Assessment of plasma leukotriene and prostaglandin levels during adjuvant arthritis and kaolin-induced paw oedema in rats

Plasma levels of some arachidonic acid metabolites were investigated in acute and chronic models of inflammation in rats. As a model of chronic inflammation, adjuvant arthritis in rats induced by the injection of Freund's complete adjuvant, and as an acute model for inflammation, kaolin-induced paw oedama were used. Plasma leukotriene(LT) C4-like and prostaglandin(PG) E2-like activities were quantitated by bioassay in guinea-pig ileum and rat stomach fundus respectively. In the course of adjuvant arthritis, plasma levels of LTC4- and PGEi2-like activities were increased. Plasma LTC4-like activity reached a maximum within 3 weeks, while PGE2-like activity reached a maximum 10 days after adjuvant injection. In the early phase of adjuvant arthritis, levels of both LTC4- and PGE2-like activities were found to be low but both activities were increased in the late phase of inflammation.     

Changes of arterial arachidonic acid metabolites in hypertensive patients during haemodialysis

It is well known that in haemodialysis patients suffering from oligoanuria, extracellular hypervolaemia develops and this hypervolaemia is the main reason for hypertension occurring in some of the patients. The absence of vasorelaxation during hypervolaemia may be secondary to an increased activity of vasoconstrictor systems and/or a decreased formation of vasodilator agents like prostaglandin E2(PGE2) and prostaglandin I2(PGI2). In the present study, arterial PGE2 and leukotriene C4(LTC4)-like activities and the effect of fluid removal on these arachidonic acid metabolites during haemodialysis were measured in normotensive and hypertensive patients. Plasma PGE2 and LTC4-like activities were significantly different between hypertensive and normotensive patients. PGE2/LTC4 ratio did not change in normotensive patients while it was increased in hypertensive patients after haemodialysis. These results indicate that haemodialysis alters the synthesis of arachidonic acid metabolites especially in hypertensive patients.     

Prostaglandin E2 and leukotriene C4 levels following different reperfusion periods in rat brain correlated with morphological changes.

Prostaglandin E2 (PGE2) and leukotriene C4 (LTC4) are the metabolites of arachidonic acid (AA) that increase in forebrain following global ischemia and reperfusion. These mediators are highly potent vasoconstrictors of cerebral arteries leading to enhanced vascular permeability that induces the formation of vasogenic edema. In this study, after developing an experimental animal model simulating the concept of ischemic penumbra in the rat, the levels of PGE2 and LTC4 produced in the forebrain were measured and the effects of these mediators in short duration and prolonged reperfusion were investigated and then correlated with neuropathological findings. We found statistically significant reduction both in PGE2 and LTC4-like activities after just 10 min ischemia (p less than 0.05, p less than 0.05). PGE2-like activity significantly increased in the 4th and 60th min of reperfusion (p less than 0.05, p less than 0.05). In the 15th min of reperfusion, PGE2 was found to be significantly reduced (p less than 0.005) that may be due to the formation of free oxygen radicals by activation of PG hydroperoxidase reaction that inhibits PGE2 production in the cyclooxygenase pathway. LTs were not significantly increased in any reperfused group. Inhibition of the lipoxygenase pathway of AA metabolism may occur as a result of 15-HPETE (15-hydroperoxyeicosatetraenoic acid) production. Pathologically, edema and degeneration of brain tissue were seen beginning from the 4th min of reperfusion that reached a peak in the 60th min of reperfusion which is in accordance with biochemical changes in the damaged tissue.(ABSTRACT TRUNCATED AT 250 WORDS)     

The combined use of isolated strips of guinea-pig lung parenchyma and ileum as a sensitive and selective bioassay for leukotriene B4

