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Inhibition of prostaglandin biosynthesis by c−5, c−8, c−11-eicosatrienoic acid
PROSTAGLANDINS INHIBITION OF PROSTAGLANDIN BIOSYNTHESIS BY c-5,c-8,c-II-EICOSATRIENOIC ACID W.C. van Evert, D.H. Nugteren and D,A. van Dorp Unilever Research, Vlaardingen The Netherlands
ABSTRACT The enzyme prostaglandin H + E-isomerase (EC 5.3.99.3), which is present in sheep vesicular gland and needs glutathione as cofactor, is inhibited by ~-5,a-8,a-ll-eicosatrienoic acid, the fatty acid accumulating during essential fatty acid deficiency. The EFA-deficiency syndrome can partly be explained from a prostaglandin deficiency caused by lack of precursors. The present finding indicates that 5,8,11eicosatrienoic acid could well be an additional factor in modifying the symptoms of EFA-deficiency. INTRODUCTION The essential fatty acid (EFA)-deficiency syndrome was first recognized in young rats and described by Burr and Burr (I) in ]929. A great number of nutritional studies on essential fatty acids have since been made (for reviews see Refs. 2-4). During the development of EFAdeficiency, the most striking change in the polyenoic acid pattern of the tissue lipids is a decrease in arachidonic acid and a pronounced increase in the amount of trienoic acid. This was first demonstrated in 1938 by Nunn and Smedley-MacLean (5), who isolated an eicosatrienoic acid as its hexabromide from the livers of fat-deficient rats. This acid was shown by Mead and Slaton (6) to be c-5,c-8,c-ll-eicosatrienoic acid. It was demonstrated that this acid can be made by the animal from oleic acid (7). Thus, when linoleic acid is absent from the diet, biosynthesis of arachidonic acid (an n-6 acid) is not possible, and the body then makes 5,8,11-eicosatrienoic acid (an n-9 acid). The discovery of the biosynthesis of prostaglandins allowed a specific function to be ascribed to arachidonic acid. It was also found that 8,11,14-eicosatrienoic acid, an intermediate in the pathway from linoleic acid towards arachidonic acid, could be converted into prostaglandin E l . However, the fatty acid accumulating in EFA-deficiency, 5,8,1l-eicosatrienoic acid, was not transformed (8). To better understand the role of prostaglandins in the EFA-deficieney syndrome, we investigated whether 5,8,1]-eicosatrienoic acid had any effect on the biosynthesis of prostaglandins from 8, ll,14-eicosatrienoic acid and arachidonic acid. We used for this study sheep vesicular gland microsomes, where two enzymes are present (9,10): a cyclo-oxygenase (EC 1.14.99.1) and an isomerase (EC 5.3.99.3) catalysing the reaction sequence: 202 arachidonic acid
~
prostaglandin H 2 ÷ prostaglandin E 2.
FEBRUARY 1978 VOL. 15 NO. 2
267
PROSTAGLANDINS
Both reactions can be studied separately. To investigate the first transformation, we measured oxygen consumption during incubations with arachidonic acid (II). The second reaction, which needs glutathione as cofactor, was studied with the labile endoperoxide, prostaglandin H2, as substrate (12).
