Stimulation Of Prostaglandin Synthesis Research Articles

Atrial natriuretic factor does not stimulate prostaglandin synthesis in isolated rat glomeruli.

The effect of alpha-human atrial natriuretic polypeptide (alpha-hANP) on the synthesis of prostaglandins was studied in isolated rat glomeruli. Glomeruli were isolated by a passive sieving technique according to the method of Misra and were incubated at 37 degrees C for 60 min in the presence (Group II: 10(-6) M, Group III: 10(-5) M) or absence (Group I: control) of alpha-hANP. Furthermore, glomeruli were incubated with arachidonic acid (10(-5), 10(-4), and 10(-3) M) and at the end of the incubation period trypan blue was added to the glomerular suspension. The presence of alpha-hANP (10(-6) and 10(-5) M) caused no significant difference in prostaglandin synthesis as compared with the control. On the other hand, arachidonic acid stimulated prostaglandin synthesis and the glomerular preparation was not stained by trypan blue, indicating that they remained viable. These results suggest that alpha-hANP does not directly affect the prostaglandin synthesis in isolated rat glomeruli.

Suppression by the cyclohexanetrione Ro 31-0521 of retinoic acid-induced teratogenicity.

The cyclohexanetrione Ro 31-0521, which stimulates prostaglandin synthesis, inhibited retinoic acid-induced cartilage degradation in vitro and suppressed the congenital forelimb malformations in rats treated with retinoic acid on day 13 of gestation in a dose-dependent manner.

Platelet-derived growth factor stimulates prostaglandin synthesis by direct effects on cyclooxygenase

It is generally believed that arachidonic acid liberation from cellular phospholipids controls formation of prostaglandins (PG). 4'~ Upon binding of many agonists to their respective high affinity cell surface receptor, arachidonic acid is released from phospholipids through activation of phospholipases. 5 There is little doubt that eicosanoid synthesis depends on phospholipase activation and arachidonic acid release. It is less clear, however, whether this step of the arachidonic acid cascade is sufficient to induce the burst of PG, thromboxane, and leukotriene formation observed in many agonist response systems. We have used cultured Swiss 3T3 cells stimulated by the potent mitogen Platelet-Derived Growth Factor (PDGF) as a model system to investigate arachidonic acid release from cellular phospholipids L2 and mechanisms of prostaglandin synthesis. 3 The question we have addressed in the present report has been whether PDGF has direct stimulatory effects on the cyclooxygenase, the key enzyme of prostaglandin, thromboxane, and prostacyclin synthesis. Figure 1 shows the effects of PDGE on prostaglandin E2 synthesis from endogenously released arachidonic acid. PDGF stimulates prostaglandin E2 synthesis in the cell cycle of Swiss 3T3 cells at two distinct time intervals: a first early increase that is complete within 10 rain, and a second increase between 2-4 hr. Quiescent cells incubated with increasing concentrations of arachidonic acid are unable to synthesize significant amounts of prostaglandin E2 (Fig. 2) while other experiments showed that PDGF has no comparable effect on the cellular pool of radiolabeled unesterified arachidonic acid (results not shown).

Mechanism of bradykinin, ADH, and cAMP interaction in rabbit cortical collecting duct.

Vasopressin (ADH) and bradykinin (BK) have been shown to stimulate prostaglandin synthesis in rabbit cortical collecting tubules. We studied ADH and BK effects on osmotic water flow (Lp), Na transport (JNa), and transepithelial voltage (VT). Bath BK but not lumen BK blunted subsequent ADH hydroosmotic responses. This BK effect was prevented by ibuprofen or pertussigen pretreatment and was overcome with exogenous cAMP, suggesting that BK, via prostaglandins, interferes with ADH action on Lp at the cAMP generation step. In contrast, bath BK had no effect on bath-to-lumen (Jb-1Na) or lumen-to-bath (Jl-bNa) Na flux or on VT. As reported by others, ADH lowered Jl-bNa and depolarized VT; however, prostaglandin synthesis inhibitors neither prevented nor reversed these ADH effects. Together, these BK and ADH data do not support regulation of JNa by peptide-stimulated prostaglandins. Moreover, cAMP alone depolarized VT but had no effect on Jl-bNa. Therefore, ADH-induced depolarization of VT may at least partly owe to cAMP effects on VT independent of accompanying changes in JNa. As with Lp, bath BK blunted subsequent ADH effects on VT and, to a lesser extent, Jl-bNa; these BK effects on ADH action were also prevented by ibuprofen or pertussigen pretreatment. The data are consistent with the following model: 1) ADH depolarizes VT and increases Lp via cAMP; 2) ADH decreases JNa via neither cAMP nor prostaglandins; and 3) BK, via prostaglandins, inhibits the actions of ADH on Lp and VT at the inhibitory guanyl-nucleotide regulatory subunit of adenylate cyclase.

