- Email: [email protected]
Human monocytes synthesize eicosanoids from T lymphocyte-derived arachidonic acid
PROSTAGLANDINS
H[RAN MDN(L-TrES SYNTHESIZE EIOOSANOIDS FROM T LYMPHDCi"rE-DERIVED ARACHIDONIC ACID Marc E. Goldyne and John D. Stobo Department of Dermatology and the Howard Hughes Medical Institute, Department of Medicine, University of Calif., San Francisco. School of Medicine, San Francisco, Calif., 94143, U.S.A. ABSTRACT T lymphocytes prelabeled with [14C] arachidonic acid failed to synthesize any eicosanoids even following stimulation with phytohemagglutinin, but they did release free [14C] arachidonic acid. Co-culture of unlabeled monocytes with the prela~led T lymphocytes resulted in the sysnthesis of [I~C] thrcmboxane B 2, a major monocytederived eicosanoid. These data show that monocytes can utilize T lymphocyte-derived arachidonic acid for the synthesis of eicosanoids. INTRD[KETIGN Products of arachidonic acid ~A) metaboli~n modulate various functions of i n m ~ t e n t cells (i). Among immunocumpetent cells, it has become apparent that the monocyte-macrophages ~4~s), but not T lymphocytes ~fcells) or B ly~phocytes (B-cells), synthesize thrcmboxane B 2 ~), various prostaglandins ~Gs), as well as other elcosanoldS (2-6). However, human T-cell phospholipids contain virtually the sane percentage of AA as do M~s (7). In addition, a lectin such as phytohemagglutinin (PHA) can enchance the release of AA from T-cells (8). These findings prcmpted our investigation of whether or not AA released by T-cells could serve as a substrate for the synthesis of eicosanoids by resident M~s.
NOVEMBER 1982 VOL. 24 NO. 5
623
PROSTAGLANDINS
MATERIALS
AND
~I~0DS
Cell Isolation Human peripheral blood mononuclear cells ~BMC) were obtained and depleted of platelets (< 1 platelet/10 cells) as previously described (9). T-cells were isolated after first removing M~s by adherence to plastic and subsequently subjecting the nonadherent cells to standard sheep red blood cell rosetting techniques (9). Adherence of the T-cell enriched population for a second time for 30 minutes resulted in a nonadherent population consisting 0f.= 98% T-cells. MQs of ~ 98% purity (determined by nonspecific esterase staining and phagocytosis of latex beads) were obtained as previously described (9). Radioimmunoassa~zs Prostaglandins E 2 (P(~2) and TXB 2 generated from endogenous AA by mixtures of M@s and T-cells were measured using established radioimmunoassays. The anti-P~ antiserum showed the following cross-reactivities: PGE 2 100%, P(~I - 15%, P(~mleC 18%, 6 - k e t o - ~ l ~ - 3%, P(~92, PC4~ a nd TMB 2 - 1%. The anti-TXB 2 antibody (Immunalysis Inc., Milton, M~) showed the following cross-reactivities: TXB 2 - 100%; PC~ 2 - 1%; P(~92- 2.5%; P(~2~-1%: 6 - k e t o - l ~ and PGA -1%; [3H] PG~? (sp. act. 160 Ci/mmol) and [ 3 ~ T~B 2 (sp. act. 150 CiTmmol) were obtained from Amersham, Arlington Hts, Ill. and from New England Nuclear, Boston, MA respectively. The techniques for the radioimmunoassays have been published (i0, ii). Radiolabeled Lymphocytes T-cel~s were prelabeled w i t h ~ l ~ ] ~ by incubating 10-20 X i0° T-cells with 1.5 X 10~ cpm [i~C] AA (sp. act. 60 mCi/mmol, New England, Boston, ~ ) for 30 minutes at 37° C in 5 ml of phosphate buffered saline ~PBS; ~H 7.4; containing 7 mM ED~I~, 15 mM Tris and 1% human AB serum). The cells were then washed twice with fresh PBS to remove unincorporated label. Aliquots of 5 X i 0 ° labeled T-cells were incubated for 40 hours with or without 5 ug/ml PHA: ~urroughs Wellccme Co.) and with or without 1 X i0o
