswhen testedat2.0 w/ml. Arachidonic acid at50 &mlwas inactivein this system.
The effectof TxB2 on hman PW is shrmnin table 3. TxB2 inmaased the directimalmigrationof cellsatboth0.5 and 2.0 w/ml. andatthelmarconoxtrationalso increasedrandmmigraticn. These findingswzresimilarineachofthreeexperimnts carriedout. Failureto increaserandcmnuvmantatthehighercmcentrationmy have been due to the disruptive effectof TxB2. Cellstich had migratedthroughthe filtertcwardsTxB2 at 2.0 w/ml showedsignsof disintegration andpyknoticnuclei. This effectwas also seen in thosecellsremainin g on theupper surfacebutti amuchlesserextent. NeitherTxB2 at 2.0 &ml nor 15-oxo-13,14-dih@ro-TxB2 at 10 w/ml had any significant effecton the migrationof nmonuclear cells. Prostaglandins E2 and F2(x(at 2.0 w/ml) and the 156x0-13,14dihydronkatabolite of thrceboxane B2(10w/ml) were ineffective in inducingchermtaxis ofhumanperipheralPMV.
Prostaglandins E2, F2a, and their correspondingrmatabolites failed Inaddition, to show cheeotactic activityfor either ratorhmanPMV. themetaboliteofTxB2 exhibitedno&emotacticactivityforhumanpEn. Hcwever,TxB2 shcweda markedchmotacticeffectfor hmmn PMI in vitro and also,althoughto a lesserextent,for rat PMi. The latterfinding oxrelateswithourprevious &se.rvations in z)it)o, that PIW accumlate in the blood vesselsof rat dermisafter localinjectionof thrcuboxane B2, and shouldbelinkedwith the~ork of others (12)thatTxB2 is genera~inlargeanouhtsfrcmth_mboxane A2 duringplatelet aggregation.Inaddition,evidencefromthesestudiesusingmuse (8) ratandhurnan~wouldindicatethat~~~ca~~tyofTxB2is not speciesspecific. TnecherrotacticactivityofTxB2isofpartia*lar interestinviewof a recent report (15)that anti-ihflmnatorydxugs can inhibitits formationin a carrageenin-induced granulcmain rats.
AUGUST 1978 VOL. 16 NO. 2
241
PROSTAGLANDINS
Table
I
Ohemtactic effect of TxB2 for rat peritoneal pclymrphonuckar leucocytes
Lowerchamber
No. of filters
zzs(Z T-2, (0.5 Klrn) 7-2, 7=2r
(1.0 (2.0
w/ml, w/m
7 6 12 12 14
Mean no. of PMN migratingper field 4.3 f 1.6 64.7 -t10.3 6.4 f 0.8 7.0 f 0.9 14.7 + 1.8
P
Table II Cutparisonof theeffect ofTxB2 on the randcxnand directimal migration of rat peritonealpolymxphonuclem leucccytes Mean no. cells/field Directional migraticm migration krpperandlmer charbar) (lowerchanbsr alone)
ChanberCcntents
1.9 f 0.5 19.2 * 4.1 4.8 + 0.5
Geys=S ZAS (5%) T-2 (2.0 w/ml) *
64.6 -I9.1* 17.4 f 1.5*
SignificantdifferenceP
Table III Ohmtactic effect of M2 for hmen peripheral polymrphonuclear leucocytes &an no. cells/field
(zhambx
1
contents
P"
1 (
*
242
Significantdifferencebetween mean values of randan and directionalmigration
AUGUST
1978 VOL. 16 NO. 2
PROSTAGLANDINS