DETERMINATION OF ENZYME ACTIVITY
Alkali reaction 8,11,14-Eicosatrienoic acid (300 ~g) was incubated at 35°C with 4 mg protein from sheep vesicular gland mierosomal particles (13) in 8 ml of a mixture of Glycine-NaOH and K-phosphate buffers (0. I mol/l; pH 8.0), hydroquinone (0.4 mmol/l) and glutathione (0.8 nm~ol/l). Incubations were done in the absence and presence of various amounts of 5,8,11-eicosatrienoic acid ~). After I, 2, 4 and 20 min, 2 ml samples were taken from the incubations and acidified with citric acid solution to pH 3. Prostaglandins were extracted twice with equal volumes of diethyl ether and estimated by the alkali reaction (13). Corrections for endogenous prostaglandins were done with equal series of blank incubations (without 8,11,14-eicosatrienoic acid). Assay of oxygenase activitx Varying amounts of 8,11)14-eicosatrienoic acid were incubated at 30°C with 4 mg sheep vesicular gland microsomal particles in 2 ml glycine-NaOH-K-phosphate buffer mixture (0.I mol/l; pH 8.0), hydroquinone (0.5 mmol/l) and glutathione (I mmol/l) in the reaction vessel of an oxygraph (Gilson Medical Electronics, Middleton, Wisconsin). The oxygen consumption was followed continuously (II). The reaction was started by adding the enzyme. The increase in the rate of oxygen consumption after adding the enzyme was a measure of the enzyme activity. Controls were performed without substrate. Incubation series were done without and with varying amounts of 5,8,11-eicosatrienoic acid. Incubations with radioactive fatt~ acid [l-14C]-8,11,14-Eicosatrienoic acid or arachidonic acid (150 ~g) were incubated with 6 mg microsomal particles in 4.5 ml glycineNaOH-K-phosphate buffer mixture (0.! mol/l) (pH 8.0) with 600 ~g glutathione and I00 ~g hydroquinone for 20 min at 30°C. Analysis of the reaction products has been described previously (12)
~)The preparation consisted of about 98% free fatty acid's (TLC), composed of about 96% of the proper acid, 3% dienoic acid and traces of monoenic and ynedienic acids (GLC). The amount of conjugated dienoic acid was less than 0.1% as measured by UV spectrophotometry.
268
FEBRUARY 1978 VOL. 15 NO. 2
PROSTAGLANDINS
Incubations with prosta~landin H 6 ~g [l-14C]-prostaglandin H l or H 2 (12) were incubated with 1.5 mg particulate fraction in 2 ml glycine-NaOH-K-phosphate buffer mixture (0.5 mol/l; pH 8.0), with GSH (0.5 mmol/l) for 25 min at 30°C with and without 200 ~g 5,8,ll-eicosatrienoic acid. After acidification to pH < 3, the lipid mixture was separated by TLC. The various hands were visualized with phosphomolybdic acid at l|0°C, and scraped off. Radioactivity was determined by liquid scintillation counting (cf. 12). For comparison an incubation of prostaglandin H 2 with 200 ~g of a bicyclic hydroxy acid (14) was done under the same conditions.
RESULTS
Considerable inhibition of prostaglandin E biosynthesis in the precence of a two-to-threefold excess of 5,8,11-eicosatrienoic acid was found by the alkali reaction (Fig. I). Some inhibition was also found when the oxygen consumption was assayed polarographically in an oxygraph. However, the kinetics of this inhibition were not straightforward, which led us to perform incubation with radioactive substrates. This allows quantification, by thin-layer chromatography, of every product of the reaction rather than prostaglandin E only (12). Such incubations of sheep vesicular gland microsomes with radioactive fatty acids, in the presence of hydroquinone and glutathione, gave nearly 100% conversion of the substrate, and hardly any formation of prostaglandins D and Fa, (Fig. 2) which is in accordance with our earlier work (13). When sufficient 5,8,11-eicosatrienoic acid was added, the substrate was still transformed completely, indicating that the cyclooxygenase was not strongly inhibited. However, the yield of prostaglandin E decreased, and that of the prostaglandins D and F~, and of 12-hydroxy-ClT-acid, which are chemical degradation products of the unstable prostaglandin H, increased. The product composition in the presence of 5,8,11-eicosatrienoic acid was to a certain extent comparable with that obtained in the absence of glutathione, which is an essential cofactor for the isomerase. These results rendered it likely that the isomerase was inhibited. To demonstrate the inhibition of the isomerase by 5,8,1|-eicosatrienoic acid more directly, we incubated the prostaglandin H itself. Fig. 3 shows that also in these experiments addition of 5,8,ll-eicosatrienoic acid or of another isomerase inhibitor, a monohydroxy acid with a bicyclo [2.2,|]-heptene group (14), yielded increased amounts of prostaglandin D and F~, which again can be explained by chemical degradation of prostaglandin H. Inhibition of the isomerase in the presence of glutathione leads to relatively large quantities of prostaglandin Fa.
DISCUSSION
Most of the prostaglandin synthetase inhibitors are non-steroidal, anti-inflannnatory drugs, antioxidants or substrate analogues (II,15), which inhibit the first enzyme in the sequence, viz. the cyclo-oxygenase.