624
NOVEMBER 1982 VOL. 24 NO. 5
PROSTAGLANDINS
unlabeled M@s in 2 ml of RPMI 1640 containing 5% heat inactivated fetal calf serum CFCS). After incubation, the supernatants were centrifuged for 10 minutes at 1800 rpm to remove cells and a lipid extraction of the supernatants was carried out as previously described (3). Thin-Layer radiochrcmato~raphy The lipid extracts were subjected to thin-layer chromatography using a solvent system consisting of ethyl acetate/acetic acid (90:1) and glass plates coated with silica gel G. Radioactive peaks were identified using a Berthold LB2760 TLC scanner. Co-chrcmatographed standards for. P(~2 , TXB2 , 6-keto-P(~l~ , P(~2 and AA (Upjohn Dzagnostlcs, Kalamazoo, MI) were visualized using a 10% phosphQmolybdic acid/ethanol spray followed by heating. RESULTS The data in Table i show that T-ceils, by themselves, failed to generate TXB9 or P(~2 even in the presence of PHA. M@s, on the other h~md, clearly generated both metabolites. Combining M~s with T-cells in the absence of PHA resulted in the generation of T~B 2 and PGE~ at concentrations equivalent to those generated by Mqs alone. However, the addition of PHA to mixtures of M~s and T-cells approximately doubled the amount of TXB 2 and P(~2 generated. For TXB 2 , this level was greater than that generated by the mixture of M~s and PHA alone (p< 0.5; paired t test), or by M~s and T cells alone (p < 0.i0; paired t test). Since PHA can stimulate the release of free AA from T-cells, we attempted to determine if the increased T~3~ synthesis noted when M~s and T-cells were exposed to P H A ~ could in part, reflect the metabolism by M~s of [14C] AA released from the prelabeled T-cells.
NOVEMBER 1982 VOL. 24 NO. 5
625
PROSTAGLANDINS
Table i: TXB 2 and PG~ 2 generation by PBMC in the presence of PHA. Cells were incubated for 24 hours in RPMI 1640 (5% FCS) at 37%. S u~ernatants from the incubation were then subjected to radzoimmunoassays for TXB 2 and PG~.
TABLE 1
Additions
Exp #I
Exp #2
Medial
1.9
1.4
0.01
0.4
T-cells2
2.0
N.D.
0.01
N.D.
T-cells + PHA3
1.9
N.D.
0.01
0.2
~4
Exp #i
Exp #2
15.9
18.6
3.5
1.5
+ PHA
21.9
24.0
6.7
8.0
+ T-cells
19.8
25.2
3.9
2.1
+ T-cells + PHA
45.6
42.0
10.4
5.6
1640 + 5% FCS; 25 X 106 cells; 3pHA = 5 ug/ml; 4 1 X 106 cells. Figure 1 shows the radiochrcmatogram obtained by incubating[14C] AA-labeled T-cells for 40 hours either Ln the absence or presence of M~s and in the absence or presence of 5 ug/ml PHA. T-cells, with or without PHA, released free [14C] AA but failed to metabolize it. The addition of unlabeled M~s however resulted in the appearance of a peak co-migrating with standard TXB 2 . Addition of 5 ug/ml of PHA increased the radioactivity in this peak from 52% to 63% of the total radioactivity recovered.
626
NOVEMBER 1982 VOL. 24 NO. 5
PROSTAGLANDINS
Figure i: Thin-layer radiochrcmatography of lipid extracts taken from 40 hour incubation of[14C 3 AA-labeled T cells in the presence or absence of unlabeled M~s and PHA. The solid circles represent the location of co-chrcmatographed standards. The addition of unlabeled M~s to labeled Tcells (third chrcmatogram from top) resulted in a peak co-chrcmatographing with TXB 2 and representing 51% of the total radioactivity recovered. The addition of PHA to these cells increased the radioactivity in the TXB 2 peak to 63% of the total recovered (bottom chrcmatogram).