Itappears tinthatTxB2 my cmtribute to the initialinfluxof m into an inflamnatory site,but it is unlikelythat it is the sole factorinvolvedsincetheamcentrationsusedintheseexperimnts were high. Fur&mmre,ccsparisonintheseexpfzimantswith!ZAS indicatesthatTxB2isamchwx&er chemotaxinthauactivated cqxnentsof cosplemant, whichmay be expe&ed to play a major role in cell accmulation. It is interestingthatbothZAS andTxB2 increasedrandmnDvemntinthesestudies. TxB2 is also releasedby phagocytosinghmsnperipheral blcodm (13)in mounts inexcess of tboseproducedbyresting~ and alsobyphagocytcsingrabbit peritonealcells (14). Itispossible thathrB2 maybe fmctioning as an inhibitorofxnigration of PPN alreadyin the tissues,rather thanas aprimaryrrediator of chemtaxis. Acmsistentbutlailevel of releaseduringactivephagocybosis couldmaintaina polymrphcmuclear leucocytepopulatim at the inf1amatoz.y focusprior to the influxof mmmuclear cells. TxA2,PGG;!orPGB2mayhave activitysimilarto,orgreaterthan, TxB2 in thesesystemsbut theirshorthalf-livesin aqueoussolution preventstheirexaminaticm in in vitro chemtsxis experixents.For these ampounds to have a significant effectin uivo,therewould need tobe acontirmus synthesisbythe cellsin the inflamatorysite in or&x to mintsin mncentraticm necessaryfor biologicalactivityof thiskind. Conclusion ~~B2hasbeen~tobe~adicforbothratand humanpolymrphcmuclear leucocytes in vitroand thesedata correlated with similarfindingsin in uivo studiesin the rat. It is suggestedthatthrmboxaneB2, releasedduringplatelet aggregatim,maybe contributing to the accmulationof polynorphonuclear leuaxytesby retainingthesecellsat the inflmimatory site ratherthan by elicitingtheirinitialinflux.
References 1.
Turmr, S.R.,Gaqbell, J.A. and Lynn,W.S. J. Exp. I&d.,3, 1437 1975
2.
Pazzaglia, A., Barker,A., Warin,A.P., Greaves,M.W. Brit. J. Dematol., 96, 533 1977
3.
Mcclatchey, W. ard Snydeman, R. Prostaglandins, 12 No. 3., 415 1976 -
4.
Kaley,G. and Weiner,R. NatureNew Biology,234, 114 1971
5.
Biggs,G.A.,-1, E. and Youlten,L.J.F. Br. J. Phamac. 53, 539 1975
AUGUST 1978VOL. 16 NO. 2
243
6.
Walker, J.R., Smith, M.J.H., Ford-Hutchinson,A.W. J. Pham. Phannac.,28, 745 1976
7.
Gcetzl, E.J., woods, J.M., and Goman, R.R. J. Clin. Invest.,2, 179 1977
8.
Boot, J.R., Dawsm, W. and Kitchen, E.A. J. Physiol.257, 47-48P 1976
9.
Dawson, W., Boot, J.R., Codxrill, A.F., Mallen, D.N.B. Osbcrne, D.J. Nature, 262, 699 1976
10.
Wilkinson,P.C. InCkmtaxis andInflamation,p. Edinburgh,Londcm. 1974
11.
Thaqxon, R.A. and Rowe, D.S. mmmology, 14, 745 1968
12.
Harhrg, M. and Samelssm, B. Pmt. natn. Acad. Sci. U.S.A. 2,
42. Churchill,Livingstcme :
3400
1974
13.
Goldstein,I.M., Malmten, C.L., Kaplan, H-B., Jhdahl, H., San~~&sson,B. andWeissmn,G. Clin. l&s..2, No. 3, 518A 1977
14.
Davidson,E.M., Ford-Hutchinson,A.W., smith, M.J.H. and Walker, J.R. Br. J. Pharmac.,63, 407P 1978
15.
Chang, W.C., Murota, S. and Tsurufuji,S. BiochemPhamawl. 2, 109 1978
244
AUGUST 1978 VOL. 16 NO. 2
ME!l'ABOLITESF0RPOLYMXPBCNDCLEMR
E. Ann Kitchen, J. R. Boot, W. Dawson
Lilly F+asearch Centre Limited ErlwoodManor
Abstract
(1)
The cheemtacticactivitiesof thrcmbaxaneB2 (m2)r =2, PGF2c(,the 15-oxo, 15-oxo-13,14-dihydro and 13,14-dihm metabolitesof PSE2, PGF2ar and a mataboliteof TxB2 for polymrphonuclearleumcytes (PMV)havebeen investigated.
(2)
ThrmaxmaneB2 increasedthe directicmalmigrationofrat peritonealPI@Iataamcentrationof 2.Ow/ml audofhman peripheralneutrophilsat a concentrationof 0.5 rJ,s/ml.