Effects of pH on in vitro ovulation of goldfish (Carassius auratus) oocytes.

The in vitro effects of pH on goldfish (Carassius auratus) ovulation were investigated. Final oocyte maturation and follicular detachment were induced in vivo in gravid goldfish by HCG injections and elevated holding temperatures. Females were biopsied to determine the time of germinal vesicle breakdown and when oocytes should be removed for in vitro incubations. Prior to ovulation, the ovaries were removed, dissected and mature intrafollicular oocytes were incubated in media of varying pH (7.3-8.9). There was a significant increase in ovulation as the pH of the incubation increased and this ovulation could be blocked by indomethacin. Indomethacin did not inhibit the actual mechanism of oocyte expulsion since exogenous PGF2 alpha induced ovulation in indomethacin blocked incubates. Increased pH did not increase the ovulatory response observed with exogenous PGF2 alpha. The combined results suggest that an increase in incubation pH stimulates prostaglandin synthesis that, in turn, stimulates ovulation.

A postulated mechanism of beta-sympathomimetic induction of rib and limb anomalies in rat fetuses.

Treatment of gravid rats (days 6-15 of gestation) with the beta-sympathomimetic doxaminol resulted in wavy ribs and bent limbs in the offspring. The fetuses also exhibited defective mineralization. These anomalies were produced by pharmacologically effective doses of the drug. Prior treatment with the beta-receptor blocker carazolol prevented their formation, so that the beta-sympathomimetic action of doxaminol is evidently a causative factor. Various hypotensive agents whose activity is not mediated by beta-receptors failed to produce abnormalities. This eliminates the possibility of a non-specific etiology such as diminished placental perfusion. The cyclooxygenase inhibitor indomethacin lowered the incidence of wavy ribs. Furosemide, a loop diuretic that stimulates renal prostaglandin synthesis, increased the incidence of abnormalities when combined with doxaminol. The nature of the anomalies found suggests that 1) fetal compression by the myometrium and 2) defective mineralization are prerequisites for their development. The first condition could be produced via the complex mechanism of beta-sympathomimetic-induced stimulation of prostaglandin synthesis. Defective mineralization can result directly from cAMP-mediated activation of osteoclasts and possibly be further promoted by beta-sympathomimetic-mediated prostaglandin action on the osteoclast. The pathological findings in the fetal rat skeleton cannot be correlated with corresponding findings in human neonates whose mothers were subjected to prolonged therapeutic uterine relaxation with beta 2-sympathomimetics, for example. Since the anomalies in the rat disappear spontaneously in the post-natal period, their clinical relevance appears to be slight.

Human breast milk stimulates prostaglandin synthesis in cultured human skin fibroblasts

Effect of human breast milk or its fractions on prostaglandin synthesis was investigated in cultured human skin fibroblasts. Prostaglandins released into the media were measured by radioimmunoassay. Incorporation of breast milk (2% level) into 10% fetal calf serum media (for 48 hours) stimulated the synthesis of 6-keto-PGF 1α (stable product of prostacyclin) by 800%. This stimulating effect of milk persisted after cold acetone extraction to remove phospholipids and potentiated further after dialysis. Stimulation by one of the commercial formulas (Similac) was less than 50% of the milk effect. Milk also stimulated PGE 2 synthesis, although to a much lesser degree. These studies show for the first time that a) human breast milk contains potent factor(s) capable of influencing prostaglandin synthesis and suggest that b) these factors might have a role in the development of lipid synthetic pathways during early life.

