Changes in prostaglandin concentration in blood subjected to repetitive freezing and thawing.

Successive freezing and thawing of whole blood results in a consistently higher yield of various prostaglandin (PG) compounds. Evaluations were made with radioimmunological assay. The increase in PG concentrations seems to be more associated with cell fragmentation and not with the dissociation of albumin-PG complex. Our data suggest that there may be some dissociation of non-albumin-PG complexes. Artifactually high PG concentrations due to in vitro PG synthetase activity appears minimal at least with respect to indomethacin blocking of this enzyme. There are, in general, only slight differences in PG concentrations in samples with and without indomethacin.     

Effects of indomethacin on intestinal secretion, prostaglandin E and cyclic AMP: Evidence against a role for prostaglandins in cholera toxin-induced secretion

The effects of indomethacin on intestine mucosal cAMP, intestinal fluid secretion, and mucosal and fluid PGE were studied in rabbits in vivo following challenge with cholera toxin. Indomethacin had no effect on cholera toxin-induced fluid secretion or cAMP accumulation. Inhibition of PGE synthesis was achieved by the administration of two but not one injection of indomethacin. These studies provide evidence against a role for PGE in mediating cholera toxin-induced secretion and point out the need to measure prostaglandin levels when using prostaglandin synthetase inhibitors in vivo.     

Methyl xanthine phosphodiesterase inhibitors behave as prostaglandin antagonists in a perfused rat mesenteric artery preparation.

The methyl xanthines, theophylline, caffeine and 3-isobutyl-1 methyl xanthine (MIX) inhibited the pressure responses to noradrnealine, angiotensin II and potassium ions in the isolated perfused mesenteric vascular bed of the male rat. The ID50s for inhibition of responses to noradrenaline were 1.85 mug/ml (0.83 x 10(-5) M) for MIX, 18 mug/ml (1 x 10(-4)M) for theophylline and 133 mug/ml (6.8 x 10(-4) M) for caffeine. Similar ID50 concentrations were found for responses to angiotensin II and potassium. We have previously found that substances which inhibit the three pressor agents equally may be prostaglandin (PG) synthesis inhibitors or PG antagonists. Xanthine itself, cyclic AMP and dibutyrl cyclic AMP had no inhibitory effects on the preparation up to concentrations of 10-2 M. Partial inhibition of PG synthesis by indomethacin shifted the % inhibition/log concentration curve to the left, while addition of exogenous PGE2 shifted it to the right. In preparations completely inhibited by sufficient indomethacin added to the perfusate to block PG synthesis, and then restored by adding 1 or 5 ng/ml PGE2 in addition to the indomethacin, the methyl xanthines again inhibited responses suggesting that they were PG antagonists rather than inhibitors of synthesis or release. In preliminary experiments MIX also inhibited effects of PGF2alpha on rat uterus and PGE1 on guinea pig ileum. Effective concentrations of theophylline were similar to the therapeutic levels in human plasma. PG antagonists may be a major action of methyl xanthines requiring reinterpretation of many experiments which have attributed their effects to PDE inhibition. PGs may also be involved in regulating PDE action.     

Compared effects of isoxicam and indomethacin on the urinary excretion of prostaglandins in degenerative articular diseases

The effects of a 7 day-treatment with isoxicam (200 mg/24 h) on the urinary excretion of prostaglandins (PG) were compared to those of indomethacin (150 mg/24 h) in a double-blind randomized study conducted in 18 patients with degenerative arthritic disease and normal renal function. Indomethacin decreased the urinary excretion of PGF2 alpha by about 70% and 6-keto-PGF1 alpha and thromboxane (Tx)B2, the stable break-down products of prostacyclin and TxA2 respectively, by about 40%. Isoxicam effects on urinary PG did not significantly differ from those of indomethacin. During both treatments, urinary gamma-glutamyl transferase and N- acetyl-glucosaminidase remained stable and none of the changes in the urinary excretion of PGs could be related to either plasma or urinary drug concentrations. In conclusion, chronic administration of isoxicam inhibited the renal PG biosynthesis to a similar extent than indomethacin which suggests that non steroidal anti-inflammatory drugs of the oxicam group ought also be used cautiously in patients with renal impairment.     