The biological effects of leukotriene (LT)B4 were compared, on a molar basis, with those of LTC4, LTD4, LTE4, 5-hydroxyeicosatetraenoic acid (5-HETE), PGD2, PGE1, PGE2, PGF2 alpha, PGI2, 6-oxo-PGF1 alpha, bradykinin (BK) and angiotensin II (Ang II) on isolated strips of guinea-pig lung parenchyma (GPP) and ileum smooth muscle (GPISM) superfused in series. LTB4 was similar to LTC4 and LTD4 on GPP, in relation to potency and contractions induced, but differed from LTE4 in being ten times more active and causing contractions of a much shorter duration of action on this tissue. However, unlike the other LTs, LTB4 produced contractions which were resistant to FPL 55712 (1.9 microM) and, when given repeatedly, caused tachyphylaxis in GPP. LTB4 was considerably more active on GPP than the other substances investigated. Further, PGD2, PGF2 alpha and PGI2 contracted GPP, the order of potency being PGD2 greater than PGF2 alpha approximately equal to PGI2, whereas PGE1 and PGE2 relaxed this tissue. In contrast to all other agonists tested which contracted GPISM, LTD4 displaying the highest activity, LTB4 was inactive on this tissue. 5-HETE and 6-oxo-PGF1 alpha were inactive on both GPP and GPISM. On the basis of differential effects of LTB4 on GPP and GPISM, this assay represents a simple and selective means to distinguish LTB4-like materials from other naturally-occurring substances likely to be generated in inflammatory fluids.     

The protective effect of thromboxane synthetase inhibitor UK 38485 against bile duct ligation induced liver injury

In order to elucidate the relation between tissue eicosanoids and liver injury due to bile duct obstruction, we have examined the effects of iloprost, a stable analogue of prostaglandin I₂ (PGI₂), and UK 38485 (UK), an inhibitor of thromboxane synthetase, on prostaglandin E₂ (PGE₂) and leukotriene C₄ (LTC₄) in guinea pig liver. 56 male guinea pigs were divided into the following groups: (i) sham operations (SHAM), (ii) bile duct ligated (BDL) group, (iii) guinea pigs given UK (5 μg/kg body wt intraperitoneally 10 min, 8 h and 16 h after bile duct ligation), and (iv) guinea pigs treated with iloprost (ILO) (2 μg/kg body wt intraperitoneally 10 min, 8 h and 16 h after bile duct ligation). Liver damage was assessed by blind quantitation of liver cell necrosis. Bile duct ligation caused an increase in tissue PGE₂-like activity and a decrease LTC₄-like activity. But the most pronounced elevation of PGE₂ was observed in ILO treated group. The LTC₄-like activity level improved significantly in the UK-treated BDL group compared with the BDL only and ILO treated animals. Also, UK was found to be beneficial in preventing the liver cell necrosis due to cholestasis. It is concluded that the ratio of PGE₂/LTC₄ in liver is a valuable marker for cholestatic injury.     

Sulfidopeptide-leukotrienes are major mediators of arachidonic acid-induced mouse ear edema

The inflammatory response of the mouse ear to topical application of arachidonic acid (2 mg/ear) was examined to study the roles of sulfidopeptide-leukotrienes (LTs) and prostaglandin (PG) E2 as mediators of edema. The increase in ear thickness caused by arachidonic acid (AA) (edema), reached a maximum at 45 to 60 min after AA application. The amounts of immunoreactive LTC4 and immunoreactive PGE2 produced increased significantly in 5 to 10 min, and then diminished gradually over 60 min. 5-lipoxygenase inhibitors, dual cyclooxygenase/lipoxygenase inhibitors and anti-histamines significantly inhibited AA-induced ear edema. Both production of PGE2 and LTC4 were suppressed by NDGA at 1 mg/ear which also inhibited ear swelling. However aspirin, which enhanced LTC4 production in AA-induced ear edema did not inhibit the ear swelling. Hypodermic injection of LTC4 at 25 ng or PGE2 at 500 ng/ear did not cause swelling, but edema was induced when both compounds were injected simultaneously. Moreover ear swelling was induced by injection of both LTD4 at 50 ng and PGE2 at 500 ng/ear. Furthermore, concomitant injection of histamine, at 500 ng or serotonin at 50 ng/ear with LTC4 at 25 ng caused ear swelling but both compounds at the same dose alone did not induce swelling. These results suggest that AA-induced ear edema is predominantly mediated by LTC4 and other lipoxygenase products while PGE2 (in the presence of LTs) acts to facilitated ear swelling, although serotonin and histamine may also contribute.     