Prostanoid synthesis in peripheral nerve

The transformation of [1- 14C]arachidonic acid into radiolabeled prostanoids was studied with homogenates and desheathed sciatic nerves of rats and frogs. All of the preparations studied were shown to synthesize prostaglandins; the specific prostanoids made were characterized by their migration on thin-layer chromatograms in three separate solvent systems. Both desheathed rat nerve and homogenates synthesize prostaglandin E 2, prostaglandin F 2α, prostaglandin D 2, 6-ketoprostaglandin F 1α and thromboxane B 2. With preparations from frog nerve, prostaglandin E 2 was the major prostanoid product formed. Several conditions were able to modulate the production of prostaglandin E 2 with desheathed frog nerve. Electrical stimulation at high frequency (100 Hz) for 30 min increased the formation of labeled prostaglandin E 2. Inclusion of glutathione also affected prostaglandin E 2 formation. A lower concentration (0.1 mM) stimulated prostaglandin synthesis, while 1 mM glutathione was partially inhibitory. In both the rat and frog system, prostanoid synthesis was suppressed by indomethacin and aspirin.

Human platelet-derived growth factor stimulates prostaglandin synthesis by activation and by rapid de novo synthesis of cyclooxygenase.

Human platelet-derived growth factor (PDGF) stimulated prostaglandin (PG) E2 synthesis in the cell cycle of Swiss 3T3 cells at two distinct time intervals, with a first plateau within 10 min and a second plateau within 2-4 h after addition of PDGF. At 4 h, the concentration of PGE2 in PDGF-stimulated cultures exceeded the quiescent control cells by a factor of 10-15. Quiescent cells incubated with up to 16 microM exogenous arachidonic acid (AA) synthesized only small amounts of PGE2. In contrast, 4 h after addition of PDGF, the concentration of PGE2 synthesized from exogenous AA exceeded that in quiescent cultures by a factor of 28. The effect of PDGF stimulation on PG synthesis from exogenous AA could not be explained by growth factor-mediated increase in the cellular free AA pool as shown in experiments using [14C]AA. PDGF also stimulated synthesis of PGI2 (prostacyclin), thromboxane, and PGF2 alpha from exogenous AA. While inhibition of protein synthesis by 10 micrograms/ml cycloheximide had no effect on the early increase in PGE2 synthesis, the second increase was completely prevented. Additionally, cycloheximide treatment at 6 h after PDGF stimulation resulted in rapid decline of PGE2 synthesis from exogenous AA. Quiescent cultures pretreated with 100 microM aspirin and stimulated by PDGF thereafter recovered from cyclooxygenase inhibition within 180 min. Our results suggest that phospholipase activation and resultant AA release is not sufficient to induce the burst of PG synthesis observed in PDGF-stimulated cells. Instead, PDGF stimulates PG synthesis by direct effects on the PG-synthesizing enzyme system, one involving a protein synthesis-independent mechanism and another that requires rapid translation of cyclooxygenase.

Interactions of vasopressin, cAMP, and prostaglandins in toad urinary bladder.

Prostaglandin (PG) inhibits the hydroosmotic effect of vasopressin. We therefore reexamined the interaction of vasopressin (VP), cAMP, and prostaglandins in toad bladder epithelial cells. Vasopressin slightly, but reproducibly, stimulated PGE2 and thromboxane B2 (TXB2) synthesis in cells prepared by the use of collagenase. When cells were prepared in the presence of a readily reversible cyclooxygenase inhibitor, ibuprofen, subsequent PGE2 synthesis was enhanced sevenfold but that of TXB2 was not. Increasing cAMP by either phosphodiesterase inhibition or 8-bromo-cAMP significantly inhibited both basal and VP-stimulated PGE2 synthesis. This inhibition was overcome by addition of arachidonic acid. Future studies employing these agents will have to consider these effects. VP enhanced 32P labeling of phosphatidylinositol (PI) and phosphatidic acid. This effect was prevented by the phosphodiesterase inhibitor, which also decreased phosphatidylcholine labeling. The results indicate that the phosphodiesterase inhibitor for cAMP may decrease PG formation by interfering with phospholipase activation. Furthermore, VP, similar to its effect in the liver, also increases PI turnover in toad bladder. This may initiate PG synthesis and provide a link among VP, cAMP, and calcium. A double-reciprocal feedback is proposed, whereby VP stimulates PG synthesis in a cAMP-independent manner and also inhibits PG synthesis in a cAMP-dependent manner.