The role of endogenous prostaglandins in the regulation of gastric secretion in rhesus monkeys

Prostaglandins (PG) are known to alter a variety of gastrointestinal functions, but the physiological role of endogenous PG remains unclear. This experiment was designed to evaluate changes in gastric secretion following both acute and chronic inhibition of PG synthesis with indomethacin (5 mg/kg s.c.). Gastric juice was collected by continuous aspiration in 8 conscious chair-adapted male rhesus monkeys following treatment with saline or indomethacin for one or four days. The gastric juice was analyzed for H+, Na+, K+, and Cl- concentrations. The amount of soluble mucus in the gastric juice was estimated using Alcian Blue dye binding of acidic glycoproteins and Periodic Acid Schiff reaction with neutral glycoproteins. PG levels were measured in the plasma and in biopsy samples of fundus, antrum and duodenum. Both one and four days of indomethacin significantly (p less than 0.05) decreased tissue PG levels in the fundus, antrum and duodenum. Plasma levels of PGF2 alpha were significantly (p less than 0.05) decreased after both one and four days of indomethacin, while PGE2 and 6-keto PGF1 alpha were significantly inhibited only after four days of indomethacin. Both acute and chronic inhibition of PG synthesis was accompanied by a decrease in the concentration of sodium and mucus in the gastric juice but by an increase in the output and concentration of hydrogen ion. These changes suggest a possible mechanism by which endogenous PG play a role in the regulation of gastric secretion and in the protection against gastrointestinal damage.     

Evidence for the involvement of prostaglandins throughout the decidual cell reaction in the rat

Previous studies in which prostaglandin (PG) production was inhibited for a limited time by the s.c. administration of indomethacin have suggested that PGs are involved in the initiation of decidualization as well as the growth and differentiation of decidual cells. To reduce PG production during decidualization, in the present study indomethacin was infused from Alzet osmotic minipumps into the uterine lumen of ovariectomized rats with uteri sensitized for decidualization. To determine the effect of route of indomethacin administration on decidualization, rats received a single s.c. injection of indomethacin or its vehicle, and unilateral intrauterine infusion of indomethacin or its vehicle, in a factorial experiment. The inhibitory effects on decidualization, as assessed 5 days later by uterine weights, were greatest when both treatments were combined. Prostaglandins E and F concentrations 24 and 48 h after the insertion of the pumps were lower in the indomethacin-infused horns, suggesting that the indomethacin reduced uterine PG production. By contrast, subcutaneously administered indomethacin reduced uterine PG concentrations at 24 h but not at 48 h. Prostaglandin E2 and PGF2 alpha alone or combined, infused with indomethacin into the uterine lumen of rats treated subcutaneously with indomethacin, overrode the inhibitory effects of indomethacin. The dose-response relationships between these PGs and decidualization did not differ. These data suggest that PGs are required during the growth and differentiation of decidual cells from endometrial stromal cells.     

Methyl xanthine phosphodiesterase inhibitors behave as prostaglandin antagonists in a perfused rat mesenteric artery preparation

The methyl xanthines, theophylline, caffeine and 3-isobutyl-1 methyl xanthine (MIX) inhibited the pressure responses to noradrenaline, angiotensin II and potassium ions in the isolated perfused mesenteric vascular bed of the male rat. The ID50s for inhibition of responses to noradrenaline were 1.85 μg/ml (0.83 × 10⁻⁵M) for MIX, 18 μg/ml (1 × 10⁻⁴M) for theophylline and 133 μg/ml (6.8 × 10⁻⁴M) for caffeine. Similar ID50 concentrations were found for responses to angiotensin II and potassium. We have previously found that substances which inhibit the three pressor agents equally may be prostaglandin (PG) synthesis inhibitors or PG antagonists. Xanthine itself, cyclic AMP and dibutyryl cyclic AMP had no inhibitory effects on the preparation up to concentrations of 10⁻²M. Partial inhibition of PG synthesis by indomethacin shifted the % inhibition/log concentration curve to the left, while addition of exogeneous PGE2 shifted it to the right. In preparations completely inhibited by sufficient indomethacin added to the perfusate to block PG synthesis, and then restored by adding 1 or 5 ng/ml PGE2 in addition to the indomethacin, the methyl xanthines again inhibited responses suggesting that they were PG antagonists rather than inhibitors of synthesis or release. In preliminary experiments MIX also inhibited effects of PGF2α on rat uterus and PGE1 on guinea pig ileum. Effective concentrations of theophylline were similar to the therapeutic levels in human plasma. PG antagonism may be a major action of methyl xanthines requiring reinterpretation of many experiments which have attributed their effects to PDE inhibition. PGs may also be involved in regulating PDE action.     