5 x 10 6
14C-AA labeled T-ceils
O0 6 keto PGFI~,PGF2~
NOVEMBER 1982 VOL. 24 NO. 5
O0 PGE2,TXB2
• PGO2
• AA
627
PROSTAGLANDINS
DISCUSSION Our studies indicate that when M@s are added to Tcells in the presence of PHA, synthesis of TXB 2 is greater than the total amount of TXB 2 generated when ~ s or T-cells are individually activated. To explain these results, it could be argued that only activated T-cells can synthesize eicosanoids and that in the absence of M~s or soluble ~ products, the T-cell enriched populations could not be sufficiently activated. This is unlikely because activation of the T-enriched cells, as determined by PHA-induced incorporation of tritiated thymidine, was equivalent to that found among mixtures of T-cells and ~ s (data not shown). Therefore we assume that the 1-2% M~ contamination of the T-enriched populaton provided accessory cell function sufficient for activation. Furthermore, stimulation of AA metabolism among T-cells by soluble products from M~s is unlikely. Addition of supernatants taken from 24 hour incubations of purified M~s, which were exposed for 3 hours prior to incubation to 10 -6 M indcmethacin to block endogenous PG synthesis, failed to induce PHA-stimulated T-cells to generate ~ r e a c t i v e PGE 2 [fXB2 was not assayed in this experiment). The explanation most ccmpatable with our results is that the activation of T-cells by PHA releases a soluble factor or factors that enhance the generation TXB 2 by M~s. Our data demonstrate that one soluble factor in free AA. Since Tcells do not synthesize TXB 2 , the [14C] TXB 2 could only derive from M~ metabolism of the T-cell-derived [14C] AA. Precedent for ~ r a t i o n between heterogeneous cell groups in the generation of eicosanoids has been provided by Marcus et al. who demonstrated that the PG endoperoxide (P(~ 2 ) released by activated platelets could serve as a substrate for prostacyclin synthesis by vascular endothelial cells (12). OUr studies are the first to describe a similar interaction among i m m u ~ t e n t cells. Since TXB 2 may enhance T-cell proliferation (13), the ability of T-cells to influence TXB 2 generation by M~s may exert a positive feedback on their own proliferatlon. While the radioimmunoassay data also suggest an interaction between T-cells and M~s in the generation of
628
NOVEMBER 1982 VOL. 24 NO. 5
PROSTAGLANDINS
P(~2 , the radiochrQmatograms fail to show a peak oochrcmatographing with standard PGE 2 . This discrepancy may be a technical one in that the quantity of TXB 2 generated by the M#s is approximately 4 to 8 fold greater than the quantity of P(~2 , and in cur chrcmlatograms one would not expect to .see a peak for PGE 2 that could be differentiated from therebackground activity. In summary, this study demonstrates for the first time the synthesis of eicosanoids by M~s utilizing AA released by T-cells. The immunoregulatory consequences of such an interaction remain to be defined. ACKNO~ ~ m X ~ P S This work was supported in part by NIH Grant AM30803. The authors wish to thank Mr. Gunther Benthin for his technical assistance.
I.
Goldyne, M.E., and J.D. Stobo. Immunoregulatory role of prostaglandins and related lipids, CRC Crit. Rev. Immunol. 2:189-233, 1981.
2.
Kurland, J.l., and R. Bockman. Prostaglandin E production by human blood monocytes and mouse peritoneal macrophages, J. Exp. Med. 147:952-955, 1978.
3.
Kennedy, M.S., J.D. Stobo, and M.E. Goldyne, In vitro synthesis of prostaglandins and related lipids by populations of human peripheral blood mononuclear cells, Prostaglandins 20:135-145, 1980.
4.
Dy, M., M. Astoin, M. Rigand, and J. Hamburger. Prostaglandin (PG) release in the mixed lymphocyte culture; effect of presensitization by a skin allograft, nature of the PG-producing Cell, Eur. J. Immunol. I0:121-126, 1980.
NOVEMBER 1982 VOL. 24 NO. 5
629
PROSTAGLANDINS
5.
