Effect of fatty acid modification on prostaglandin production by cultured 3T3 cells

We have investigated the extent to which modifications in the essential fatty acid content of mammalian cells can affect prostaglandin production. Swiss mouse 3T3 cells stimulated with the calcium ionophore A23187 produced 1.7 to 7 times more prostaglandin E(2) (PGE(2)) when the cultures were supplemented with linoleic acid. Increases in PGE(2) production as a result of linoleic acid supplementation occurred under all culture conditions except during the first 24 hr after attachment, when prostaglandin production was very high. Arachidonic acid supplementation produced a similar enhancement in the capacity of the cells to produce PGE(2), but no appreciable increase occurred when the cultures were supplemented with oleic acid. The phospholipids of the cells exposed to the linoleate-enriched medium contained 4 times more arachidonic acid and twice as much linoleic acid as compared with the corresponding controls. The choline phosphoglycerides were most highly enriched in arachidonic acid, but 2- to 3-fold increases also occurred in the inositol and ethanolamine phosphoglycerides. When cultures initially enriched with linoleic acid were transferred to an unsupplemented medium, the fatty acid composition as well as the capacity of the cells to produce PGE(2) reverted almost to control values. The amount of exogenous arachidonic acid converted to PGE(2) as measured by radioimmunoassay also was greater when the cells were enriched with linoleic acid. Studies with radioactive arachidonic acid indicated that the distribution of prostaglandin metabolites was not affected appreciably by linoleic acid enrichment. These findings suggest that at least two factors contribute to the increased capacity of the cultures supplemented with linoleate to produce PGE(2). One is enrichment of the phospholipid substrate pools with arachidonic acid. The other is an increased ability of the cells to synthesize PGE(2) from unesterified arachidonic acid, perhaps because the prostaglandin-forming enzymes are more active.-Denning, G. M., P. H. Figard, and A. A. Spector. Effect of fatty acid modification on prostaglandin production by cultured 3T3 cells.     

Enrichment of human platelet phospholipids with linoleic acid diminishes thromboxane release

We have investigated whether exposure of human platelets to elevated concentrations of linoleic acid, the principal dietary polyunsaturate, would influence platelet thromboxane A₂ release. Platelets were incubated with albumin-bound linoleic acid at 30°C for 24 h, with prostaglandin E₁ added to prevent aggregation. The linoleic acid supplemented platelets released, on averaged, 50% less thromboxane A₂ in response to stimulation with thrombin than corresponding control platelets. Other fatty acids were without appreciable effect. The inhibition of thrombin-stimulated thromboxane A₂ release was dependent on the time and temperature of incubation, as well as on the concentration of added linoleic acid. Supplementation increased the amount of linoleic acid in the platelet phospholipids, but the arachidonic acid content of the phospholipids was reduced. [1-¹⁴C]Linoleic acid was not converted to arachidonic acid by the platelets. Linoleic acid was released exclusively form the inositol phosphoglycerides when the enriched platelets were stimulated with thrombin. The linoleate-enriched platelets converted less [1-¹⁴C]arachidonic acid to all prostaglandin products, suggesting that the platelet cyclooxygenase was partially inhibited.     

Influence of dietary polyunsaturated fatty acids on renal & aortic prostaglandin synthesis in 1 kidney 1 clip goldblatt hypertensive rats

To study the influence of dietary modification on prostaglandin synthesis and on blood pressure regulation, the effects of dietary enrichment with linolenic or linoleic acid was compared with standard rat chow in 3 groups of 13 rats before and after renal artery constriction and contralateral nephrectomy. Before renal artery constriction 4 weeks supplementation with 40 en% linseed oil (53% linolenic acid) increased renal linolenic acid, decreased arachidonic acid, and suppressed synthesis of 6-keto-PGF_1α and PGE₂ by renal homogenates (33% and 38% respectively, p<0.01) compared with standard diet. Rats fed on 40 en % sunflower seed oil (63% linoleic acid) increased renal prostaglandin synthesis (p<0.05) compared with linseed oil, but not compared with standard diet. Seven weeks after renal artery constriction renal and aortic 6-keto-PGF_1α and PGE₂ were suppressed 30% to 50% (p<0.05) by linseed oil supplements compared with sunflower seed oil and standard diets. In the sunflower seed oil group aortic 6-keto-PGF_1α correlated (r = 0.75, p<0.02) with final systolic blood pressure. Final systolic blood pressures were similar in linseed oil (152.9 mmHg ± se 3.3, sunflower oil (155.1 ± se 6.6) and standard diet group (159.0 ±se 4.2). Thus dietary linseed oil suppressed renal and aortic prostaglandin synthesis but did not accentuate renal hypertension, and linoleic acid supplementation did not protect against 1 kidney 1 clip renal hypertension.     