Effects of prostaglandins and of leukotriene C4 on the metabolism of labelled glucose in uteri isolated from ovariectomized-diabetic rats. Influences of 17-beta-estradiol and of insulin.

The influences of exogenous PGE1, PGE2, PGF2 alpha, LTC4 and insulin (INS) on glucose oxidation in uterine strips isolated from ovariectomized-diabetic (OVD) and ovariectomized-estrogenized-diabetic (OVED) rats, were studied. The spayed animals were made diabetic by a single injection of streptozotocin (65 mg.kg-1 body weight). The effects of prostaglandins were studied in the presence of indomethacin (INDO) in the incubation medium and the effects of LTC4 in the presence of INDO and nordihydroguaretic acid (NDGA). These procedures were followed in order to avoid the possible influences of endogenous derivatives of arachidonic acid formed by the activity of cyclooxygenase and of lipoxygenases. INDO and NDGA did not modify significantly the formation of 14CO2 from U-14C-glucose in uteri from OVD and from OVED rats. INS (0.5 U.ml-1) augmented significantly labelled glucose metabolism, both in OVD as well as in OVED rats. On the other hand, added PGE1, PGE2, PGF2 alpha or LTC4 failed to alter glucose metabolism in uteri from OVD rats. Only PGE1 was able to increase significantly (p less than 0.05) 14CO2 production from labelled glucose in uterine strips from OVED rats. In OVD rats the stimulatory action of INS on uterine glucose metabolism was significantly enhanced by exogenous PGE1, but not modified by PGE2, by PGF2 alpha or by LTC4. PGE1, PGE2 and LTC4 sensitized uterine strips obtained from OVED rats to the effects of INS. The possible importance of PGE1 in improving uterine glucose metabolism in diabetic animals is discussed.     

Nicardipine reduces the levels of leukotriene C4 and prostaglandin E2, following different ischemic periods in rat brain tissue.

Ischemic depolarization of nerve membranes is associated with a rapid influx of calcium into the cell, resulting in production of arachidonic acid (AA) metabolites. These metabolites, particularly leukotriene C4 (LTC4) have a very potent vasoconstrictor effect on cerebral arteries inducing vasogenic edema that may damage the ischemic penumbra. Calcium antagonists are assumed to prevent or reduce metabolic disturbances associated with ischemia. In this study, after developing an experimental animal model simulating the concept of the ischemic penumbra in the rat, the levels of LTC4 and prostaglandin E2 (PGE2) produced in the forebrain following different ischemic periods, such as 4th, 15th, 60th and 240th min were measured by a bioassay method, including 6 rats for each ischemic group. Then the effect of the 1-4 dihydropyridine nicardipine (1 mg/kg) on these mediators was investigated by giving it to the rat 30 min before the development of the ischemic model in each corresponding group (n = 6). We showed that nicardipine significantly reduced the high levels of LTC4 and PGE2 in the 4th min and 4th h of cerebral ischemia (p less than 0.005, p less than 0.0005). So it may be concluded that institution of nicardipine may be helpful in protecting the ischemic penumbra during the early hours of cerebral ischemia.     

Leukotriene C4 and prostaglandin E2 activities in the serum and cerebrospinal fluid during acute cerebral ischemia

Lipoxygenase pathway products of arachidonic acid (AA) metabolism (known as leukotrienes, LTs) are produced in the brain during pathologic conditions such as ischemia, hemorrhage, trauma, and seizure in which the release of AA is sustained by the activation of local phospholipases. The most common type of LT in the central nervous system is an LTC4 which is a highly potent vasoconstrictor leading to increase in vascular permeability. In this study, we compared the serum (S) and cerebrospinal fluid (CSF) prostaglandin E2 (PGE2) and LTC4 levels in 13 consecutively admitted patients with acute cerebral ischemia aged 55-80 years with 10 age-matched controls. Patients with previous glucocorticosteroid and antiinflammatory drug usage were not included in the study. S and CSF samples were drawn during the first 72 h of the attack, and samples were evaluated by bioassay. There was no significant difference in S PGE2 and LTC4 values, whereas a significant difference was observed between CSF PGE2 and LTC4 values as compared with the control group. The high levels of CSF PGE2 and LTC4-like activity in acute cerebral ischemia may indicate that these mediators have a role to play in cerebral edema. The CSF PGE2/LTC4 ratio was also found to be reduced in the ischemic group implying higher LTC4 synthesis than PGE2 synthesis. In the light of these findings, we suggest that use of a selective antagonist of LTs may be helpful in reducing the ischemic penumbra during acute cerebral ischemia by controlling the vasogenic edema.     