FEBRUARY 1978 VOL. 15 NO. 2
269
PROSTAGLANDINS
F~E 1 (pg)
2001 •/¢-
--O -&
lOOitl/II.--,--- 0
/ 2O I
t/rain
Figure I Biosynthesis of prostaglandin E l from 300 ~g 8,11,14-eicosatrienoic acid with 4 mg protein from sheep vesicular gland microsomal particles in the presence of 0 (e), 300 (A), 600 (D) or 900 (o) hg c-5,c-8,c-11eicosatrienoic acid. Prostaglandin E; was determined by the alkali reaction. The enzyme responsible for the prostaglandin E formation, the prostaglandin H + prostaglandin E isomerase, needs glutathione for full activity, and is specifically inhibited by p-chloromercuribenzoate, N-ethylmaleimide and other thio-reagents. Apart from these reagents, only one specific isomerase inhibitor has been described thus far, a bicyclo [2.2.1]heptene derivative (14), which is a kind of prostaglandin H analogue. The present study shows that 5,8,1]-eicosatrienoic acid is also a potent isomerase inhibitor. Our finding is especially of interest for a better understanding of the essential fatty acid deficiency symptoms and for explaining the effects of variable amounts of linoleic acid in the diet. It is known, for instance, that kidney medulla is relatively rich in prostaglandin E2-forming enzymes, and there is evidence of the involvement of kidney prostaglandins in the regulation of blood pressure. During essential fatty acid deficiency there is not only shortage of substrate for prostaglandin biosynthesis but there is probably also a change in the prostaglandin E2/D2/F2a ratio because of the accumulation of 5,8,11eicosatrienoic acid.
270
FEBRUARY 1978 VOL. 15 NO. 2
PROSTAGLANDINS
20:4
20:3
radioactivityl% 100
80
HA j
!
60
'
~ -H.H~ ..'.~'.!!! /-- F~3~'--/ ......,,:~2.-:.~. / - P G D
40
-/
! PGE
:
!
!I
I
-
-
2O ............... ~
C
/--FA
~.
C
*I -GSH
÷I -GSH
Figure 2 Incubation of radioactive 8,;l,]4-eicosatrienoic acid (20:3) or arachidonic acid (20:4): C = complete system; +I = 300 ~g 5,8,1]eicosatrienoic acid added; -GSH = glutathione omitted; FA = nonconverted fatty acid substrate; HA = 12-hydroxy C20-acid + 12-hydroxyC]7-acid; R = sum of % radioactivity on other zones of the plate.
radioactivityl%
PGH1
PGH 2
100 R
i :
/-
HH-/ ;-----]!~
IHII,,(II
6O 4O
~
20
-E
C .I
F~3E
-GSH
C
,,I
-OSH*B
-E
Figure 3 Inhibition of the enzymic isomerisation of 6 ~g l]-]4C[-prostaglandin H I (PGH I) or H 9 (PGHg) , in the presence of 300 pg GSH, by 200 ~g 5~8,11-~icosat~ienoi~ acid (I) or 200 ~g bicyclie hydroxy acid (14) (B). C = complete system; -GSH = glutathione omitted; -E = enzyme omitted HH = 12-hydroxy-C17-acid. Further abbreviations: see Fig. 2.
FEBRUARY
1 9 7 8 V O L . 15 N O . 2
271
PROSTAGLAND1NS
It has long been recognised that the 8,11,14-eicosatrienoic acid/ arachidonic acid ratio in the phospholipids determines the prostaglandins El/E2, Fla/F2e ratio, etc. Our results indicate that the 5,8, ll-eicosatrienoic acid/arachidonic acid ratio could be one of the factors determining the E2/D2/F2~ ratio. Other natural fatty acids possibly have the same effect as 5,8,1l-eicosatrienoic acid (16,17). An increase in 5,8,11-eicosatrienoic acid in the phospholipids of the body can occur through diets which, although low in linoleic acid, give no visible symptoms of essential fatty acid deficiency (3,4). These relatively small changes in fatty acid composition may influence the very subtle regulatory mechanisms in which prostaglandins participate.
REFERENCES I. Burr, G.O. and Burr, M.M. (1929) J. Biol. Chem. 82, 345. 2. Aaes-J6rgensen, E.