The role of endogenous prostaglandins in the regulation of gastric secretion in rhesus monkeys

Prostaglandins (PG) are known to alter a variety of gastrointestinal functions, but the physiological role of endogenous PG remains unclear. This experiment was designed to evaluate chanes in gastric secretion following both acute and chronic inhibition of PG synthesis with indomethacin (5 mg/kg s.c.). Gastric juice was collected by continuous aspiration in 8 concious chair-adapted male rhesus monkeys following treatment with saline or indomethacin for one or four days. The gastric juice was anzlyzed for H⁺, Na⁺, K⁺ and Cl⁻ concentrations. The amount of soluble mucus in the gastric juice was estimated using Alcian Blue dye binding of acidic glycoproteins and Periodic Acid Schiff reaction with neutral glycoproteins. PG levels were measured in the plasma and in biopsy samples of fundus, antrum and duodenum. Both one and four days of indomethacin significantly (p < 0.05) decreased tissue PG levels in the fundus, antrum and duodenu. Plasma levels of PGF_2α were significantly (p < 0.05) decreased after both one and four days of indomethacin, while PGE₂ and 6-keto PGF_1α were significantly inhibited only after four days of indomethacin. Both acute and chronic inhibition of PG synthesis was accompanied by a decrease in the concentration of sodium and mucus in the gastric juice but by an incrase in the output and concentration of hydrogen ion. These changes suggest a possible mechanism by which endogenous PG play a role in the regulation of gastric secretion and in the protection against gastrointestinal damage.     

Effect of indomethacin in vivo on prostaglandin content of several rabbit tissues

The prostaglandin (PG) content of several tissues and fluids from 6 day pregnant rabbits was evaluated following treatment with indomethacin or vehicle in vivo. PGE and PGF were measured by radioimmunoassay. More complete depletion of PGE and PGF was accomplished by 3 injections of indomethacin (s.c.) given during the 18 h before sacrifice at a dose of 10 mg indomethacin per kg body weight than was accomplished by 1 injection of the same amount of indomethacin (i.v.) 1.5 h before sacrifice. Levels of PGF were more easily depressed by indomethacin than were those of PGE. PG levels in the kidney and blastocysts were depressed to a greater extent by indomethacin than were those in the uterus, uterine fluid or peritoneal fluid. Evaluation of the effect of indomethacin on a particular physiological function should be interpreted with caution unless the extent of PG depletion in that tissue is also measured.     

Effect of indomethacin on prostaglandin content of several rabbit tissues

The prostaglandin (PG) content of several tissues and fluids from 6 day pregnant rabbits was evaluated following treatment with indomethacin or vehicle . PGE and PGF were measured by radioimmunoassay. More complete depletion of PGE and PGF was accomplished by 3 injections of indomethacin (s.c.) given during the 18 h before sacrifice at a dose of 10 mg indomethacin per kg body weight than was accomplished by 1 injection of the same amount of indomethacin (i.v.) 1.5 h before sacrifice. Levels of PGF were more easily depressed by indomethacin than were those of PGE. PG levels in the kidney and blastocysts were depressed to a greater extent by indomethacin than were those in the uterus, uterine fluid or peritoneal fluid. Evaluation of the effect of indomethacin on a particular physiological function should be interpreted with caution unless the extent of PG depletion in that tissue is also measured.     