Arachidonic acid metabolism by adult human osteoblast-like cells exhibits sexually dimorphic characteristics

The eicosanoids, including prostaglandin E₂ (PGE₂) and other bioactive arachidonic acid metabolites, are important local mediators of bone remodeling. Presumably, the limited or excessive synthesis of the eicosanoids could compromise bone homeostasis. We have noted that the stimulated release of arachidonic acid by adult male donor derived human osteoblast-like (hOB) cells exceeded the stimulated release measured for female-derived hOB cells by 1.5-fold. Assays of PGE₂ biosynthesis by cytokine-stimulated hOB cells also demonstrated a sex-linked difference, such that male hOB cell PGE₂ production exceeded female cell production by 1.6–2.2-fold. The calcium-dependent cytoplasmic phospholipase A₂ activity in subcellular fractions prepared from hOB cell homogenates was higher in both the cytosolic (1.6-fold) and particulate (1.5-fold) fractions from the male cells than in those prepared from female hOB cells, suggesting a molecular basis for the observed sexually dimorphic characteristics related to arachidonic acid metabolism by hOB cells. The relatively limited capacity of the female cells may limit needed intracellular and intercellular signaling during bone remodeling, thereby contributing to the development of bone pathology. J. Cell. Biochem. 71:74–81, 1998. © 1998 Wiley-Liss, Inc.     

Differences in types of prostaglandins produced by two MDCK canine kidney cell sublines

The pattern of prostaglandins produced from arachidonic acid by two sublines of MDCK canine kidney epithelia cells was different. In one subline designated MDCK₁, the most prevalent prostaglandin product was PGE₂, whereas the most prevalent product in the subline designated MDCK₂ was PGF_2α. This difference was observed when cells previously labeled with [1^?14C]arachidonic acid were stimulated with either bradykinin or the calcium ionophore A23187, or when prostaglandins were produced from labeled arachidonic acid added directly to the assay medium. In the latter case, the difference was maintained over a 38-fold range of extracellular arachidoante concentrations. These findings indicate the there is a persistent difference in the distribution of prostaglandins produced by the two commonly used sublines of MDCK cells.     

The antihypertensive action of arachidonic acid in the spontaneous hypertensive rat and its antagonism by anti-inflammatory agents

Changes in arterial blood pressure and heart rate were observed in the spontaneous hypertensive (SH) rat following the intravenous administration of arachidonic acid, the precursor of prostaglandin E₂ (PGE₂). The pronounced fall in blood pressure and the increase in heart rate induced by arachidonic acid were also observed in SH rats receiving either prostaglandin E₁ (PGE₁) or PGE₂. In SH rats receiving various anti-inflammatory agents the cardiovascular responses to arachidonic acid were inhibited, but the blood pressure responses to the E-type prostaglandins were not altered. The data are interpreted to suggest that cardiovascular changes induced by arachidonic acid are mediated via its conversion to PGE₂.     

Prostaglandin E2 is a Potential Mediator of Extracellular ATP Action in Osteoblast-Like Cells

Extracellular ATP dose dependently stimulated ⁴⁵Ca²⁺ influx even in the presence of nifedipine, a Ca²⁺ antagonist that inhibits voltage-dependent Ca²⁺ channel, in osteoblast-like MC3T3-E1 cells. ATP stimulated arachidonic acid release and the synthesis of prostaglandin E₂ (PGE₂). However, the ATP-induced arachidonic acid release was significantly reduced by chelating extracellular Ca²⁺ with EGTA. On the other hand, ATP induced DNA synthesis of these cells in a dose-dependent manner in the range between 1μM and 1 mM. The pretreatment with indomethacin, a cyclooxygenase inhibitor, suppressed both ATP-induced PGE₂ synthesis and DNA synthesis in these cells. The inhibitory effect by 50μM indomethacin on the DNA synthesis was reversed by adding 10μM PGE₂. These results strongly suggest that extracellular ATP stimulates Ca²⁺ influx resulting in the release of arachidonic acid in osteoblast-like cells and that extracellular ATP-induced proliferative effect is mediated, at least in part, by ATP-stimulated PGE₂ synthesis.     

