Inhibition of histamine-sensitive adenylate cyclase from guinea pig fundic gastric mucosa by arachidonic acid and by an analog of prostaglandin endoperoxide PGH2

The effects of prostaglandin precursors, namely an analog of prostaglandin endoperoxide PGH₂ [(15S)-hydroxy-9α,11α-(epoxymethano)prosta-5, 13-dienoic acid] and arachidonic acid, were assessed on gastric adenylate cyclase activity from cell-free preparations of guinea pig fundic mucosa. The two precursors were tested against basal adenylate cyclase activity and that stimulated by histamine (10^?4M), by PGE₂(10^?4M), by 5′-guanylylimidodiphosphate [Gpp(NH)p] (10^?4M) and by NaF(10^?2M). PGH₂ analog (10^?4M) and arachidonic acid (10^?4M) both inhibited to a similar extent adenylate cyclase stimulated by histamine or by NaF, but not that stimulated by PGE₂ or by Gpp(NH)p. Neither agent significantly affected basal adenylate cyclase levels. In the presence of indomethacin (10^?4M), basal adenylate cyclase activity remained unchanged but the inhibitory effect of arachidonic acid was almost entirely abolished, suggesting that such inhibitory effect may be caused by prostaglandin endoperoxides generated from arachidonic acid in the course of assay. Moreover, indomethacin did not attenuate PGH₂ inhibition of histamine action. Unlike arachidonic acid, which is a natural metabolic precursor of PGE₂, arachidic acid did not significantly influence histamine-stimulated adenylate cyclase activity. These results suggest that the prostaglandin endoperoxides may have an inhibitory effect on histamine-sensitive ciclic AMP generation in gastric mucosa.     

Effects of prostaglandin H2, prostaglandin E2, and arachidonic acid on parathyroid hormone and antidiuretic hormone activation of rat kidney adenylate cyclase

These experiments examine interactions of arachidonic acid; the substrate for prostaglandin cyclooxygenase, prostaglandin (PG)H₂, a key endoperoxide intermediate in prostaglandin synthesis; and prostaglandin (PG)E₂, an important prostaglandin produced within the kidney; with adenylate cyclase activity in renal cortex, outer medulla, and inner medulla. In addition, the effects of arachidonic acid, PGH₂, and PGE₂ on parathyroid hormone (PTH) activation of adenylate cyclase in cortex, and of antidiuretic hormone (ADH) activation of that enzyme in outer and inner medulla are examined. Arachidonic acid elicited a concentration-dependent inhibition of basal and PTH-stimulated adenylate cyclase activity in renal cortex. Concentration-dependent inhibition by arachidonic acid of basal and ADH-stimulated adenylate cyclase activity was observed in outer and inner medulla. PGH₂ inhibited basal activity in all three areas of the kidney. There was also inhibition by PGH₂ of medullary ADH and cortical PTH stimulation. PGE₂ stimulated adenylate cyclase in all three areas. PGE₂ had no effect upon PTH stimulation in cortex and was additive with ADH in outer and inner medulla. PGE₂ stimulation was inhibited by arachidonic acid, and this inhibition seemed competitive. Inhibition by both arachidonic acid and PGH₂ was not destructive. Experiments with [1-¹⁴C]arachidonic acid and indomethacin suggest that the inhibition by arachidonic acid was actually mediated by arachidonic acid and not a metabolite. Both PGH₂ and arachidonic acid inhibition was independent of phosphodiesterase. This activation by product, PGE₂, and inhibition by its precursors, arachidonic acid and PGH₂, provide a possible mechanism by which the prostaglandin system could modulate adenylate cyclase responsiveness to hormonal activation.     