Bray, M.A., R.G. Powell, and P.M. Lydyard. Prostaglandin generation by separated human blood mononuclear cell fractions, Int. J. Immunopharm. 3:377381, 1981.
6.
Bankhurst, A.D., E. Hastain, J.S. Goodwin, and G.T. Peake. The nature of the prostaglandin-producing mononuclear cell in human peripheral blood, J. Lab. Clin. Med. 97:179-186, 1981.
7.
Stossel, T.P., R.J. Mason, and A.L. Smith. Lipid peroxidation by human blood phagocytes, J. Clin. Invest. 54: 638-645, 1974.
8.
Parker, C.W., J.P. Kelly, S.F. Falkenhein, and M.G. Huber. Release of arachidonic acid frcm human lymphocytes in response to mitogenic lectins, J. Exp. Med. 149:1487-1503, 1979.
9.
Picker, L.J., H.V. Raff, M.E. Goldyne, and J.D. Stobo. Metabolic heterogeneity among human monocytes and its modulation by P(~2, J" Immunol. 124:25572562, 1980.
i0.
Goldyne, M.E., R.K. Winkelmann, and R.J. Ryan. Prostaglandin activity in human cutaneous inflanlnation: detection by radioimmunoassay Prostaglandins 4:737749, 1973.
ii.
Van Orden, D., and D.B. Farley. Prostaglandin F radioimmunoassay utilizing polyethylene glycol separation technique, Prostaglandins 4:215-233, 1973.
12.
Marcus, A.J., B.B. Weksler, E.A. Jaffe, and M.J. Broekman. Synthesis of prostacyclin from plateletderived endoperoxides by cultured human endothelial cells. J. Clin. Invest. 66:979-986, 1980.
13.
Kelly, J.P., M.C. Johnson, and C.W. Parker. Effects of inhibitors of arachidonic acid metabolism on mitogenesis in human lymphocytes: possible role of thrc~boxanes and products of the lipoxygenase pathway. J. Immunol. 122:1563-1571, 1979.
Editor: Peter W. Ramwell Received: 9-28-82 Accepted: 9-30-82
630
NOVEMBER 1982 VOL. 24 NO. 5
H[RAN MDN(L-TrES SYNTHESIZE EIOOSANOIDS FROM T LYMPHDCi"rE-DERIVED ARACHIDONIC ACID Marc E. Goldyne and John D. Stobo Department of Dermatology and the Howard Hughes Medical Institute, Department of Medicine, University of Calif., San Francisco. School of Medicine, San Francisco, Calif., 94143, U.S.A. ABSTRACT T lymphocytes prelabeled with [14C] arachidonic acid failed to synthesize any eicosanoids even following stimulation with phytohemagglutinin, but they did release free [14C] arachidonic acid. Co-culture of unlabeled monocytes with the prela~led T lymphocytes resulted in the sysnthesis of [I~C] thrcmboxane B 2, a major monocytederived eicosanoid. These data show that monocytes can utilize T lymphocyte-derived arachidonic acid for the synthesis of eicosanoids. INTRD[KETIGN Products of arachidonic acid ~A) metaboli~n modulate various functions of i n m ~ t e n t cells (i). Among immunocumpetent cells, it has become apparent that the monocyte-macrophages ~4~s), but not T lymphocytes ~fcells) or B ly~phocytes (B-cells), synthesize thrcmboxane B 2 ~), various prostaglandins ~Gs), as well as other elcosanoldS (2-6). However, human T-cell phospholipids contain virtually the sane percentage of AA as do M~s (7). In addition, a lectin such as phytohemagglutinin (PHA) can enchance the release of AA from T-cells (8). These findings prcmpted our investigation of whether or not AA released by T-cells could serve as a substrate for the synthesis of eicosanoids by resident M~s.