(196l) Physiol. Rev. 41, l-Sl.
3. Holman, R.T. (1968) Prog. Chem. Fats Other Lipids 9, (2) 279-348. 4. Holman, R.T. (1976) in Lipids, vol. l (Paoletti, R., Porcellati, G. and Jacini, G., eds.) Raven Press, New York pp 215-226. 5. Nunn, L.C.A. and Smedley-MacLean,
I (1938) Bioehem. J. 32, 2178.
6. Mead, J.F. and Slaton, W.H. (1956) J. Biol. Chem. 219, 705-709. 7. Fulco, A.J. and Mead, J.F. (1959) J. Biol. Chem. 234, 1411-1416. 8. Struijk, C.B., Beerthuis, R.K., Pabon, H.J.J. and Van Dorp, D.A. (1966) Recl. Tray. Chim. Pays-Bas 85, 1233-1250. 9. Miyamoto, T., Ogino, N., Yamamoto, S. and Hayaishi, O. (]976) J. Biol. Chem. 251, 2629-2636. 10. Van der Ouderaa, F.J. Buytenhek, M., Nugteren, D.H. and Van Dorp, D.A. (1977) Biochim. Biophys. Acta 487, 315-331. II. Nugteren~ D.H. (1970) Biochim. Biophys. Acta 210, 171-176. 12. Nugteren, D.H. and Hazelhof, E. (1973) Biochim. Biophys. Acta 326, 448-461. 13. Nugteren, D.H., Beerthuis, R.K. and Van Dorp, D.A. (1966) Recl. Trav. Chim. Pays-Bas 85, 405-419. 14. Wlodawer, P., Samuelsson, B., Albonico, S.M. and Corey, E.J. (1971) J. Am. Chem. Soc. 93, 2815-2816. 15. Robinson, H.J. and Vane, J.R., Prostaglandin Synthetase Inhibitors (1974) Raven Press, New York. 16. Flower, R.J., Cheung, H.S. and Cushman, D.W. (1973) Prostaglandins 4, 325-341. 17. Ziboh, V.A., Vanderhoek, J.Y. and Lands, W.E.M. (1974) Prostaglandins 5, 233-240.
272
FEBRUARY 1978 VOL. 15 NO. 2
ABSTRACT The enzyme prostaglandin H + E-isomerase (EC 5.3.99.3), which is present in sheep vesicular gland and needs glutathione as cofactor, is inhibited by ~-5,a-8,a-ll-eicosatrienoic acid, the fatty acid accumulating during essential fatty acid deficiency. The EFA-deficiency syndrome can partly be explained from a prostaglandin deficiency caused by lack of precursors. The present finding indicates that 5,8,11eicosatrienoic acid could well be an additional factor in modifying the symptoms of EFA-deficiency. INTRODUCTION The essential fatty acid (EFA)-deficiency syndrome was first recognized in young rats and described by Burr and Burr (I) in ]929. A great number of nutritional studies on essential fatty acids have since been made (for reviews see Refs. 2-4). During the development of EFAdeficiency, the most striking change in the polyenoic acid pattern of the tissue lipids is a decrease in arachidonic acid and a pronounced increase in the amount of trienoic acid. This was first demonstrated in 1938 by Nunn and Smedley-MacLean (5), who isolated an eicosatrienoic acid as its hexabromide from the livers of fat-deficient rats. This acid was shown by Mead and Slaton (6) to be c-5,c-8,c-ll-eicosatrienoic acid. It was demonstrated that this acid can be made by the animal from oleic acid (7). Thus, when linoleic acid is absent from the diet, biosynthesis of arachidonic acid (an n-6 acid) is not possible, and the body then makes 5,8,11-eicosatrienoic acid (an n-9 acid). The discovery of the biosynthesis of prostaglandins allowed a specific function to be ascribed to arachidonic acid. It was also found that 8,11,14-eicosatrienoic acid, an intermediate in the pathway from linoleic acid towards arachidonic acid, could be converted into prostaglandin E l . However, the fatty acid accumulating in EFA-deficiency, 5,8,1l-eicosatrienoic acid, was not transformed (8). To better understand the role of prostaglandins in the EFA-deficieney syndrome, we investigated whether 5,8,1]-eicosatrienoic acid had any effect on the biosynthesis of prostaglandins from 8, ll,14-eicosatrienoic acid and arachidonic acid. We used for this study sheep vesicular gland microsomes, where two enzymes are present (9,10): a cyclo-oxygenase (EC 1.14.99.1) and an isomerase (EC 5.3.99.3) catalysing the reaction sequence: 202 arachidonic acid
~
prostaglandin H 2 ÷ prostaglandin E 2.