PGE2 secretion from organ cultured gastric mucosa: Correlation with cyclooxygenase activity and endogenous substrate release

In gastrointestinal research the in vitro release of prostaglandins from incubated or cultured biopsies is a widely used method to estimate prostaglandin synthesis. We therefore investigated the rate limiting mechanisms of PGE₂ release in organ cultured gastric mucosa of the rabbit, determining PGE₂ secretion from organ cultured mucosal biopsies by radioimmunoassay and prostaglandin synthesizing capacity by in vitro incubation of mucosal homogenate or microsomes with [¹⁴C]-arachidonic acid.Freshly taken biopsies secreted PGE₂ at an initial high rate, that decreased during the following 4 hrs of culture. This PGE₂ release was dose dependently reduced by inhibitors of the prostaglandin cyclooxygenase. 5mM acetylsalicylic acid (ASA) maximally suppressed PGE₂ secretion to 7% of controls, and the inhibition by ASA was quantitatively similar at every given culture period. PGE₂ release was markedly increased by carbenoxolone but was only slightly activated by extracellular calcium and the Ca⁺⁺-ionophore A23187. However, Ca⁺⁺/A23187 were unable to maintain PGE₂ secretion at the initial rate.PGE₂ secretion was undisturbed in calcium-free medium but was reduced to 50–60% of controls by excess EDTA. The intracellular calcium chelator 1,2-bis-(2-aminophenoxy)-ethane-N,N,N′,N′,-tetraacetic acid-acetoxymethyl ester (BAPTA-AM) similarly inhibited PGE₂ release to 72% of controls. In contrast, PGE₂ release was unaffected by the intracellular calcium antagonist 3,4,5-trimethylene-bis(4-formylpyridinium bromide) dioxime (TMB-8), the calmodulin antagonists N-(6-aminohexyl)-1-5-chloro-1-naphthalenesulfonamide (W-7) and calmidazolium (compound R24571) or various direct inhibitors of endogenous arachidonic acid release like tetracaine, bromophenacyl bromid, neomycine or low dose quinacrine, indicating that the reduction of PGE₂ release by EDTA or BAPTA may be mediated by mechanisms different from substrate release. In contrast, an inhibition of PGE₂ secretion by quinacrine at high concentrations (≥ 0.8mM) was attributed to a direct inhibition of the prostaglandin cyclooxygenase, similar to ASA. Finally, the reduction of the prostaglandin synthesizing capacity by ASA was strongly correlated with the inhibition of PGE₂ secretion, also at low concentrations and minor degrees of inhibition.From these data we conclude, that the activity of the prostaglandin cyclooxygenase is rate limiting for PGE₂ secretion from organ cultured mucosal biopsies rather than arachidonic acid release by a phospholipase A₂. This should be considered for interpretation of studies based on prostaglandin release from cultured mucosa.     

Urinary excretion of and recent N-6-3 fatty acid intake

Epidemiological studies were performed in a Japanese fishing village when catches of fish were highest and in a Japanese farming village with usual fish consumption. Intake of eicosapentaenoic, docosahexaenoic and also arachidonic acid were significantly higher in the fishing village during the 3 days of the study than in the farming village. The correlation between eicosapentaenoic acid intake on the day when urine was collected and excreion of Δ 17-2,3-dinor-6-keto-prostaglandin F_1α, the main urinary metabolite of prostaglandin I₃, was highly significant, whereas there was no correlation between arachidonic or linoleic acid intake and excretion of 2,3-dinor-6-keto-prostaglandin F_1α, the main urinary metabolite of prostaglandin I₂. We suggest that the arachidonic acid pool for prostaglandin I₂ production is not quickly influenced by dietary linoleic or arachidonic acid because of a large pool size of arachidonic acid and a slow conversion of linoleic acid to arachidonic acid, while prostaglandin I₃ formation is directly related to the intake of eicosapentaenoic acid.     