Microcirculatory effects of prostaglandins E1, E2, and A1 in the rat mesentery and cremaster muscle

The present investigation was carried out to determine the site(s) of vasomotor activity of Prostaglandins E₁ (PGE₁), E₂ (PGE₂), and A₁ (PGA₁) in the terminal vascular bed of the rat mesentery and cremaster (skeletal) muscle. Studies were also performed to determine the effects of PGE₁ and PGA₁ on arteriolar responsiveness to vasoconstrictor agents. In vivo changes in microvascular diameters in response to topical application of prostaglandins and the vasoconstrictor agents were determined quantitatively with an image shearing television microscope and recording system. In the mesentery, PGE₁ elicited arteriolar dilator responses, while similar doses of PGE₂ and PGA₁ were ineffective. The responses of mesenteric arterioles to PGE₁ were not entirely dose-dependent. In the cremaster muscle, the three prostaglandins studied produced a dose-dependent dilation of all muscular microvessels; the order of potency was PGE₁ > PGE₂ > PGA₁.Mesenteric and cremasteric arteriolar responses to epinephrine, norepinephrine, angiotensin, and vasopressin were inhibited by PGE₁, and this inhibition was demonstrable after the dilator action of PGE₁ had terminated. PGA₁ did not alter vascular responsiveness in either tissue. It was concluded that prostaglandins, by virtue of their vasodilator actions and inhibitory effects on microvascular responsiveness, may contribute to local control of blood flow.     

Effects of trinucleotides on hormonally responsive adenylate cyclase activity in rat kidney

The effects of trinucleotides on basal and hormonally stimulated adenylate cyclase activity in rat kidney homogenates was evaluated. In the absence of added trinucleotides, parathyroid hormone (PTH) increased cortical adenylate cyclase activity, while antidiuretic hormone (ADH) increased medullary activity; however, prostaglandin (PG)E₂ did not stimulate adenylate cyclase activity in the renal cortex, outer medulla, or inner medulla. However, with exogenous guanosine 5′-triphosphate (GTP) at concentrations as low as 8 × 10^?7M, PGE₂ did activate adenylate cyclase. This effect was seen over a concentration range of 8 × 10^?7 to 8 × 10^?4M PGE₂. In addition, deoxyGTP and cytosine 5′-triphosphate (CTP) were also effective in the cortex, and deoxyGTP was effective in the inner medulla. GTP also augmented PTH stimulation slightly in the cortex, but had no effect on ADH stimulation in medullary tissue. Therefore, a diference in the GTP requirement was observed for prostaglandins compared to polypeptide hormones. When stimulation by PTH or ADH was examined over a wide concentration range, no dependency upon GTP for stimulation was observed. Furthermore, when the ATP concentration was reduced from 2 to 0.2 mM and Mg⁺⁺ concentration was varied from 1 to 5 mM, a dependency of either PTH or ADH stimulation upon GTP was still not observed. Guanylyl 5′-imidodiphosphate (8 μM) elevated both basal and polypeptide hormone activity in all three tissues but did not elevate PGE₂ above basal values for that group. GTP appears to be important in renal regulation of PGE₂-stimulated adenylate cyclase activity.     

The metabolism of arachidonic acid in the isolated tracheal and lung strip preparations of guinea-pigs

The effect of arachidonic acid (AA) on tension in guinea-pig isolated lung strips and tracheas was investigated. AA (10^?5 M) caused relaxation in the trachea and contraction in the lung strip. In lung strips the AA-induced contractions were inhibited by indomethacin (10^?5 M), 5, 8, 11, 14-eicosatetraynoic acid (5 × 10^?5 M) and L 8027 (5 × 10^?7 M) but not by imidazole (up to 2 × 10^?3 M). Material which co-chromatographed with thromboxane B₂ (TXB₂) was shown to be released from lung strips challengend with¹⁴CAA. L 8027 abolished TXB₂ production whereas imidazole did not. These results indicate that AA is metabolised differently in central and peripheral airways. In the trachea prostaglandin E₂ may mediate relaxation, whereas in lung strips material which co-chromatographs with TXB₂ is associated with a contractile response to AA.     