FEBRUARY 1978 VOL. 15 NO. 2
267
PROSTAGLANDINS
Both reactions can be studied separately. To investigate the first transformation, we measured oxygen consumption during incubations with arachidonic acid (II). The second reaction, which needs glutathione as cofactor, was studied with the labile endoperoxide, prostaglandin H2, as substrate (12).
DETERMINATION OF ENZYME ACTIVITY
Alkali reaction 8,11,14-Eicosatrienoic acid (300 ~g) was incubated at 35°C with 4 mg protein from sheep vesicular gland mierosomal particles (13) in 8 ml of a mixture of Glycine-NaOH and K-phosphate buffers (0. I mol/l; pH 8.0), hydroquinone (0.4 mmol/l) and glutathione (0.8 nm~ol/l). Incubations were done in the absence and presence of various amounts of 5,8,11-eicosatrienoic acid ~). After I, 2, 4 and 20 min, 2 ml samples were taken from the incubations and acidified with citric acid solution to pH 3. Prostaglandins were extracted twice with equal volumes of diethyl ether and estimated by the alkali reaction (13). Corrections for endogenous prostaglandins were done with equal series of blank incubations (without 8,11,14-eicosatrienoic acid). Assay of oxygenase activitx Varying amounts of 8,11)14-eicosatrienoic acid were incubated at 30°C with 4 mg sheep vesicular gland microsomal particles in 2 ml glycine-NaOH-K-phosphate buffer mixture (0.I mol/l; pH 8.0), hydroquinone (0.5 mmol/l) and glutathione (I mmol/l) in the reaction vessel of an oxygraph (Gilson Medical Electronics, Middleton, Wisconsin). The oxygen consumption was followed continuously (II). The reaction was started by adding the enzyme. The increase in the rate of oxygen consumption after adding the enzyme was a measure of the enzyme activity. Controls were performed without substrate. Incubation series were done without and with varying amounts of 5,8,11-eicosatrienoic acid. Incubations with radioactive fatt~ acid [l-14C]-8,11,14-Eicosatrienoic acid or arachidonic acid (150 ~g) were incubated with 6 mg microsomal particles in 4.5 ml glycineNaOH-K-phosphate buffer mixture (0.! mol/l) (pH 8.0) with 600 ~g glutathione and I00 ~g hydroquinone for 20 min at 30°C. Analysis of the reaction products has been described previously (12)
~)The preparation consisted of about 98% free fatty acid's (TLC), composed of about 96% of the proper acid, 3% dienoic acid and traces of monoenic and ynedienic acids (GLC). The amount of conjugated dienoic acid was less than 0.1% as measured by UV spectrophotometry.
268
FEBRUARY 1978 VOL. 15 NO. 2
PROSTAGLANDINS
Incubations with prosta~landin H 6 ~g [l-14C]-prostaglandin H l or H 2 (12) were incubated with 1.5 mg particulate fraction in 2 ml glycine-NaOH-K-phosphate buffer mixture (0.5 mol/l; pH 8.0), with GSH (0.5 mmol/l) for 25 min at 30°C with and without 200 ~g 5,8,ll-eicosatrienoic acid. After acidification to pH < 3, the lipid mixture was separated by TLC. The various hands were visualized with phosphomolybdic acid at l|0°C, and scraped off. Radioactivity was determined by liquid scintillation counting (cf. 12). For comparison an incubation of prostaglandin H 2 with 200 ~g of a bicyclic hydroxy acid (14) was done under the same conditions.