Structural and physicochemical requirements of endotoxins for the activation of arachidonic acid metabolism in mouse peritoneal macrophages in vitro

Lipopolysaccharides of different wild-type and mutant gram-negative bacteria, as well as synthetic and bacterial free lipid A, were studied for their ability to activate arachidonic acid metabolism in mouse peritoneal macrophages in vitro. It was found that lipopolysaccharides of deep-rough mutants of Salmonella minnesota and Escherichia coli (Re to Rc chemotypes) stimulated macrophages to release significant amounts of leukotriene C4 (LTC4) and prostaglandin E2 (PGE2). Lipopolysaccharides of wild-type strains (S. abortus equi, S. friedenau) only induced PGE2 and not LTC4 formation. Unexpectedly, free bacterial and synthetic E. coli lipid A were only weak inducers of LTC4 and PGE2 production. Deacylated Re-mutant lipopolysaccharide preparations were inactive. However, co-incubation of macrophages with both deacylated lipopolysaccharide and lipid A lead to the release of significant amounts of LTC4 and PGE2, similar to those obtained with Re-mutant lipopolysaccharide. The significance of the lipid A portion of lipopolysaccharide for the induction of LTC4 was indicated by demonstrating that peritoneal macrophages of endotoxin-low-responder mice or of mice rendered tolerant to endotoxin did not respond with the release of arachidonic acid metabolites on stimulation with Re-mutant lipopolysaccharide and that polymyxin B prevented the Re-lipopolysaccharide-induced LTC4 and PGE2 release. Physical measurements showed that the phase-transition temperatures of both free lipid A and S-form lipopolysaccharide were above 37 degrees C while those of R-mutant lipopolysaccharides were significantly lower (30-35 degrees C). Thus, with the materials investigated, an inverse relationship between the phase-transition temperature and the capacity to elicit LTC4 production was revealed.     

The effect of cocaine on gastric mucosal PGE(2), LTC(4) and ulcerations

The association between cocaine use and acute gastroduodenal perforation is known. The effect of cocaine and stress on gastric mucosal ulceration and the levels of prostaglandin E(2) (PGE(2)) and leukotriene C(4) (LTC(4)) was studied in 40 Sprague-Dawley rats. Controls received intraperitoneal (i.p.) saline, ten received i.p. cocaine (35 mg/kg), ten were stressed by the cold restraint method, and ten had i.p. cocaine and stress. Cocaine alone did not induce ulceration, but decreased PGE(2) levels. Stress alone caused ulceration, but was not associated with a change in either PGE(2) or LTC(4) levels. When combined with stress, however, cocaine caused a three-fold increase in ulceration and a significant increase in PGE(2) and LTC(4) levels. Stress may predispose the cocaine addict to loss of gastroduodenal mucosal integrity, which is related to an imbalance of PGE(2) and LTC(4) synthesis.     

The extraction of leukotrienes (LTC4, LTD4, and LTE4) from tissue fluids: the metabolism of these mediators during IgE-dependent hypersensitivity reactions in lung