NOVEMBER 1982 VOL. 24 NO. 5
623
PROSTAGLANDINS
MATERIALS
AND
~I~0DS
Cell Isolation Human peripheral blood mononuclear cells ~BMC) were obtained and depleted of platelets (< 1 platelet/10 cells) as previously described (9). T-cells were isolated after first removing M~s by adherence to plastic and subsequently subjecting the nonadherent cells to standard sheep red blood cell rosetting techniques (9). Adherence of the T-cell enriched population for a second time for 30 minutes resulted in a nonadherent population consisting 0f.= 98% T-cells. MQs of ~ 98% purity (determined by nonspecific esterase staining and phagocytosis of latex beads) were obtained as previously described (9). Radioimmunoassa~zs Prostaglandins E 2 (P(~2) and TXB 2 generated from endogenous AA by mixtures of M@s and T-cells were measured using established radioimmunoassays. The anti-P~ antiserum showed the following cross-reactivities: PGE 2 100%, P(~I - 15%, P(~mleC 18%, 6 - k e t o - ~ l ~ - 3%, P(~92, PC4~ a nd TMB 2 - 1%. The anti-TXB 2 antibody (Immunalysis Inc., Milton, M~) showed the following cross-reactivities: TXB 2 - 100%; PC~ 2 - 1%; P(~92- 2.5%; P(~2~-1%: 6 - k e t o - l ~ and PGA -1%; [3H] PG~? (sp. act. 160 Ci/mmol) and [ 3 ~ T~B 2 (sp. act. 150 CiTmmol) were obtained from Amersham, Arlington Hts, Ill. and from New England Nuclear, Boston, MA respectively. The techniques for the radioimmunoassays have been published (i0, ii). Radiolabeled Lymphocytes T-cel~s were prelabeled w i t h ~ l ~ ] ~ by incubating 10-20 X i0° T-cells with 1.5 X 10~ cpm [i~C] AA (sp. act. 60 mCi/mmol, New England, Boston, ~ ) for 30 minutes at 37° C in 5 ml of phosphate buffered saline ~PBS; ~H 7.4; containing 7 mM ED~I~, 15 mM Tris and 1% human AB serum). The cells were then washed twice with fresh PBS to remove unincorporated label. Aliquots of 5 X i 0 ° labeled T-cells were incubated for 40 hours with or without 5 ug/ml PHA: ~urroughs Wellccme Co.) and with or without 1 X i0o
624
NOVEMBER 1982 VOL. 24 NO. 5
PROSTAGLANDINS
unlabeled M@s in 2 ml of RPMI 1640 containing 5% heat inactivated fetal calf serum CFCS). After incubation, the supernatants were centrifuged for 10 minutes at 1800 rpm to remove cells and a lipid extraction of the supernatants was carried out as previously described (3). Thin-Layer radiochrcmato~raphy The lipid extracts were subjected to thin-layer chromatography using a solvent system consisting of ethyl acetate/acetic acid (90:1) and glass plates coated with silica gel G. Radioactive peaks were identified using a Berthold LB2760 TLC scanner. Co-chrcmatographed standards for. P(~2 , TXB2 , 6-keto-P(~l~ , P(~2 and AA (Upjohn Dzagnostlcs, Kalamazoo, MI) were visualized using a 10% phosphQmolybdic acid/ethanol spray followed by heating. RESULTS The data in Table i show that T-ceils, by themselves, failed to generate TXB9 or P(~2 even in the presence of PHA. M@s, on the other h~md, clearly generated both metabolites. Combining M~s with T-cells in the absence of PHA resulted in the generation of T~B 2 and PGE~ at concentrations equivalent to those generated by Mqs alone. However, the addition of PHA to mixtures of M~s and T-cells approximately doubled the amount of TXB 2 and P(~2 generated. For TXB 2 , this level was greater than that generated by the mixture of M~s and PHA alone (p< 0.5; paired t test), or by M~s and T cells alone (p < 0.i0; paired t test). Since PHA can stimulate the release of free AA from T-cells, we attempted to determine if the increased T~3~ synthesis noted when M~s and T-cells were exposed to P H A ~ could in part, reflect the metabolism by M~s of [14C] AA released from the prelabeled T-cells.
NOVEMBER 1982 VOL. 24 NO. 5
625
PROSTAGLANDINS
Table i: TXB 2 and PG~ 2 generation by PBMC in the presence of PHA. Cells were incubated for 24 hours in RPMI 1640 (5% FCS) at 37%. S u~ernatants from the incubation were then subjected to radzoimmunoassays for TXB 2 and PG~.