Vasoconstrictor-evoked prostaglandin synthesis in cultured human mesangial cells.

We examined the influence of angiotensin II (ANG II), arginine vasopressin (AVP), and platelet activating factor (PAF) on prostaglandin (PG) synthesis and cell contractility in human glomerular mesangial cells in culture. Addition of sodium butyrate to the culture medium for 40 h significantly increased synthesis of both 6-keto-PGF1 alpha and PGE2 in the presence of exogenous arachidonic acid and of PGE2 under basal conditions. To optimize conditions in all further experiments, cells cultured with butyrate were studied. Under basal conditions, cultured mesangial cells produced predominantly 6-keto-PGF1 alpha and much less PGE2. Addition of either ANG II, AVP, or PAF all resulted in a rapid (within minutes) two- to threefold stimulation of 6-keto-PGF1 alpha and PGE2. Threshold stimulations were obtained at 10 pM for ANG II, 1 nM for AVP, and 10-100 pM for PAF. Preincubation of the cells with [Sar1,Ala8]ANG II, an antagonist of ANG II, inhibited ANG II-enhanced PG production, and preincubation with 1-desamino-8-D-arginine vasopressin, an antidiuretic analogue, blunted AVP-enhanced PG production. Under phase-contrast microscopy, PAF, ANG II, and, to a lesser degree, AVP caused decrease in cell surface area of mesangial cells cultured without butyrate at concentrations similar to those stimulating PG synthesis. Only PAF contracted cells cultured with butyrate, indicating attenuation of the vasoactive effects of ANG II and AVP when synthesis of PG was increased. However, a lower dose of PAF was only active when PG synthesis was inhibited, suggesting the same feedback mechanism for the three agonists.

Mechanism of action of angiotensin II and bradykinin on prostaglandin synthesis and vascular tone in the isolated rat kidney. Effect of Ca++ antagonists and calmodulin inhibitors.

We have studied the effect of angiotensin II and bradykinin on prostaglandin output and vascular tone during extracellular calcium depletion and administration of calcium antagonists and calmodulin inhibitors to elucidate the mechanism of action in the isolated rat kidney perfused with Tyrode's solution. Administration of angiotensin II (0.028-0.28 nmol) or bradykinin (0.28-2.8 nmol) enhanced the output of prostaglandin E2 and 6-keto-prostaglandin F1 alpha in a dose-dependent manner. Angiotensin II, but not bradykinin, produced renal vasoconstriction. Omission of calcium from the medium or infusion of calcium entry blockers, diltiazem (60 microM), or nimodipine (47 microM), failed to alter prostaglandin output elicited by angiotensin II or bradykinin; however, the effect of angiotensin II to produce renal vasoconstriction was inhibited. If calcium was omitted from the medium, the intracellular calcium antagonists, 8-(diethylamino)octyl 3,4,5-trimethoxybenzoate hydrochloride (23 microM), dantrolene sodium (31 microM), or ryanodine (2 microM), attenuated prostaglandin output caused by angiotensin II but not bradykinin. Calmodulin inhibitors, trifluoperazine (2 microM), napthalene sulfonamide hydrochloride (2 microM), or calmidazolium (2 microM), diminished prostaglandin output elicited by angiotensin II, but not that caused by bradykinin. Trifluoperazine, but not naphthalene sulfonamide or calmidazolium, attenuated the renal vasoconstrictor effect of angiotensin II. Prostaglandin output induced by angiotensin II and bradykinin were inhibited by mepacrine and indomethacin, whereas, the prostaglandin output caused by exogenous arachidonic acid (33 nmol) was abolished by indomethacin but was unaltered by mepacrine, calcium antagonists, and calmodulin inhibitors. From these data, we conclude that angiotensin II produces renal vasoconstriction by a mechanism dependent on extracellular calcium but not calmodulin, whereas angiotensin II-induced prostaglandin output depends on intracellular calcium and calmodulin. In contrast, bradykinin appears to stimulate prostaglandin synthesis by a calcium/calmodulin-independent mechanism.