Bradykinin-stimulated release of [3H]arachidonic acid from phospholipids of HSDM1C1 cells: Comparison of diacyl phospholipids and plasmalogens as sources of prostaglandin precursors

Ethanolamine plasmalogens (1-alk-1′-enyl-2-acyl-sn-glycero-3-phosphoethanolamines) of many tissues contain high levels of arachidonate at their 2-position, and in certain tissues have been implicated as possible donors of arachidonate required in the synthesis of prostaglandins and thromboxanes. In the present study, [³H]arachidonate-labeled phospholipids of HSDM₁C₁ cells, a cell line derived from a mouse fibrosarcoma, were examined to determine the donor of the arachidonic acid released upon bradykinin stimulation of the synthesis of PGE₂. HSDM₁C₁ cells labeled with [³H]arachidonic acid for 24 hr in serum-free medium were used in most of the experiments and had the following distribution of label among the cellular lipids; phosphatidylcholine (33%), phosphatidylinositol (20%), diacyl-sn-glycero-3-phosphoethanolamine (15%), ethanolamine plasmalogen (15%), and less polar lipids (16%). Bradykinin treatment stimulated a rapid hydrolysis of [³H]arachidonate from the cellular lipids and conversion of the released acid to PGE₂, which was secreted into the medium. The label was released predominantly from phosphatidylinositol and possibly from phosphatidylcholine with no detectable change in the labeling of diacyl- or 1-alk-1′-enyl-2-acyl-sn-glycero-3-phosphoethanolamine. The ethanolamine plasmalogens, therefore, do not appear to be involved in the stimulated release of arachidonate in the HSDM₁C₁ cells. Indomethacin blocked the bradykinin-stimulated synthesis of PGE₂ and to a lesser degree inhibited the release of [³H]-arachidonate from the cellular lipids into the medium.     

Prostaglandin synthesis in human placenta and fetal membranes

The conversion of exogenous arachidonic acid into prostaglandins was studied in human placenta and fetal membrane microsomes. Only one prostaglandin was formed, prostaglandin E₂ (PGE₂), in fetal membrane microsomes. In placental microsomes PGE₂ was further transformed into 15 keto-PGE₂. Cofactor requirements and some characteristics of the system were studied. 1 to 3% conversion of arachidonic acid into prostaglandins was observed in placental microsomes and 5 to 8% conversion in fetal membrane microsomes.     

Prostaglandin production by methylcholanthrene-transformed mouse BALB/3T3: Inhibition by cytochalasin B

Cytochalasin B inhibits the production of prostaglandins by serum-, thrombin-, and bradykinin-stimulated MC5-5 cells. The serum-stimulated release of arachidonic acid from cellular phospholipids also is inhibited. Cytochalasin B does not affect the cells' prostaglandin synthetase activity when exogenous arachidonic acid is present. Deacylation of phospholipids may be the step affected by cytochalasin B possibly as a result of disruption of microfilament organization. Colchicine and vinblastine, two drugs that can disrupt microtubule organization, do not inhibit prostaglandin production by cells.     

Modification of upper gastrointestinal prostaglandin synthesis by dietary fatty acids

The effects of dietary iols on gastric, duodenal mucosa and liver were investigated ina rat model. Unsaturated fatty acid profles and in vitro prostaglandin (PG) synthesis (PGE₂, PGF_2α, 6-oxo-PGF_1α and thromboxane B₂). were measured after 14 days of dietary oil supplements.There were no significant differences in prostanoid synthesis between rats fed coconut oil (high saturated fat content) and standard diet. After fish oil supplement, tissue eicosapentaenoic acid and docosahexaenoic acid levels were higher, arachidonic acid levels were lower, and prostanoid synthesis was reduced in both stomach and duodenum. After corn oil and evening primrose oil, linoleic acid levels were variaby increased, bt there were no significant differences in arachidonic acid or prostanoid synthesis. Dihomogamma-linolenic acid levels were slightly increased after evening primrose oil.Dietary incorporation of fatty acids into gastroduodenal tissue is not uniform. When incorporated, fatty acids can modify prostaglandin synthesis.     