The metabolites of arachidonic acid known as the leukotrienes are a class of lipid mediators which have potent and diverse biological effects in pulmonary tissue. Leukotrienes C, D, and E (LTC4, LTD4, and LTE4) are known to be principal mediators of immunoglobulin E (IgE)-mediated hypersensitivity reactions in lung tissue. It is therefore important to develop reliable and quantitative isolation techniques for estimating levels of these mediators in tissue. In this study, LTC4, LTD4, and LTE4 were separated from other arachidonate metabolites by organic extraction procedures. 5-Hydroxyeicosatetraeonic acid and leukotriene B4 extract efficiently into the organic layer of aqueous:ether or aqueous:chloroform extractions, whereas arachidonate metabolites containing conjugated peptides (e.g., LTC4, LTD4, and LTE4) failed to extract into these organic solvents. An extraction step was therefore developed that affords quantitative extraction of LTC4, LTD4, and LTE4 into the organic phase of an isopropanol:ether:H2O mixture. This step is the key for a two-step extraction method that isolates histamine, LTC4, LTD4, and LTE4 with a recovery of 100, 85, 75, and 57%, respectively. One advantage of this separation procedure for obtaining these mediators by organic extraction is an ability to expediently process many samples. Furthermore, the leukotriene content of extracted samples can be analyzed using the guinea pig ileum bioassay without interference from vasoamines or platelet-activating factor. These later substances are eliminated from leukotriene-enriched fractions by this extraction process. When histamine and LTC4 were added to supernatant fluids recovered from isolated lung tissue, they were quantitatively recovered using this extraction method.(ABSTRACT TRUNCATED AT 250 WORDS)     

Intraperitoneal injection of zymosan in mice induces pain, inflammation and the synthesis of peptidoleukotrienes and prostaglandin E2

Intraperitoneal injection of zymosan in mice induced rapid extravasation and accumulation of plasma protein in the peritoneal cavity. Neutrophils began to appear in the peritoneal cavity after a lag period of approximately 3 hours. The injected mice exhibited a pain response (writhing) during the first 30 minutes after injection, but writhing ceased before protein or cell accumulation had reached maximum levels. The injection of zymosan induced synthesis of PGE2 (measured by RIA) which reached maximum levels at 30 minutes, then declined slowly. Peptido-leukotriene levels (detected by bioassay, RIA and HPLC) increased rapidly after injection, reached a peak within an hour of injection and declined to undetectable levels within 4 hours. The early peptido-LT was predominantly LTC4, while later, LTE4 was the major component. LTD4 levels remained low throughout and no LTB4 was detected at any time. Indomethacin treatment elevated levels of peptido-LTs, reduced PGE2 levels and inhibited writhing. Phenidone reduced peptido-LT levels. In vitro studies demonstrated that zymosan stimulates LTC4 synthesis by peritoneal cells whereas LTE4, LTD4, LTB4 or monoHETES were not detectable (using HPLC methods). The source of enzymes responsible for the in vivo metabolism of LTC4 to LTD4 and LTE4 could not be identified.     

Specific binding of a stable prostacyclin analogue (Iloprost) to rat oxyntic mucosa

It has been reported that prostacyclin (PGI2) is the predominant species of prostanoid in rat oxyntic mucosa. However since PGI2 is inactivated under physiological conditions it has not been possible to demonstrate specific PGI2 binding to the rat stomach. Therefore a stable PGI2 analogue, Iloprost, was chosen as ligand in this study. Binding of labelled Iloprost to the 20,000 xg homogenate fraction of rat oxyntic mucosa was specific, dissociable, saturable and dependent upon the temperature and time of incubation. Neither tritiated PGE2 nor 6 keto PGF1 alpha displayed any significant specific binding to rat stomach. A Scatchard plot of the equilibrium binding data for Iloprost was curvilinear and could be resolved into at least two binding sites. The average parameters determined from Scatchard analysis were: dissociation constants of 1.8 X 10(-11) M and 7.1 X 10(-8) M and corresponding binding site concentrations of 12.0 pmole/mg and 4800 pmoles/mg protein respectively. PGI2 was less potent than unlabelled Iloprost in displacing 3H-Iloprost from its binding site. The addition of PGE2 to the incubation medium resulted in an increase in 3H-Iloprost binding. It is concluded that rat oxyntic mucosa has specific binding sites for PGI2-like agents but not for either PGE2 or 6 keto PGF1 alpha.     