Identification of C3b as the major serum protein that stimulates prostaglandin and thromboxane synthesis by macrophages

We have previously reported that heterologous, homologous and autologous sera, all stimulated rabbit alveolar macrophages to synthesize prostaglandins (PG). Gel permeation chromatography of serum showed multiple fractions possessing this stimulatory activity, with the major one at 150–160 K daltons. In the present study, we have shown that: (a) Fresh rabbit serum stimulated PG release by macrophages. (b) Serum depleted of C3 and C5 lost its stimulatory activity. (c) Trypsinized serum, sera activated by aggregated IgG and zymosan, partially purified C3, C5 and the C3, C5 preparation or purified C3 activated by zymosan, all stimulated PG release by macrophages with the following order of potency: activated C3, C5 = activated C3 = zymosan-activated serum > trypsinized serum = aggregated IgG-activated serum > partially purified C3, C5 = serum. PGE 2 was the predominant PG synthesized by stimulated macrophages. However, thromboxane (TX) production seemed to be more selectively enhanced i.e., increase in TX production was more pronounced than the increase in PGE release. To further identify the active complement component, we blocked the C3b receptor (C3bR) by preincubating macrophages with anti-C3bR, and showed that subsequent treatment with activated C3 and C5 failed to elicit any PG release. This pretreatment with anti-C3bR had no inhibitory effect on subsequent zymosan stimulation of PG release. Thus we concluded that C3b was the major serum protein that stimulates PG synthesis by macrophages.

Altering hydrodynamic variables influences PGI2 production by isolated lungs and endothelial cells.

Because deformation of lung tissue stimulates prostaglandin synthesis, we wanted to investigate whether hydrodynamic forces would affect lung prostacyclin (PGI2) production. To test the hypothesis that lung prostacyclin synthesis was flow dependent, we examined lung prostacyclin production after flow alterations. Using a salt solution that contained either Ficoll or albumin as a perfusate, we changed the flow to half and to double the control flow. When flow was changed, lung prostacyclin production followed changes in flow and pressure drop. When flow was varied in lungs treated with indomethacin, prostacyclin production was too low to be measurable. Variations in pressure pulsatility at constant mean flow had no influence on lung prostacyclin production. Since vascular distension may also stimulate prostacyclin production, we increased venous pressure. An increase in venous pressure (from 2.1 to 4.8 mmHg) had no effect on prostacyclin production; a further increase in venous pressure (to 7.5 mmHg) initiated edema and caused a large increase in prostacyclin production. When we subjected monolayers of endothelial cells cultured in wells to defined shear rates, the prostacyclin concentration in the supernatant quickly increased to a maximum. The absence of further increase with greater shear may have reflected feedback control of prostacyclin synthesis. The results indicated that hydrodynamic disturbances affect endothelial cells and stimulate arachidonate metabolism. Lung prostacyclin production may be related to flow. However, this effect is small compared with the lung prostacyclin production during edema formation.

Effect of glucocorticoids on vascular reactivity to vasoactive hormones in rat isolated kidney: lack of relationship to prostaglandins.

The relationship between the effects of glucocorticoids on renal vascular reactivity and prostaglandin synthesis elicited by noradrenaline (NA), angiotensin II (AII), arginine vasopressin (AVP) and bradykinin (Bk) was investigated in the isolated kidney of the rat perfused with Tyrode solution. Administration of NA 0.3-3.0 nmol, AII 0.028-0.28 nmol, AVP 0.027-0.27 nmol and Bk 0.28-2.8 nmol enhanced in a dose-dependent manner the renal output of immunoreactive prostaglandin E2 (PGE2) and 6-keto-PGF1 alpha. NA, AII and AVP, but not Bk, produced renal vasoconstriction and increased perfusion pressure. In the presence of dexamethasone (2.6 X 10(-5)M) or corticosterone (2.9 X 10(-5) M), the effects of NA and AII, in enhancing prostaglandin synthesis and producing renal vasoconstriction, were reduced. In contrast, stimulation of prostaglandin synthesis by Bk and AVP and the renal vasoconstriction produced by AVP were not altered by the glucocorticoids. Dexamethasone or corticosterone did not alter the output of prostaglandins elicited by A-23187 or arachidonic acid (AA). Addition of mepacrine (2.1 X 10(-5)M) to the perfusion fluid reduced the renal output of prostaglandins elicited by the vasoactive hormones and by A-23187, but not by AA; the vasoconstrictor response to NA and AII, but not to AVP was reduced. In kidneys in which prostaglandin synthesis was inhibited by indomethacin (2.8 X 10(-6)M), administration of dexamethasone also reduced the renal vasoconstrictor effect of NA and AII. These data indicate that in Tyrode-perfused rat kidney the glucocorticoids dexamethasone and corticosterone exert a differential effect on the renal vascular reactivity to vasoactive hormones, and that their inhibitory effect on NA and AII-induced renal vasoconstriction appears to be unrelated to prostaglandin synthesis.