Effects of all CIS-5, 8,11,14,17 eicosapentaenoic acid and PGH3 on platelet aggregation

In human platelet-rich plasma (PRP) eicosapentaenoic acid (EPA) inhibited platelet aggregation induced by a stable analogue of PGH₂ (U46619), arachidonic acid, collagen or ADP. EPA was more potent than oleic, linoleic, α-linolenic or γ-linolenic acids. In aspirin-treated platelets, aggregation induced by U46619 was inhibited to a similar extent by arachidonic acid or by EPA over a range of concentrations of 0.05–0.3 mM. EPA incubated with PRP did not induce the generation of a thromboxane (TXA)-like activity; indeed it prevented the formation of TXA₂ induced by arachidonic acid or by collagen. The anti-aggregatory activity of EPA was not influenced by inhibitors of cyclo-oxygenase and lipoxygenase. The anti-aggregatory action of EPA may be caused by a rapid occupancy by EPA of TXA₂/PGH₂ “receptors” on platelet membrane as well as by a slower displacement of arachidonic acid from platelet phospholipids by chemically unchanged molecules of EPA.Not all samples of PRP were irreversibly aggregated by PGH₂, but in those that were, PGH₃ also induced an immediate dose-dependent but reversible aggregation. After a 4 min incubation of non-aggregating doses of PGH₂ or PGH₃ (100–300 nM) with PRP a stable anti-aggregatory compound was detected. The inhibitory activity produced from PGH₃ was apparently more potent (ca 10 times) than that obtained from PGH₂. The anti-aggregating compounds were identified by TLC and GLC-MS as PGD₂ and PGD₃. The apparent difference of potency between PGD₂ and PGD₃ was attributed to the concurrent production of PGE₂ and PGE₃. PGE₂ prevented the inhibitory effect of PGD₂ whereas PGE₃ did not affect the activity of PGD₃.It is concluded that one of the reasons for the low incidence of myocardial infarction in Eskimos could be that the pro-aggregatory arachidonic acid is replaced in their phospholipids by the anti-aggregatory EPA.     

Bradykinin-stimulated release of [H]arachidonic acid from phospholipids of HSDM1C1 cells: Comparison of diacyl phospholipids and plasmalogens as sources of prostaglandin precursors

Ethanolamine plasmalogens (1-alk-1′-enyl-2-acyl-sn-glycero-3-phosphoethanolamines) of many tissues contain high levels of arachidonate at their 2-position, and in certain tissues have been implicated as possible donors of arachidonate required in the synthesis of prostaglandins and thromboxanes. In the present study, [³H]arachidonate-labeled phospholipids of HSDM₁C₁ cells, a cell line derived from a mouse fibrosarcoma, were examined to determine the donor of the arachidonic acid released upon bradykinin stimulation of the synthesis of PGE₂. HSDM₁C₁ cells labeled with [³H]arachidonic acid for 24 hr in serum-free medium were used in most of the experiments and had the following distribution of label among the cellular lipids; phosphatidylcholine (33%), phosphatidylinositol (20%), diacyl-sn-glycero-3-phosphoethanolamine (15%), ethanolamine plasmalogen (15%), and less polar lipids (16%). Bradykinin treatment stimulated a rapid hydrolysis of [³H]arachidonate from the cellular lipids and conversion of the released acid to PGE₂, which was secreted into the medium. The label was released predominantly from phosphatidylinositol and possibly from phosphatidylcholine with no detectable change in the labeling of diacyl- or 1-alk-1′-enyl-2-acyl-sn-glycero-3-phosphoethanolamine. The ethanolamine plasmalogens, therefore, do not appear to be involved in the stimulated release of arachidonate in the HSDM₁C₁ cells. Indomethacin blocked the bradykinin-stimulated synthesis of PGE₂ and to a lesser degree inhibited the release of [³H]-arachidonate from the cellular lipids into the medium.     

Cyclic AMP and platelet prostaglandin synthesis

The present study has investigated the influence of agents which elevate intracellular levels of endogenous platelet adenosine 3′5′-cyclic monophosphate (cyclic AMP), and the effect of the exogenous cyclic AMP analog, dibutyryl cyclic AMP, on the conversion of ¹⁴C-arachidonic acid by washed platelets. Prostaglandin E₁ (PGE₁), PGE₁ with theophylline, or dibutyryl cyclic AMP incubated with washed platelets prevented arachidonic acid induced platelet aggregation, but had no effect on the conversion of arachidonic acid to 12L-hydroxy-5,8,10, 14-eicosatetraenoic acid (HETE), 12L-hydroxy-5,8,10 heptadecatrienoic acid (HHT), or thromboxane B₂. Ultrastructural studies of the platelet response revealed that agents acting directly or indirectly to increase the level of cyclic AMP inhibited the action of arachidonic acid on washed platelets and prevented internal platelet contraction as well as aggregation. The influence of PGE₁ with theophylline, and dibutyryl cyclic AMP on the thrombin induced release of ¹⁴C-arachidonic acid from platelet membrane phospholipids was also investigated. These agents were found to be potent inhibitors of the thrombin stimulated release of arachidonic acid from platelet phospholipids, due most likely to an inhibition of platelet phospholipase A activity. The results show that dibutyryl cyclic AMP and agents which elevate intracellular cyclic AMP levels act to inhibit platelet activation at two steps 1) internal contraction and 2) release of arachidonic acid from platelet phospholipids.     