TABLE 1
Additions
Exp #I
Exp #2
Medial
1.9
1.4
0.01
0.4
T-cells2
2.0
N.D.
0.01
N.D.
T-cells + PHA3
1.9
N.D.
0.01
0.2
~4
Exp #i
Exp #2
15.9
18.6
3.5
1.5
+ PHA
21.9
24.0
6.7
8.0
+ T-cells
19.8
25.2
3.9
2.1
+ T-cells + PHA
45.6
42.0
10.4
5.6
1640 + 5% FCS; 25 X 106 cells; 3pHA = 5 ug/ml; 4 1 X 106 cells. Figure 1 shows the radiochrcmatogram obtained by incubating[14C] AA-labeled T-cells for 40 hours either Ln the absence or presence of M~s and in the absence or presence of 5 ug/ml PHA. T-cells, with or without PHA, released free [14C] AA but failed to metabolize it. The addition of unlabeled M~s however resulted in the appearance of a peak co-migrating with standard TXB 2 . Addition of 5 ug/ml of PHA increased the radioactivity in this peak from 52% to 63% of the total radioactivity recovered.
626
NOVEMBER 1982 VOL. 24 NO. 5
PROSTAGLANDINS
Figure i: Thin-layer radiochrcmatography of lipid extracts taken from 40 hour incubation of[14C 3 AA-labeled T cells in the presence or absence of unlabeled M~s and PHA. The solid circles represent the location of co-chrcmatographed standards. The addition of unlabeled M~s to labeled Tcells (third chrcmatogram from top) resulted in a peak co-chrcmatographing with TXB 2 and representing 51% of the total radioactivity recovered. The addition of PHA to these cells increased the radioactivity in the TXB 2 peak to 63% of the total recovered (bottom chrcmatogram).
5 x 10 6
14C-AA labeled T-ceils
O0 6 keto PGFI~,PGF2~
NOVEMBER 1982 VOL. 24 NO. 5
O0 PGE2,TXB2
• PGO2
• AA
627
PROSTAGLANDINS
DISCUSSION Our studies indicate that when M@s are added to Tcells in the presence of PHA, synthesis of TXB 2 is greater than the total amount of TXB 2 generated when ~ s or T-cells are individually activated. To explain these results, it could be argued that only activated T-cells can synthesize eicosanoids and that in the absence of M~s or soluble ~ products, the T-cell enriched populations could not be sufficiently activated. This is unlikely because activation of the T-enriched cells, as determined by PHA-induced incorporation of tritiated thymidine, was equivalent to that found among mixtures of T-cells and ~ s (data not shown). Therefore we assume that the 1-2% M~ contamination of the T-enriched populaton provided accessory cell function sufficient for activation. Furthermore, stimulation of AA metabolism among T-cells by soluble products from M~s is unlikely. Addition of supernatants taken from 24 hour incubations of purified M~s, which were exposed for 3 hours prior to incubation to 10 -6 M indcmethacin to block endogenous PG synthesis, failed to induce PHA-stimulated T-cells to generate ~ r e a c t i v e PGE 2 [fXB2 was not assayed in this experiment). The explanation most ccmpatable with our results is that the activation of T-cells by PHA releases a soluble factor or factors that enhance the generation TXB 2 by M~s. Our data demonstrate that one soluble factor in free AA. Since Tcells do not synthesize TXB 2 , the [14C] TXB 2 could only derive from M~ metabolism of the T-cell-derived [14C] AA. Precedent for ~ r a t i o n between heterogeneous cell groups in the generation of eicosanoids has been provided by Marcus et al. who demonstrated that the PG endoperoxide (P(~ 2 ) released by activated platelets could serve as a substrate for prostacyclin synthesis by vascular endothelial cells (12). OUr studies are the first to describe a similar interaction among i m m u ~ t e n t cells. Since TXB 2 may enhance T-cell proliferation (13), the ability of T-cells to influence TXB 2 generation by M~s may exert a positive feedback on their own proliferatlon. While the radioimmunoassay data also suggest an interaction between T-cells and M~s in the generation of
628
NOVEMBER 1982 VOL. 24 NO. 5
PROSTAGLANDINS
P(~2 , the radiochrQmatograms fail to show a peak oochrcmatographing with standard PGE 2 . This discrepancy may be a technical one in that the quantity of TXB 2 generated by the M#s is approximately 4 to 8 fold greater than the quantity of P(~2 , and in cur chrcmlatograms one would not expect to .see a peak for PGE 2 that could be differentiated from therebackground activity. In summary, this study demonstrates for the first time the synthesis of eicosanoids by M~s utilizing AA released by T-cells. The immunoregulatory consequences of such an interaction remain to be defined. ACKNO~ ~ m X ~ P S This work was supported in part by NIH Grant AM30803. The authors wish to thank Mr. Gunther Benthin for his technical assistance.