Short-term angiotensin converting enzyme inhibition reduces basal tone and dilator reactivity in skeletal muscle arterioles

Alterations in resting tone, maximum diameter, and dilator reactivity to acetylcholine (ACH) and sodium nitroprusside (SNP) were assessed in cremaster muscle microvessels of Sprague-Dawley rats receiving angiotensin converting enzyme (ACE) inhibition with captopril for 4 days and in untreated time-control rats. The transilluminated in situ cremaster muscle was superfused with physiologic salt solution (PSS) and viewed via television microscopy; arteriolar diameter was measured using a videomicrometer. Before agonist challenge, resting arteriolar diameter was significantly increased in captopril-treated rats. Although maximum arteriolar diameter (determined during superfusion of the cremaster muscle with Ca²⁺-free PSS containing 10⁻⁴ mol/L adenosine) was not altered with ACE inhibition, the maximum possible arteriolar dilation was reduced in captopril-treated rats. Captopril administration reduced both ACH- and SNP-induced dilation of cremasteric arterioles compared with responses in control rats, although this was partially a function of the reduced capacity for dilation, primarily to SNP. These observations indicate that short-term ACE inhibition reduces both resting tone and agonist-induced dilator responses of skeletal muscle arterioles.     

Antioxidant inhibition of prostaglandin production by rat renal medulla.

Antioxidant effects upon renal production of both prostaglandins and cAMP were investigated using slices of rat inner medulla. Synthetic antioxidants were more potent inhibitors of prostaglandin production than were naturally occurring antioxidants. Synthetic compounds 2,7-naphthalenediol, and Santoquin^® (Ethoxyquin) caused a 60% inhibition of prostaglandin E₂(PGE₂) synthesis at a concentration of 0.01 mM. Ascorbic acid caused only a 30% inhibition at a concentration of 10 mM. Antioxidant inhibition of prostaglandin production was also observed following arachidonic acid addition. Antioxidants that reduced PGE₂ synthesis also reduced PGF_2α synthesis. Test agents found to reduce prostaglandin synthesis also lowered cAMP content. 2,7-Naphthalenediol elicited a dose-dependent decrease in both prostaglandin synthesis and cAMP content. While PGE₁ did not increase cAMP in control slices, the low cAMP level produced by Santoquin was increased to control values by PGE₁. Furthermore, Santoquin and 2,7-naphthalenediol did not alter arginine vasopressin-stimulated cAMP content. By contrast, inhibition of the arginine vasopressin stimulation by butylated hydroxyanisole suggested additional effects by this agent. These results are consistent with the hypothesis that endogenously produced PGE₂ can exert a hormonelike action in the inner medulla by increasing cAMP content. Advantages of the inner medullary slice system compared to homogenates for investigation of the actions of antioxidants or other agents thought to alter prostaglandin synthesis are discussed.     

Arachidonic acid metabolites and glucocorticoid regulatory mechanism in cultured porcine tracheal smooth muscle cells

To elucidate the signal transduction system in the production of prostaglandin E₂ (PGE₂) by porcine tracheal smooth muscle cells in culture (PTSMC), we examined the pattern of arachidonic acid metabolites released from PTSMC and the relationship between bradykinin-stimulated rises in intracellular calcium concentration ([Ca²⁺]_i) and PGE₂ production by PTSMC. We next examined the effect of dexamethasone on these parameters. Bradykinin induced a dose-dependent increase in both the rise in [Ca²⁺]_i and PGE₂ production by PTSMC. The increase in [Ca²⁺]_i paralleled an increase in PGE₂ production. High-performance liquid chromatography (HPLC) revealed that dexamethasone-treated PTSMC were suppressed to release arachidonic acid metabolites such as PGE₂ and prostaglandin F₂ (PGF₂). Incubation of PTSMC with 10^–6M dexamethasone for 24 h significantly suppressed both the rise in [Ca²⁺]_i and PGE₂ production by PTSMC in response to bradykinin, and also significantly suppressed bradykinin-stimulated release of radioactivity from PTSMC prelabeled with ³H-labeled arachidonic acid (³H-AA). When PTSMC pretreated with dexamethasone were incubated with 170 nM prostaglandin H₂ (PGH₂) or 20 M arachidonic acid; PTSMC synthesized less PGE₂ than control PTSMC. Results suggest that bradykinin stimulates PTSMC to produce PGE₂ via the signal transduction system including Ca²⁺, and dexamethasone appeared to suppress PGE₂ production by reducing the activity of cytosolic phospholipase A₂ (cPLA₂) and PGE₂ synthase. However, we failed to demonstrate the suppression of the activity of cyclooxygenase in PTSMC by dexamethasone. Since the elevation of [Ca²⁺]_i is necessary for the contraction of airway smooth muscles, dexamethasone seems to reduce the contraction of airway.Offprint requests to: Hideki Tanaka     