Adenosine as a natural prostaglandin antagonist in vascular smooth muscle

Adenosine has actions on smooth muscle similar to those of prostaglandin (PG) antagonists. Like some PG antagonists it is a phosphodiesterase inhibitor and seems to interfere with calcium effects. It has agonist/antagonist interactions with theophylline, a PG antagonist. In rat mesenteric vascular smooth muscle adenosine blocked responses to noradrenaline which depend on release of intracellular calcium but not those to potassium ions which depend on calcium entry from extracellular fluid. Partial inhibition of endogenous PG synthesis by indomethacin enhanced the adenosine effect. In preparations in which vascular reactivity had been abolished by indomethacin and then partly restored by 1 or 5 ng/ml PGE2, adenosine also inhibited responses to noradrenaline: the curve for the 5 ng/ml PGE2 concentration was to the right of and parallel to the 1 ng/ml curve consistent with a competitive interaction between adenosine and PGE2. Similar interactions between adenosine and PGE2 were shown in human lymphocytes in which activation also depends on calcium release. These findings suggest how calcium-dependent metabolic responses may be controlled and indicate further reasons for caution in the interpretation of cyclic AMP experiments.     

Prostaglandin F-2 alpha receptors in corpora lutea of pregnant rats and relationship with induction of 20 alpha-hydroxysteroid dehydrogenase

Luteal receptors for PGF-2 alpha in the pregnant rat were characterized. No changes in the Kd were found during pregnancy, whereas capacity increased to a maximum on Day 19, decreasing thereafter. The decrease in binding sites seen from Days 20 to 22 may be due to down regulation of the receptor by its ligand, since it was prevented by inhibition of PG synthesis by indomethacin treatment. Likewise, in-vivo treatment with PGF-2 alpha reduced the apparent number of PG binding sites. PG receptor concentration seems to be modulated by oestrogens since an increment was found on Day 19, associated with the known increase in plasma oestradiol concentrations, and since receptor concentration on Day 16 was significantly increased by oestradiol benzoate. The uterus also had a negative influence on the appearance of the PG receptor, since hysterectomy on Day 16 increased the number of binding sites on Day 18. However, receptor concentration and 20 alpha-hydroxysteroid dehydrogenase induction by hysterectomy was not affected by indomethacin, indicating that these events are probably not related to prostaglandin withdrawal. However, treatment with hCG, which diminishes enzyme induction by hysterectomy, did not produce changes in receptor concentration. The present results suggest that PGF-2 alpha, acting through a specific receptor site, is the physiological luteolytic signal. The consequence of its receptor binding seems to be the blockade of a gonadotrophic stimulus, which in turn determines (1) the decrease in progesterone synthesis and (2) the induction of 20 alpha-hydroxysteroid dehydrogenase.     

Effects of arachidonic acid and indomethacin on the in vitro release of prostaglandins by aortic strips of spontaneously hypertensive rats

Aortic strips removed from spontaneously hypertensive (SH) rats and preincubated with arachidonic acid (1.0 X 10(-5) g/ml) for 15 min produced two times more prostaglandin (PG) like material than aortae unexposed to the precursor of PG biosynthesis. The stimulating effect of arachidonic acid was largely inhibited by indomethacin (1.0 X 10(-5) g/ml). Also, the release of PG-like material by aortic strips derived from SH rats treated with an intravenous injection of indomethacin (10 mg/kg) was inhibited by 74% compared with the control tissues. These results raised the possibility that the in vivo conversion of arachidonic acid by large arteries of SH rats may contribute to the hypotensive effect of this PG precursor in SH rats.     

Effect of indomethacin invivo on prostaglandin content of several rabbit tissues

The prostaglandin (PG) content of several tissues and fluids from 6 day pregnant rabbits was evaluated following treatment with indomethacin or vehicle $\text{in}$$\text{vivo}$. PGE and PGF were measured by radioimmunoassay. More complete depletion of PGE and PGF was accomplished by 3 injections of indomethacin (s.c.) given during the 18 h before sacrifice at a dose of 10 mg indomethacin per kg body weight than was accomplished by 1 injection of the same amount of indomethacin (i.v.) 1.5 h before sacrifice. Levels of PGF were more easily depressed by indomethacin than were those of PGE. PG levels in the kidney and blastocysts were depressed to a greater extent by indomethacin than were those in the uterus, uterine fluid or peritoneal fluid. Evaluation of the effect of indomethacin on a particular physiological function should be interpreted with caution unless the extent of PG depletion in that tissue is also measured.     