(3) Neither PCZ2, PGF2a nor their mtabolites shm.ed chemtactic activity for rat peritonealm. (4) PGF2a and 15-oxc~13,14-dihydro-t chemtactic activity forhmanperipheral
B2 showed no M.
(5) The possible role of thxnboxaneB2ininflamnaticmisdiscussed. Introduction Prostaglantis (PGs) havebeen iqlicatedinmanyoftheevents whichoccurduringanacute inflamnatoryreaction. chleof themany roles attributedto a minority of the prostaglandinsis that of cherlotaxis. Mst of the studies carried out on the chexrotactic prcpwties of theprostaglandins have involvedtheuseofhman periphe.ralbloodPFNandresultshave generally shcm that the par&z PGs of the A B D E and F series are not cheaotacticfor either mononuclearcells or PMV (1,2,3). PGEl,hmever, ischemtactic for rabbit peritmeal M (4,5)but not for rabbit peripheralblood Pm nor for rat PMV derived f_m either peripheralblood or peritoneum (6). HE!l?E(12-Lhydmxy-5,8,10,14 eicosatetraenoicacid) hasbeenshawntobecherrotacticforhua~nblood~s(7).
AUGUST
1978 VOL. 16 NO. 2
239
PROSTAGLANDINS
ThrmboxaneB2has previously been shmn tobe chemYzacti.c for museperitonealm in vitroand tc causethe accum lationof rat PIN in vim after I.D. injection(8). 'Iheaimof thispresentstudywas toextendthese findingsand assessthe chemtacticactivityof TxB2 in vitro for rat peritoneal PkN and forhmanperiphe-calbloodPMJinordertoestablishwhether TxB2ismre generallychemtacticthan,forinstance,FCElwhich appearsto be specificfor rabbitP&N. Inaddition,we have investigated the chemtacticactivities of PG2 and PGF2cr for both rat andhumanbloodPM\Iand15-oxo-13,14-dihydro-TxB2 forhman P&N. Method.5
thrmboxaneB2 and the Thranboxane B2, 15-oxo-13,14-dihydroE-oxo, E-oxo-13,14-dihydx-o and 13,14-dihyd~o mtabolites of m2 and PGF2awerep~paredbiosyntheticallyand characterisedby GLCmass spectrcmatqas describedpreviously(9). PGS2 andPGF2,were obtai.nedfrmCambrianChemicals.Glycogen(exoyster)wasobtained frm Koch Light. Preparation of cell suspensionsA mixedpopulationof cellgcontaining 60-70%PkN,was harvestedfmantheperitinealcavityof rats (Lilly Lodge&&Wistar, male, 150-170g bodyweight) 24 h-afteran intraperitoneal injectionof2 mls 2% glycogenin saline. H~P~wereo$tainedfrantheperipheralbloodofhealthyfiale volunteersand the red blood cellssedimentid with 0.5% methyl cellulosefor 45/60mins. Chemtaxis Assay Chemtaxis was studiedin vitrousing a modification of theE!oydenchanbertechniqueas describedby Wilkinson (10). Polycarbonate filterswith a pore size of 5.0 p (Nuclepore Corporation) wereused. CellswerewashedandsuspendedinGeysbalancedsalt solution4BS.S) , pH 7.3, to give a finalconcentration of 5.0 x 106 cellsml- andO. mls of this cellsuspensionwereplacedin the upperchanber. Agentsundertestwerepreparedin Geys BSS and of TxB2 were used placedin the lmer chanber. Threepreparations rat serm randanlythroughouttheseexperin-mts.Zymsan-activated (11)(ZAS)at a concentration of 5% was used in all testsas the positive chesotactic amtmland Geys BSS as thenegativecontrol.In experiments todistinguishbetween chataxis and&emkinesis (stimulabsdrandm novement)agents~~rtestwereaddedtobothupperandl~~~s. At the end of this Thechanberswereincubatedat3~ for 2h. time,the filtersweresirmltaneouslydetachedandfixed,byimrr?rsion in 95% ethanolfor a minimm periodof 60 min and stainedwith and absolutecountswere carriedout on haematoxylin.Differential cellswhichhadmicratedto thelmer surfaceof'chefilter. Between 4-6 filterswere u&d for each sampleand a minimm of 10 fieldsper filterwerecomtedatamagnificationof 630X.