Stimulatory and inhibitory effects of prostaglandin E2 on prolactin release in the domestic fowl

In vivo prolactin secretion was increased in immature cockerels 20–30 min after the intravenous administration of prostaglandin (PG) E₂ at a dose of 200 μg/kg. The addition of PGE₂ to incubation medium had no direct effect on the release of pituitary prolactin during short-term (3-hr) culture, but augmented the stimulatory effect of hypothalamic tissue on prolactin secretion. The stimulatory effect of serotonin, noradrenaline, acetylcholine, and histamine on hypothalamus-induced prolactin release was also increased when pituitaries were coincubated with 10^?7M PGE₂, as was the stimulatory effect of thyrotrophin-releasing hormone (TRH) and hypothalamic extract (HE). The long-term (24-hr) preincubation of pituitaries with 10^?7M PGE₂ reduced the responsiveness of the prolactin-secreting cells to TRH or HE stimulation. PGE₂ treatment also reduced the stimulatory effect of hypothalamic tissue on prolactin release and diminished the stimulatory effect of serotonin on hypothalamus-induced prolactin secretion. A 24-hr preincubation of hypothalamic tissue with 10^?7M PGE₂ also reduced its stimulatory effect on prolactin release when subsequently incubated with control pituitary glands. These results demonstrate that PGE₂ initially stimulates in vivo and in vitro prolactin secretion in the fowl, possibly by increasing the release of hypothalamic prolactin-releasing activity and/or by increasing pituitary sensitivity to provocative stimuli. Chronic PGE₂ stimulation appears to result in a reduction in pituitary responsiveness to stimulatory influences and in the release of hypothalamic-releasing activity.     

Prostaglandins mediate arteriolar dilation to increased blood flow velocity in skeletal muscle microcirculation

In cremaster muscle of pentobarbital-anesthetized rats, temporary occlusion of an arteriole increased red blood cell velocity (mean increase, 8.2 +/- 1.0 mm/sec from a control velocity of 7.9 +/- 0.7 mm/sec) in proximal parallel arteriolar branches (mean control diameter, 19.4 +/- 0.6 microns). Increases in flow velocity were consistently followed by proportional delayed (6-15 seconds) increases in arteriolar diameter (5.8 +/- 0.7 microns). Administration of NG-monomethyl-L-arginine (200 microM), an inhibitor of the synthesis of endothelium-derived relaxing factor that blocked the arteriolar responses to acetylcholine (1 microM) but not to arachidonic acid (10 microM), did not affect the dilation (mean increase, 8.9 +/- 1.1 microns) due to increases in red blood cell velocity (13.4 +/- 1.5 mm/sec). However, the cyclooxygenase inhibitor indomethacin (or meclofenamate), which completely blocked the dilator response to arachidonic acid but did not change the response to acetylcholine, inhibited the arteriolar dilation (mean increase, 0.3 +/- 0.2 micron) due to increases in red blood cell velocity (9.3 +/- 1.0 mm/sec). Inhibition of prostaglandin synthesis also reduced the increase in calculated blood flow by 57% during occlusion. These results suggest that the arterioles are sensitive to increases in blood flow velocity (wall shear stress), in response to which they release prostaglandins, eliciting vasodilation. The existence of this phenomenon in the skeletal muscle microcirculation suggests a new regulatory mechanism that, by modulation of vascular resistance in the microvascular network, has the role of normalizing wall shear stress and providing for substantial increases in tissue blood flow.     