Myo-inositol in physiological concentrations stimulates production of prostaglandin-like material

In physiological concentrations myo-inositol stimulated production of prostaglandin (PG)-like material in a rat mesenteric vascular bed preparation. There were five lines of evidence: 1. Inositol potentiated pressor responses to both norepinephrine and potassium in a manner similar to PGE2. 2. Inositol had no potentiating effect in preparations in which endogenous PG production was blocked by indomethacin. 3. Inositol caused no further potentiation in preparations already potentiated by arachidonic acid, the PG precursor. 4. The inhibitory effect of the PG antagonist chloroquine was reduced in an apparently competitive manner by inositol. 5. As indicated by rat stomach bioassay inositol caused a three fold rise in the outflow of PG-like material from the preparation.     

Effects of indomethacin on moloney sarcoma virus-induced tumors

Chronic administration of indomethacin to weanling (20 day old) BALB/c mice inoculated with Moloney sarcoma virus (MSV) delayed the onset of the tumor and suppressed the tumor growth. As expected, the prostaglandin (PG) levels associated with the MSV-injected leg muscle of the indomethacin treated mice were greatly depressed when compared to the elevated PG content associated with the tumors of MSV injected control mice. There was no effect of indomethacin on the cyclic AMP or cyclic GMP levels.The elevated levels of PG and cyclic AMP associated with the tumor were found to parallel the tumor growth.Administration of antilymphocyte serum (ALS) to the mice at the time of MSV inoculation resulted in accelerated and enhanced tumor growth. Indomethacin treatment of these mice similarly suppressed the tumor growth, but less dramatically.     

The role of alpha-adrenergic receptors in the vasoconstrictor response induced by indomethacin in the kidney.

The effect of indomethacin, an inhibitor of prostaglandin (PG) synthesis, was studied on the renal circulation, Na+ and water excretion in anaesthesized dogs during alpha-receptor inhibition. Indomethacin decreased cortical blood flow (CBFcontr, 454 +/- 142; CBFindo, 332 +/- 51 ml per min per 100 g; p less than 0.02) as well as medullary blood flow (OMBFcontr, 339 +/- 95; OMBFindo, 183 +/- 46 ml per min per 100 g; p less than 0.001), salt and water excretion, further it caused a shift in the intrarenal blood flow distribution toward the cortex. Alpha-blockade prevented the indomethacin-induced vasoconstriction in the cortex (CBF alpha inhibition + indo, 455 +/- 76 ml per min per 100 g) but not in the medullar (OMBF alpha inhibition + indo, 259 +/- 102 ml per min per 100 g, p less than 0.05). Alpha-blockade failed to prevent the indomethacin-induced antidiuresis, antinatriuresis and the intrarenal blood flow redistribution. GFR remained unaffected in all three series of studies. Our experimental findings are in line with the presumption that alpha-receptors are involved in the renal circulatory changes caused by indomethacin, probably as a result of an enhanced NE release during the inhibition of PG production. A NE--PG feed back mechanism is suggested in the regulation of renal circulation. The reduction of salt and water output induced by indomethacin appears to be independent of the alterations in renal haemodynamics, and seems rather to be the result of enhanced Na+ reabsorption, predominantly at the distal segment of the nephron, in the absence of PG, and/or a direct action of indomethacin.     

Effects of gonadotropin-releasing hormone agonists on meiotic maturation of follicle-enclosed oocytes in rabbits