Physiological aspects of blastocyst uterine interaction

An interaction between the blastocyst and the uterus is essential for establishment of pregnancy. Because maternal estrogen is not an absolute requirement, estrogen of embryonic origin has been implicated in this process in the pig and the rabbit. Furthermore, estrogen forming capacity has been documented in the blastocyst of these species. However, while the complete machinery for steroid synthesis in the pig balstocyst has been demonstrated, the issue is still unresolved for the rabbit blastocyst. In the present communication we have shown that 17α-hydroxylase and C17–20-lyase, enzymes involved in the formation of androgens (C19-steroids) from C21-steroids (progestins), are present in day-6 rabbit blastocysts. C17–20-lyase activity was undetectable to low in day-5 and increased in day-6 balstocysts. The activity was further increased in day-6 blastocysts cultured for 24 h. Because prostaglandins have been implicated in uterine vascular changes at about the time of implantation and pregnancy establishment, and because catechol estrogens are more potent than phenolic estrogens in stimulating prostaglandin synthesis in the blastocyst and the uterus, we determined catechol estrogen forming capacity in the rabbit and pig blastocyst. Catechol estrogen forming capacity (estrogen-2/4-hydroxylase) in the pig blastocyst appears on day 10 of pregnancy, peaks on day 12 and then declines. Our preliminary experiments also indicate that day-6 rabbit blastocysts have catechol estrogen forming capacity. On the basis of our present findings and of others, we propose that catechol estrogens of embryonic origin mediate the stimulatory effect of estrogens on prostaglandin synthesis in the embryo and/or the uterus and thus participate in the process of establishment of pregnancy.

The role of calcium in the stimulation of prostaglandin synthesis by vasopressin in rabbit renal-medullary interstitial cells in tissue culture.

The role of Ca2+ in the stimulation of prostaglandin (PG) biosynthesis by vasopressin was investigated in rabbit renal-medullary interstitial cells in tissue culture. A decrease in extracellular Ca2+ to less than 25 microM did not affect basal PGE2 production, but inhibited PGE2 synthesis stimulated by vasopressin, angiotensin and the Ca2+ ionophore A23187 by 55, 65 and 95% respectively. The study of vasopressin-stimulated PGE2 synthesis in the absence of extracellular Ca2+ demonstrated that: (a) hormone-sensitive phospholipase activity was inhibited as measured by [3H]arachidonic acid release; (b) the maximal rate of vasopressin-stimulated activity was decreased without a change in the vasopressin concentration that evoked half-maximal stimulation of PGE2 synthesis; and (c) the Ca2+-channel blocker verapamil and the Ca2+-calmodulin antagonist trifluoperazine mimicked the inhibitory effects of removing extracellular Ca2+. These agents had no effect in the absence of Ca2+. In contrast with their effects on vasopressin action, neither the removal of extracellular Ca2+ nor the addition of verapamil altered the ability of hyperosmotic mannitol to increase PGE2 synthesis. These data are consistent with the hypothesis that a component of vasopressin-stimulated PGE2 biosynthesis involves the influx of extracellular Ca2+, followed by the activation of Ca2+-calmodulin-stimulated phospholipase(s).