RESULTS
Considerable inhibition of prostaglandin E biosynthesis in the precence of a two-to-threefold excess of 5,8,11-eicosatrienoic acid was found by the alkali reaction (Fig. I). Some inhibition was also found when the oxygen consumption was assayed polarographically in an oxygraph. However, the kinetics of this inhibition were not straightforward, which led us to perform incubation with radioactive substrates. This allows quantification, by thin-layer chromatography, of every product of the reaction rather than prostaglandin E only (12). Such incubations of sheep vesicular gland microsomes with radioactive fatty acids, in the presence of hydroquinone and glutathione, gave nearly 100% conversion of the substrate, and hardly any formation of prostaglandins D and Fa, (Fig. 2) which is in accordance with our earlier work (13). When sufficient 5,8,11-eicosatrienoic acid was added, the substrate was still transformed completely, indicating that the cyclooxygenase was not strongly inhibited. However, the yield of prostaglandin E decreased, and that of the prostaglandins D and F~, and of 12-hydroxy-ClT-acid, which are chemical degradation products of the unstable prostaglandin H, increased. The product composition in the presence of 5,8,11-eicosatrienoic acid was to a certain extent comparable with that obtained in the absence of glutathione, which is an essential cofactor for the isomerase. These results rendered it likely that the isomerase was inhibited. To demonstrate the inhibition of the isomerase by 5,8,1|-eicosatrienoic acid more directly, we incubated the prostaglandin H itself. Fig. 3 shows that also in these experiments addition of 5,8,ll-eicosatrienoic acid or of another isomerase inhibitor, a monohydroxy acid with a bicyclo [2.2,|]-heptene group (14), yielded increased amounts of prostaglandin D and F~, which again can be explained by chemical degradation of prostaglandin H. Inhibition of the isomerase in the presence of glutathione leads to relatively large quantities of prostaglandin Fa.
DISCUSSION
Most of the prostaglandin synthetase inhibitors are non-steroidal, anti-inflannnatory drugs, antioxidants or substrate analogues (II,15), which inhibit the first enzyme in the sequence, viz. the cyclo-oxygenase.
FEBRUARY 1978 VOL. 15 NO. 2
269
PROSTAGLANDINS
F~E 1 (pg)
2001 •/¢-
--O -&
lOOitl/II.--,--- 0
/ 2O I
t/rain
Figure I Biosynthesis of prostaglandin E l from 300 ~g 8,11,14-eicosatrienoic acid with 4 mg protein from sheep vesicular gland microsomal particles in the presence of 0 (e), 300 (A), 600 (D) or 900 (o) hg c-5,c-8,c-11eicosatrienoic acid. Prostaglandin E; was determined by the alkali reaction. The enzyme responsible for the prostaglandin E formation, the prostaglandin H + prostaglandin E isomerase, needs glutathione for full activity, and is specifically inhibited by p-chloromercuribenzoate, N-ethylmaleimide and other thio-reagents. Apart from these reagents, only one specific isomerase inhibitor has been described thus far, a bicyclo [2.2.1]heptene derivative (14), which is a kind of prostaglandin H analogue. The present study shows that 5,8,1]-eicosatrienoic acid is also a potent isomerase inhibitor. Our finding is especially of interest for a better understanding of the essential fatty acid deficiency symptoms and for explaining the effects of variable amounts of linoleic acid in the diet. It is known, for instance, that kidney medulla is relatively rich in prostaglandin E2-forming enzymes, and there is evidence of the involvement of kidney prostaglandins in the regulation of blood pressure. During essential fatty acid deficiency there is not only shortage of substrate for prostaglandin biosynthesis but there is probably also a change in the prostaglandin E2/D2/F2a ratio because of the accumulation of 5,8,11eicosatrienoic acid.
270
FEBRUARY 1978 VOL. 15 NO. 2
PROSTAGLANDINS
20:4
20:3
radioactivityl% 100
80
HA j
!
60
'
~ -H.H~ ..'.~'.!!! /-- F~3~'--/ ......,,:~2.-:.~. / - P G D
40
-/
! PGE
:
!
!I
I
-
-
2O ............... ~
C
/--FA
~.