240
AUGUST 1978 VOL. 16 NO. 2
PROSTAGLANDINS
*suits
The effectof !I%32 cm the migrationof rat peritonealPM+ is sumwisedin table1. TxB2 inueasedthemigrationofrat PMNat 2.0 &ml but failedto show any effectat loweromcentrationsof 1.0 and 0.5 w/ml. These findingswaresimLlarineachofthree expsriments c!axried out. Arandcanmigraticn-~studysha&that this effectwasdue to an jmxease inthedirectionalmigraticm (dmmtaxis) in additiontoa slight increase in c!henokinesis. Neither-2 at 1.0 &ml nor PGF2u at 2.0 and 1.0 w/ml nor the metaboliteof !CxB2at 5.0 and 10.0 pg/ml shmed chemtacticactivity for PIW. Similarlynoeffectwasseenwiththe oxreqxmding matabolites of thesePGswhen testedat2.0 w/ml. Arachidonic acid at50 &mlwas inactivein this system.
The effectof TxB2 on hman PW is shrmnin table 3. TxB2 inmaased the directimalmigrationof cellsatboth0.5 and 2.0 w/ml. andatthelmarconoxtrationalso increasedrandmmigraticn. These findingswzresimilarineachofthreeexperimnts carriedout. Failureto increaserandcmnuvmantatthehighercmcentrationmy have been due to the disruptive effectof TxB2. Cellstich had migratedthroughthe filtertcwardsTxB2 at 2.0 w/ml showedsignsof disintegration andpyknoticnuclei. This effectwas also seen in thosecellsremainin g on theupper surfacebutti amuchlesserextent. NeitherTxB2 at 2.0 &ml nor 15-oxo-13,14-dih@ro-TxB2 at 10 w/ml had any significant effecton the migrationof nmonuclear cells. Prostaglandins E2 and F2(x(at 2.0 w/ml) and the 156x0-13,14dihydronkatabolite of thrceboxane B2(10w/ml) were ineffective in inducingchermtaxis ofhumanperipheralPMV.
Prostaglandins E2, F2a, and their correspondingrmatabolites failed Inaddition, to show cheeotactic activityfor either ratorhmanPMV. themetaboliteofTxB2 exhibitedno&emotacticactivityforhumanpEn. Hcwever,TxB2 shcweda markedchmotacticeffectfor hmmn PMI in vitro and also,althoughto a lesserextent,for rat PMi. The latterfinding oxrelateswithourprevious &se.rvations in z)it)o, that PIW accumlate in the blood vesselsof rat dermisafter localinjectionof thrcuboxane B2, and shouldbelinkedwith the~ork of others (12)thatTxB2 is genera~inlargeanouhtsfrcmth_mboxane A2 duringplatelet aggregation.Inaddition,evidencefromthesestudiesusingmuse (8) ratandhurnan~wouldindicatethat~~~ca~~tyofTxB2is not speciesspecific. TnecherrotacticactivityofTxB2isofpartia*lar interestinviewof a recent report (15)that anti-ihflmnatorydxugs can inhibitits formationin a carrageenin-induced granulcmain rats.