Binding of [3H]bradykinin in isolated nephron segments of the rabbit

Binding of intrinsically labeled [3H]bradykinin was studied in isolated nephron segments of the rabbit. Highest binding was observed in the cortical collecting tubule (5.76 +/- 0.34 X 10(-18) mol/mm) and the outer medullary collecting tubule (5.24 +/- 0.25 X 10(-18) mol/mm, means +/- SE, n = 6). Small but significant binding was also seen in the glomerulus, proximal straight tubule, cortical thick ascending limb of Henle's loop, and distal convoluted tubule. Lysyl-bradykinin, methionyl-lysyl-bradykinin, and tyrosine-8-bradykinin (but not des-arginine-9-bradykinin, vasopressin, angiotensins I and II, or prostaglandins) competed with [3H]bradykinin. The site of highest kinin binding (collecting tubule) is downstream from the highest concentration of kallikrein (granular portion of distal convoluted tubule). The binding data indicate that the major sites of kinin action in the kidney are the cortical and medullary collecting tubules. One action of kinins could be the stimulation of prostaglandin synthesis in the collecting tubules, which are known to actively synthesize prostaglandins.

Stimulation of prostaglandin synthesis in cultured rat synovial cells by a factor derived from polymorphonuclear leukocytes

Stimulation of synovial cell prostaglandin production by a factor obtained from casein-induced peritoneal polymorphonuclear (PMN) cells has been investigated. Both the extract and short time cultured medium of rat peritoneal PMN cells stimulate prostaglandin (PG) E 2 production as well as collagenase production in the culture of rat synovial cells. PGE 2 production by the cells in the presence of the PMN factor is much faster (5 to 24 hr) than collagenase production (24 hr or later, Biochemical Res. 3 , 506–516, 1982). This stimulating factor is confirmed to be derived from PMN cells, based on the purification of the cells from peritoneal exudate cells by the Ficoll-Urographin method. Elution profile of the factor on gel filtration has indicated that both PGE 2 and collagenase production by synovial cells are stimulated by the same effluent fractions corresponding to molecular weights of 15,000 – 20,000 daltons and 30,000 – 40,000 involved in PG production as well as collagenase production in the inflamed tissue by stimulating connective tissue cells such as synovial cells.

Stimulation of vascular smooth muscle cell prostacyclin and prostaglandin E2 synthesis by plasma high and low density lipoproteins.

We studied the effects of plasma high density and low density lipoproteins upon the synthesis of prostacyclin and prostaglandin E2 by vascular smooth muscle cells. Prostaglandin synthesis was measured in 24-hour cultures by radioimmunoassay of the stable metabolites of prostacyclin, 6-keto-prostaglandin F1 alpha and of prostaglandin E2. High density lipoproteins induced dose-dependent increases in the release of 6-keto-prostaglandin F1 alpha and of prostaglandin E2 from smooth muscle cells to values 14- and 50-fold above control. Incubations with low density lipoproteins at comparable cholesterol concentrations also induced dose-dependent release of 6-keto-prostaglandin F1 alpha and prostaglandin E2, but to a lesser extent. Rat high density lipoprotein, which contained 2.5 times more cholesteryl arachidonate than human high density lipoproteins, stimulated 6-keto-prostaglandin F1 alpha and prostaglandin E2 release 2- to 3-fold more than human high density lipoproteins, whereas the delipidated apoproteins of high density lipoproteins had no significant effect on prostaglandin synthesis. Recombinant high density lipoproteins containing cholesteryl-[1- 14C]arachidonate stimulated release of [14C]-6-keto-prostaglandin F1 alpha and [14C]prostaglandin E2 by smooth muscle cells. The ionophore, A 23187, released labeled 6-keto-prostaglandin F1 alpha and prostaglandin E2 from cells preincubated with recombinant high density lipoprotein containing cholesteryl-[1- 14C]arachidonate. Unlabeled high density lipoproteins, in contrast, did not cause release of radioactive prostaglandins from cells preincubated with [1- 14C]arachidonate. Phospholipase activators were synergistic (bradykinin) or additive (angiotensin II) with high density lipoprotein in stimulation of prostaglandin synthesis. The data indicate that both high and low density lipoproteins stimulate the synthesis of prostacyclin and prostaglandin E2 by vascular smooth muscle cells. The results suggest that the lipoproteins provide arachidonate to a phospholipase-sensitive pool accessible to cyclooxygenase.