C
*I -GSH
÷I -GSH
Figure 2 Incubation of radioactive 8,;l,]4-eicosatrienoic acid (20:3) or arachidonic acid (20:4): C = complete system; +I = 300 ~g 5,8,1]eicosatrienoic acid added; -GSH = glutathione omitted; FA = nonconverted fatty acid substrate; HA = 12-hydroxy C20-acid + 12-hydroxyC]7-acid; R = sum of % radioactivity on other zones of the plate.
radioactivityl%
PGH1
PGH 2
100 R
i :
/-
HH-/ ;-----]!~
IHII,,(II
6O 4O
~
20
-E
C .I
F~3E
-GSH
C
,,I
-OSH*B
-E
Figure 3 Inhibition of the enzymic isomerisation of 6 ~g l]-]4C[-prostaglandin H I (PGH I) or H 9 (PGHg) , in the presence of 300 pg GSH, by 200 ~g 5~8,11-~icosat~ienoi~ acid (I) or 200 ~g bicyclie hydroxy acid (14) (B). C = complete system; -GSH = glutathione omitted; -E = enzyme omitted HH = 12-hydroxy-C17-acid. Further abbreviations: see Fig. 2.
FEBRUARY
1 9 7 8 V O L . 15 N O . 2
271
PROSTAGLAND1NS
It has long been recognised that the 8,11,14-eicosatrienoic acid/ arachidonic acid ratio in the phospholipids determines the prostaglandins El/E2, Fla/F2e ratio, etc. Our results indicate that the 5,8, ll-eicosatrienoic acid/arachidonic acid ratio could be one of the factors determining the E2/D2/F2~ ratio. Other natural fatty acids possibly have the same effect as 5,8,1l-eicosatrienoic acid (16,17). An increase in 5,8,11-eicosatrienoic acid in the phospholipids of the body can occur through diets which, although low in linoleic acid, give no visible symptoms of essential fatty acid deficiency (3,4). These relatively small changes in fatty acid composition may influence the very subtle regulatory mechanisms in which prostaglandins participate.
REFERENCES I. Burr, G.O. and Burr, M.M. (1929) J. Biol. Chem. 82, 345. 2. Aaes-J6rgensen, E.
(196l) Physiol. Rev. 41, l-Sl.
3. Holman, R.T. (1968) Prog. Chem. Fats Other Lipids 9, (2) 279-348. 4. Holman, R.T. (1976) in Lipids, vol. l (Paoletti, R., Porcellati, G. and Jacini, G., eds.) Raven Press, New York pp 215-226. 5. Nunn, L.C.A. and Smedley-MacLean,
I (1938) Bioehem. J. 32, 2178.
6. Mead, J.F. and Slaton, W.H. (1956) J. Biol. Chem. 219, 705-709. 7. Fulco, A.J. and Mead, J.F. (1959) J. Biol. Chem. 234, 1411-1416. 8. Struijk, C.B., Beerthuis, R.K., Pabon, H.J.J. and Van Dorp, D.A. (1966) Recl. Tray. Chim. Pays-Bas 85, 1233-1250. 9. Miyamoto, T., Ogino, N., Yamamoto, S. and Hayaishi, O. (]976) J. Biol. Chem. 251, 2629-2636. 10. Van der Ouderaa, F.J. Buytenhek, M., Nugteren, D.H. and Van Dorp, D.A. (1977) Biochim. Biophys. Acta 487, 315-331. II. Nugteren~ D.H. (1970) Biochim. Biophys. Acta 210, 171-176. 12. Nugteren, D.H. and Hazelhof, E. (1973) Biochim. Biophys. Acta 326, 448-461. 13. Nugteren, D.H., Beerthuis, R.K. and Van Dorp, D.A. (1966) Recl. Trav. Chim. Pays-Bas 85, 405-419. 14. Wlodawer, P., Samuelsson, B., Albonico, S.M. and Corey, E.J. (1971) J. Am. Chem. Soc. 93, 2815-2816. 15. Robinson, H.J. and Vane, J.R., Prostaglandin Synthetase Inhibitors (1974) Raven Press, New York. 16. Flower, R.J., Cheung, H.S. and Cushman, D.W. (1973) Prostaglandins 4, 325-341. 17. Ziboh, V.A., Vanderhoek, J.Y. and Lands, W.E.M. (1974) Prostaglandins 5, 233-240.
272
FEBRUARY 1978 VOL. 15 NO. 2