Changes in the concentrations of hepatic metabolites on administration of dihydroxyacetone or glycerol to starved rats and their relationship to the control of ketogenesis

1. Glycerol and dihydroxyacetone, both antiketogenic and readily metabolized, but differing in their effects on the redox state of the hepatic NAD couples, were given to starved rats and the contents of metabolites were measured in freezeclamped liver and in the blood. The object was to study the effects of changes in the redox state and of the availability of oxidizable substrates on the rate of ketone-body formation. 2. Intramuscular administration of dihydroxyacetone, glycerol or glucose to starved rats decreased the concentrations of acetoacetate and 3-hydroxybutyrate in the blood by 70-80% within 60min., whereas there was no major change in the free fatty acid concentration. 3. Dihydroxyacetone, but not glucose or glycerol, caused an immediate and sustained twofold increase in the blood lactate concentration. 4. Dihydroxyacetone and glycerol caused a rapid fall in the hepatic concentrations of ketone bodies, dihydroxyacetone being more effective. 5. This decrease was not accompanied by significant changes in the concentrations of acetyl-CoA, long-chain acyl-CoA or free CoA. 6. The hepatic glycerophosphate concentration rose about 40-fold on administration of glycerol, whereas with dihydroxyacetone the increase was only about 50%. The large increase in glycerophosphate concentration after administration of glycerol was completely prevented by pretreatment of the rats with tri-iodothyronine. Triiodothyronine-treated rats showed the same decrease in ketone-body concentrations after administration of glycerol as the untreated rats. 7. Glycerol and dihydroxyacetone caused an increase in the hepatic lactate concentration; the pyruvate concentration rose only after injection of dihydroxyacetone. 8. Both compounds increased liver glycogen. 9. Calculation of the [free NAD(+)]/[free NADH] ratios indicated that dihydroxyacetone increased the ratio in cytoplasm and mitochondria, whereas glycerol caused a prompt fall in both compartments, followed at 10min. by a slight rise in the mitochondrial compartment. 10. Dihydroxyacetone did not alter the hepatic content of ATP. 11. The findings suggest that the main reason for the antiketogenic effect of glycerol and dihydroxyacetone was a consequence of their ready metabolism and the provision of an increased supply of C(3) intermediates for conversion into oxaloacetate. Under the test conditions, neither the hepatic content of alpha-glycerophosphate nor the redox state of the NAD couples appeared to play a major role in the regulation of ketogenesis.     

The effects of prostacyclin analogue ZK 36374 and thromboxane synthetase inhibitor UK 38485 on mesenteric ischemia in guinea pigs.

The effects of ZK 36374, a prostacyclin analogue and UK 38485, a thromboxane synthetase inhibitor were studied in guinea pigs after performing mesenteric arterial occlusion. In this study, while ZK 36374 significantly lowered the alkaline phosphatase and creatine phosphokinase values two hours after mesenteric arterial occlusion when compared with the control group (p less than 0.005), UK 38485 did not induce any change. In guinea pigs, when given together, ZK 36374 and UK 38485 lowered the enzyme levels to preligation values and the difference was nonsignificant (p greater than 0.1). The histopathologic investigation of the small intestine after giving ZK 36374 and UK 38345 together revealed minimal changes. These findings stress the importance of preserving the PGI2 levels in the PGI2/TXA2 ratio in preventing the increase of lysosomal enzyme levels and histopathologic changes after mesenteric arterial occlusion in guinea pigs.     

Inhibition by 16,16-dimethyl PGE2 of ethanol-induced gastric mucosal damage and leukotriene B4 and C4 formation

The effects of PGE2 and its stable analogue, 16,16 dimethyl PGE2 (dmPGE2) were investigated on ethanol-induced gastric mucosal haemorrhagic lesions and leukotriene formation in the rat. Exposure of the rat gastric mucosa to ethanol in-vivo, produced a concentration-related increase in the mucosal formation of leukotriene B4 (LTB4) which was correlated with macroscopically-apparent haemorrhagic damage to the mucosa. Challenge with absolute ethanol likewise enhanced the mucosal formation of LTC4 whereas the mucosal formation of 6-keto-PGF1 alpha was unaffected. Challenge of the rat gastric mucosa in vitro with ethanol induced a concentration-dependent increase in the formation of LTB4 and LTC4, but not 6-keto PGF1 alpha. Pretreatment with PGE2 (200-500 micrograms/kg p.o.) prevented the haemorrhagic mucosal damage induced by oral administration of absolute ethanol but not the increased formation of leukotrienes by the mucosa. In contrast, pretreatment with a high dose of dmPGE2 (20 micrograms/kg p.o.) prevented both the gastric mucosal lesions and the increase mucosal leukotriene formation. The differences in the effects of these prostaglandins may be related to the nature or degree of protection of the gastric mucosa. Thus, high doses of dmPGE2 but not PGE2 may protect the cells close to the luminal surface of the mucosa and hence reduce the stimulation of leukotriene synthesis by these cells.     