AUGUST 1978 VOL. 16 NO. 2
241
PROSTAGLANDINS
Table
I
Ohemtactic effect of TxB2 for rat peritoneal pclymrphonuckar leucocytes
Lowerchamber
No. of filters
zzs(Z T-2, (0.5 Klrn) 7-2, 7=2r
(1.0 (2.0
w/ml, w/m
7 6 12 12 14
Mean no. of PMN migratingper field 4.3 f 1.6 64.7 -t10.3 6.4 f 0.8 7.0 f 0.9 14.7 + 1.8
P
Table II Cutparisonof theeffect ofTxB2 on the randcxnand directimal migration of rat peritonealpolymxphonuclem leucccytes Mean no. cells/field Directional migraticm migration krpperandlmer charbar) (lowerchanbsr alone)
ChanberCcntents
1.9 f 0.5 19.2 * 4.1 4.8 + 0.5
Geys=S ZAS (5%) T-2 (2.0 w/ml) *
64.6 -I9.1* 17.4 f 1.5*
SignificantdifferenceP
Table III Ohmtactic effect of M2 for hmen peripheral polymrphonuclear leucocytes &an no. cells/field
(zhambx
1
contents
P"
1 (
*
242
Significantdifferencebetween mean values of randan and directionalmigration
AUGUST
1978 VOL. 16 NO. 2
PROSTAGLANDINS
Itappears tinthatTxB2 my cmtribute to the initialinfluxof m into an inflamnatory site,but it is unlikelythat it is the sole factorinvolvedsincetheamcentrationsusedintheseexperimnts were high. Fur&mmre,ccsparisonintheseexpfzimantswith!ZAS indicatesthatTxB2isamchwx&er chemotaxinthauactivated cqxnentsof cosplemant, whichmay be expe&ed to play a major role in cell accmulation. It is interestingthatbothZAS andTxB2 increasedrandmnDvemntinthesestudies. TxB2 is also releasedby phagocytosinghmsnperipheral blcodm (13)in mounts inexcess of tboseproducedbyresting~ and alsobyphagocytcsingrabbit peritonealcells (14). Itispossible thathrB2 maybe fmctioning as an inhibitorofxnigration of PPN alreadyin the tissues,rather thanas aprimaryrrediator of chemtaxis. Acmsistentbutlailevel of releaseduringactivephagocybosis couldmaintaina polymrphcmuclear leucocytepopulatim at the inf1amatoz.y focusprior to the influxof mmmuclear cells. TxA2,PGG;!orPGB2mayhave activitysimilarto,orgreaterthan, TxB2 in thesesystemsbut theirshorthalf-livesin aqueoussolution preventstheirexaminaticm in in vitro chemtsxis experixents.For these ampounds to have a significant effectin uivo,therewould need tobe acontirmus synthesisbythe cellsin the inflamatorysite in or&x to mintsin mncentraticm necessaryfor biologicalactivityof thiskind. Conclusion ~~B2hasbeen~tobe~adicforbothratand humanpolymrphcmuclear leucocytes in vitroand thesedata correlated with similarfindingsin in uivo studiesin the rat. It is suggestedthatthrmboxaneB2, releasedduringplatelet aggregatim,maybe contributing to the accmulationof polynorphonuclear leuaxytesby retainingthesecellsat the inflmimatory site ratherthan by elicitingtheirinitialinflux.
References 1.
Turmr, S.R.,Gaqbell, J.A. and Lynn,W.S. J. Exp. I&d.,3, 1437 1975
2.
Pazzaglia, A., Barker,A., Warin,A.P., Greaves,M.W. Brit. J. Dematol., 96, 533 1977
3.
Mcclatchey, W. ard Snydeman, R. Prostaglandins, 12 No. 3., 415 1976 -
4.
Kaley,G. and Weiner,R. NatureNew Biology,234, 114 1971
5.
Biggs,G.A.,-1, E. and Youlten,L.J.F. Br. J. Phamac. 53, 539 1975
AUGUST 1978VOL. 16 NO. 2
243
6.
Walker, J.R., Smith, M.J.H., Ford-Hutchinson,A.W. J. Pham. Phannac.,28, 745 1976
7.
Gcetzl, E.J., woods, J.M., and Goman, R.R. J. Clin. Invest.,2, 179 1977
8.
Boot, J.R., Dawsm, W. and Kitchen, E.A. J. Physiol.257, 47-48P 1976
9.
Dawson, W., Boot, J.R., Codxrill, A.F., Mallen, D.N.B. Osbcrne, D.J. Nature, 262, 699 1976
10.
Wilkinson,P.C. InCkmtaxis andInflamation,p. Edinburgh,Londcm. 1974
11.
Thaqxon, R.A. and Rowe, D.S. mmmology, 14, 745 1968
12.
Harhrg, M. and Samelssm, B. Pmt. natn. Acad. Sci. U.S.A. 2,
42. Churchill,Livingstcme :
3400
1974
13.
Goldstein,I.M., Malmten, C.L., Kaplan, H-B., Jhdahl, H., San~~&sson,B. andWeissmn,G. Clin. l&s..2, No. 3, 518A 1977
14.
Davidson,E.M., Ford-Hutchinson,A.W., smith, M.J.H. and Walker, J.R. Br. J. Pharmac.,63, 407P 1978
15.
Chang, W.C., Murota, S. and Tsurufuji,S. BiochemPhamawl. 2, 109 1978
244
AUGUST 1978 VOL. 16 NO. 2