I.
Goldyne, M.E., and J.D. Stobo. Immunoregulatory role of prostaglandins and related lipids, CRC Crit. Rev. Immunol. 2:189-233, 1981.
2.
Kurland, J.l., and R. Bockman. Prostaglandin E production by human blood monocytes and mouse peritoneal macrophages, J. Exp. Med. 147:952-955, 1978.
3.
Kennedy, M.S., J.D. Stobo, and M.E. Goldyne, In vitro synthesis of prostaglandins and related lipids by populations of human peripheral blood mononuclear cells, Prostaglandins 20:135-145, 1980.
4.
Dy, M., M. Astoin, M. Rigand, and J. Hamburger. Prostaglandin (PG) release in the mixed lymphocyte culture; effect of presensitization by a skin allograft, nature of the PG-producing Cell, Eur. J. Immunol. I0:121-126, 1980.
NOVEMBER 1982 VOL. 24 NO. 5
629
PROSTAGLANDINS
5.
Bray, M.A., R.G. Powell, and P.M. Lydyard. Prostaglandin generation by separated human blood mononuclear cell fractions, Int. J. Immunopharm. 3:377381, 1981.
6.
Bankhurst, A.D., E. Hastain, J.S. Goodwin, and G.T. Peake. The nature of the prostaglandin-producing mononuclear cell in human peripheral blood, J. Lab. Clin. Med. 97:179-186, 1981.
7.
Stossel, T.P., R.J. Mason, and A.L. Smith. Lipid peroxidation by human blood phagocytes, J. Clin. Invest. 54: 638-645, 1974.
8.
Parker, C.W., J.P. Kelly, S.F. Falkenhein, and M.G. Huber. Release of arachidonic acid frcm human lymphocytes in response to mitogenic lectins, J. Exp. Med. 149:1487-1503, 1979.
9.
Picker, L.J., H.V. Raff, M.E. Goldyne, and J.D. Stobo. Metabolic heterogeneity among human monocytes and its modulation by P(~2, J" Immunol. 124:25572562, 1980.
i0.
Goldyne, M.E., R.K. Winkelmann, and R.J. Ryan. Prostaglandin activity in human cutaneous inflanlnation: detection by radioimmunoassay Prostaglandins 4:737749, 1973.
ii.
Van Orden, D., and D.B. Farley. Prostaglandin F radioimmunoassay utilizing polyethylene glycol separation technique, Prostaglandins 4:215-233, 1973.
12.
Marcus, A.J., B.B. Weksler, E.A. Jaffe, and M.J. Broekman. Synthesis of prostacyclin from plateletderived endoperoxides by cultured human endothelial cells. J. Clin. Invest. 66:979-986, 1980.
13.
Kelly, J.P., M.C. Johnson, and C.W. Parker. Effects of inhibitors of arachidonic acid metabolism on mitogenesis in human lymphocytes: possible role of thrc~boxanes and products of the lipoxygenase pathway. J. Immunol. 122:1563-1571, 1979.
Editor: Peter W. Ramwell Received: 9-28-82 Accepted: 9-30-82
630
NOVEMBER 1982 VOL. 24 NO. 5