The effects of indomethacin, NDGA, allopurinol and superoxide dismutase on prostaglandin E2 and leukotriene C4 levels after mesenteric ischemia-reperfusion injury

In this study, the changes of arachidonic acid metabolites after an ischemia-reperfusion (I/R) period are investigated. The cyclooxygenase and lipoxygenase metabolites were found to be significantly increased after a 45 min period of ischemia followed by 5 min of reperfusion. Prostaglandin E₂ (PGE₂)- and leukotriene C₄ (LTC₄)-like activities did not change in the ischemic period, but they both increased after reperfusion. A cyclooxygenase inhibitor indomethacin and lipoxygenase inhibitor nordehydroguaretic acid (NDGA) decreased PGE₂- and LTC₄-like activities, respectively, while allopurinol and superoxide dismutase (SOD) decreased both activities.According to our results, it can be assumed that free oxygen radicals are responsible for the elevation of PGE₂- and LTC₄-like activities and both of these arachidonic acid metabolites and free oxygen radicals are the main necrotizing agents in ischemia-reperfusion induced damage.     

Regulation of gluconeogenesis by glycerol and its phosphorylated derivatives

Glycerol, glycerol-3-phosphate (G3P), and dihydroxyacetone phosphate (DHAP) were evaluated as inhibitors of gluconeogenesis on rat liver enzymes in vitro, and for their effects on glucose formation in vivo in well-nourished and malnourished rats. DHAP was more potent as an inhibitor than G3P on fructose-1,6-diphosphatase (FDPase), phosphoenolpyruvate carboxykinase (PEPCK), and glucose-6-phosphatase (G6Pase). The I50 for DHAP was 2, 8, and 9 x 10(-3) M, respectively. No effect was observed on rat liver pyruvate carboxylase (PC). Glycerol was a weak inhibitor of FDPase and PEPCK, but did not inhibit PC and G6Pase. In vivo, when G3P was injected before a parenteral L-alanine (Ala) challenge, it produced a hypoglycemic effect in malnourished rats and a lesser, but noticeable, blood glucose level reduction in well-fed animals. Glycerol caused a smaller reduction in glucose formation from Ala. No comparable effects were observed after a fructose pretreatment. These results underscore the potential hypoglycemic effects of phosphorylated glycerol metabolites and identify the steps in gluconeogenesis where this action is exerted. The study also stresses the nutritional component in the glycerol intolerance syndrome, apparent from the far more severe effects observed in malnourished rats given G3P or glycerol prior to Ala.     

Questionable role of leukotriene B4 in monosodium urate (MSU)-induced synovitis in the dog

Monosodium urate (MSU)-induced synovitis in the dog's stifle (knee joint) is similar to an acute gouty attack in man in which a loss of function of the joint correlates with massive influx of neutrophils and the release of an assortment of inflammatory mediators (e.g. histamine, bradykinin, lysosomal enzymes, complement and eicosanoids) into the synovial space. We found in the urate-induced inflammatory exudates 3 hr post MSU the following: 88 million leukocytes/ml (approximately 95% neutrophils) and eicosanoid concentrations of LTB4, LTC4, and PGE2 of less than 0.1, 1.4 and 20 ng/ml, respectively. Isotonic saline injected knee joints at 3 hr contained 5 million leukocytes/ml (approximately 95% neutrophils) and concentrations of LTB4, LTC4, and PGE2 of less than 0.1, 0.7 and 0.2 ng/ml, respectively. Intrasynovial injections of 1 microgram LTB4, 10 micrograms PGE2 or the combination of LTB4 and PGE2 produced no reduction of paw pressure for up to 3 hr. Leukocyte concentrations measured at 3 hr in joints injected with these arachidonic acids metabolites were similar to saline controls. These results question the role of LTB4 as a chemotactic and inflammatory mediator in urate-induced synovitis in the dog but confirm the importance of PGE2 and possibly LTC4 in this model.