The Effect of Various Lipids on Flowering of Pharbitis nil in in vitro Culture

The effect of applied arachidonic acid, prostaglandin (PGE₁) and various sterols and combinations of arachidonic acid + sterols, on flowering of Pharbitis nil were ascertained by using a tissue culture technique. It was found that arachidonic acid, PGE₁ stigmasterol, testosterone, cholesterol, stigmasterol + arachidonic acid, -sitosterol + arachidonic acid and cholesterol + arachidonic acid all caused earlier flowering. Four inhibitors of prostaglandin biosynthesis (gentisic acid, acetylsalicylic acid, salicylic acid and oleic acid), inhibited flowering completely. The results confirm that the compounds tested could possibly play a role in the flowering of P. nil.     

The effect of physiological levels of non-esterified fatty acids on the radioimmunoassay of prostaglandins

Non-esterified fatty acids (NEFA) can significantly interfere with the radioimmunoassay of PGE and PGF_2α using commercially available anti-sera. PGB₁ antigen-antibody binding is 50% inhibited by 110 pg of PGB₁, 48 ng of PGE₁, 3.5 μg of PGF_2α, or 9.0 μg linoleic, 14 μg arachidonic, 22 μg δ-linoleic, 40 μg palmitoleic or 45 μg oleic acids. PGF_2α antigen-antibody binding is 50% inhibited by 270 pg of PGF_2α, 70 ng of PGE₁, or 4.2 μg arachidonic, 14 μg δ-linolenic, 22 μg linoleic, 70 μg palmitoleic or 110 μg oleic acids. Physiological levels of NEFA, such as the quantities found in small volumes of plasma, are sufficient to prohibit accurate prostaglandin measurements. Chromatography on small columns of silicic acid proved to be an effective technique for separation of NEFA and prostaglandin from lipid extracts, however, the results of this study suggest that the interference produced by the presence of NEFA in the measurement of prostaglandin from certain physiological fluids may be avoided if the prostaglandins are not extracted prior to radioimmunoassay.     

Prostaglandin production by the pregnant and non-pregnant rabbit uterus

We have studied the prostaglandin synthesis of the pregnant and non-pregnant rabbit uterus in a microsomal membrane preparation, and in an perfused uterus preparation which retains agonist stimulated prostaglandin production. In both the microsomal and isolated perfused system, prostacyclin was the major arachidonic acid metabolite produced; PGE₂ was also produced in substantial quantities while TxB₂ and PGF_2α were not detectable. Moreover, oxytocin was a specific stimulus of PGE₂ release. the steroid hormone milieu influenced the level of agonist stimulated prostaglandin release; in general, ovariectomized, estrogen treated animals were more responsive to agonist stimulation than those treated with estrogen followed by progesterone. The microsomal studies indicated that the pregnant animal had a greatly enhanced capacity to metabolize arachidonic acid when compared with the non-pregnant animal. However, this was not reflected in the ability of agonists to stimulate prostaglandin release in the perfused preparation.     

Prostaglandin Biosynthesis from Endogenous Precursors in Rabbit Kidney

THIS report describes the biosynthesis of the naturally occurring renal prostaglandins E₂ (PGE₂) and F_2α (PGF_2α)^1,2 by homogenates and slices of rabbit renal medulla, from endogenous precursors. I have confirmed that rabbit renal cortex contains little prostaglandin and cannot synthesize them from endogenous lipids³. Hamberg has reported that arachidonic acid, which is converted to PGE₂ and PGF_2α by enzymes present in ram seminal vesicles⁴, can be efficiently converted to PGE₂ and PGF_2α by homogenates of rabbit renal medulla³. I have now confirmed that arachidonic acid, added to such medullary homogenates, can increase the quantities of prostaglandins synthesized. There was no evidence that the major prostaglandin biosynthesized, PGE2, was further metabolized to inactive products.