The effects of GnRH agonists on in vitro maturation of rabbit follicle-enclosed oocytes were studied. Rabbit preovulatory follicles were cultured with or without hCG (10(2) ng/ml), buserelin (10(2)-10(5) ng/ml), or leuprolide (10(2)-10(5) ng/ml) for 14 hours in vitro. GnRH agonists induced the resumption of meiosis in the follicle-enclosed oocytes in a dose-dependent manner. The percentage of oocytes achieving GVBD following treatment with 10(5) ng/ml buserelin (87.9 +/- 6.3%) or 10(5) ng/ml leuprolide (86.0 +/- 4.1%) did not differ significantly from hCG-treated control (87.3 +/- 3.8%). Mature oocytes initially were detected within 2 hours of GnRH agonist exposure. Concomitant addition of a GnRH antagonist at 10(4) ng/ml significantly blocked the stimulatory effect of GnRH agonist on oocyte maturation. GnRH agonists significantly stimulated both prostaglandin (PG) E2 (PGE2) and PGF2 alpha production by preovulatory follicles (p less than 0.01), but secreted prostanoid levels did not differ significantly among different concentrations of GnRH agonists. Meiotic maturation of follicle-enclosed oocytes following GnRH agonist exposure began 2 hours earlier than production of PGs. PG production stimulated by GnRH agonists was reduced significantly by indomethacin. However, oocyte maturity in the presence of GnRH agonist plus indomethacin did not differ significantly from that of GnRH agonist alone. GnRH agonistic analogues induce the resumption of meiosis in follicle-enclosed oocytes in rabbits by a mechanism other than PG stimulation.     

Acute prostaglandin reduction with indomethacin and chronic prostaglandin reduction with an essential fatty acid deficient diet both decrease plasma flow to the renal papilla in the rat

Renal distribution of prostaglandin synthetase is mainly medullary, whereas the major degrading enzyme, prostaglandin dehydrogenase is primarily cortical. This suggests that prostaglandins (PG) released from the renal medulla could affect the medullary blood vessels. In two different experiments we studied the role of PG in the regulation of renal papillary plasma flow in the rat. First study: PG synthesis were stimulated in 34 adult Sprague-Dawley rats by bleeding from the femoral artery 1% of the body weight over a period of 10 minutes. Following this, indomethacin (a PG inhibitor, 10 mg/kg i.v.) was given slowly and then renal papillary plasma flow was measured 25 minutes after the end of infusion. In 17 indomethacin rats the renal papillary plasma flow averaged 18.8 ml/100 g/minute, whereas it averaged 23.0 in 17 non-indomethacin rats given diluent, an 18% reduction (p less than .025). Second study: Male Sprague-Dawley rats were made prostaglandin deficient by fasting rats for one week, followed by 10% dextrose fluid for one week and subsequent institution of an essential fatty acid (EFA) deficient diet for two weeks. With urinary PG excretion in prostaglandin deficient rats 28 ng/24 hours compared to 149 ng in control rats, they could be considered as prostaglandin deficient. When renal papillary plasma flow was measured, the 16 prostaglandin deficient rats had a 16% lower papillary plasma flow than 16 control rats, 21.6 vs 25.6 (p less than .005). These results clearly demonstrate that PG inhibition in rats decreases plasma flow to the papilla, strongly suggesting that PG are vasodilators for the vessels supplying the renal papilla.     

Indomethacin-induced augmentation of lymphoproliferative responses in patients with head and neck cancer

Augmentation of the phytohemagglutinin (PHA)-induced lymphoproliferation of peripheral blood mononuclear cells by indomethacin, a drug which blocks prostaglandin (PG) synthesis, was assessed in 37 patients with squamous cell carcinoma of the head and neck. Indomethacin enhanced the uptake of 3H-thymidine in stimulated cultures both from patients and normal individuals. However, because lymphoid cells from cancer patients were less reactive than those from normal controls, the proportionate increase in PHA-stimulated 3H-thymidine incorporation caused by indomethacin was greater in this population than in the normal population. The degree of enhancement induced by indomethacin did not correlate with the percent of esterase-positive mononuclear cells in the preparations. The amounts of PGE synthesized at 48 h by patients' or normal cells were similar. Cell populations that exhibited elevated levels of augmentation in the presence of indomethacin were approximately 3 times as sensitive to inhibition by 3 nM PGE2. The degree of augmentation detected in the presence of Ro-20-5720, which also prevents PG synthesis, was related to that produced by indomethacin. These results suggested that: the enhancing effect of indomethacin on lymphoproliferation in vitro was related to its inhibition of PG synthesis; and the sensitivity of lymphoid cells to inhibition by PGE2 was slightly, but significantly, increased in individuals with elevated augmentation values.