Relaxation effect of marmin on guinea pig tracheal smooth muscle via NO-independent mechanisms

ObjectiveTo investigate the relaxation mechanims of marmin on epithelium of guinea pig isolated trachea smooth muscle (TSM).MethodsThe study was conducted using in vitro isolated-trachea experimental. The guinea pig isolated trachea were incubated in Krebs solution-containing organ bath and supplied with a mixed gas of O₂: CO₂ (95%:5%).ResultRemoval of tracheal epithelium was associated with significant increases in the potencies of histamine and methacholine to contract guinea pig TSM. The pD₂ value of histamine increased from 6.04±0.08 on epithelial-intact to 6.32±0.06 on epithelial-denuded (P < 0.05). The pD₂ value of methacholine also increased from 5.85±0.09 on epithelial-intact to 6.15±0.07 on epithelial-denuded (P < 0.05). Marmin exhibited relaxation effects on TSM induced by methacholine (3×10^?5 mol/L) and histamine (3×10^?5 mol/L). Inhibition of prostaglandin E2 (PGE₂) through incubation with indomethacin could reduce the relaxation effect of marmin (P < 0.05) on methacholine- and histamine-induced contractions. However, no significant differenceswere shown in methylene blue, Nω-nitro-L-arginine (L-NNA) and propranolol-incubated TSM.ConclusionsThe results suggest that marmin has relaxation effect on TSM which is epithelial-dependent through the release of PGE₂. However, nitric oxide, cGMP and β2-adrenergic-mediated relaxation were not involved.     

Prostaglandin synthetase activity in bovine adrenocortical cells and subcellular preparations: Effect of ACTH

Prostaglandin (PGE₂, PGF_2α) production by bovine fasciculo-reticulata adrenocortical cell suspensions was examined using specific radioimmunoassay procedures. No detectable PGs (> 50 pg) could be measured in the extract from up to 2 × 10⁶ cell incubations after 1 h, with or without the presence of ACTH, although these cells expressed full steroidogenic capabilities under these conditions. The same preparations could produce PGs when supplemented with arachidonic acid but ACTH had no effect on this process. These negative findings could not be explained by analytical artifacts or metabolic transformation. However, an active PG synthetase system was characterized in bovine adrenocortical subcellular preparations. This system converted arachidonate and endogenous substrate(s) to PGE₂ as the major product. No thromboxane or prostacyclin pathways were detected even at high enzyme/substrate ratio. Although the microsornal adrenal cortex PG synthetase activity shares many features with those observed in other tissues (K_m = 8.3 × 10^?5 M, optimal pH at 8.0, stimulation of PGE₂ formation in the presence of glutathione and L-epinphrine), its specific activity was comparatively low (V_max = 2.5 ng PGE₂/min/mg microsornal proteins), which may explain our negative findings using cell suspensions. These findings do not provide evidence to support the hypothesis proposing a role of endogenous PGs in the mechanism of acute ACTH action in the case of bovine adrenal cortex.     

Increased arachidonic acid-induced thromboxane generation impairs skeletal muscle arteriolar dilation with genetic dyslipidemia

Objective: The aim of this study was to determine if arachidonic acid (AA)‐induced skeletal muscle arteriolar dilation is altered with hypercholesterolemia in ApoE and low‐density lipoprotein receptor (LDLR) gene deletion mice fed a normal diet. This study also determined contributors to altered AA‐induced dilation between dyslipidemic mice and controls, C57/Bl/6J (C57). Methods: Gracilis muscle arterioles were isolated, with mechanical responses assessed following a challenge with AA under control conditions and after elements of AA metabolism pathways were inhibited. Conduit arteries from each strain were used to assess AA‐induced production of PGI₂ and TxA₂. Results: Arterioles from ApoE and LDLR exhibited a blunted dilation to AA versus C57. While responses were cyclo‐oxygenase‐dependent in all strains, inhibition of thromboxane synthase or blockade of PGH₂/TxA₂ receptors improved dilation in ApoE and LDLR only. AA‐induced generation of PGI₂ was comparable across strains, although TxA₂ generation was increased in ApoE and LDLR. Arteriolar reactivity to PGI₂ and TxA₂ was comparable across strains. Treatment with TEMPOL improved dilation and reduced TxA₂ production with AA in ApoE and LDLR. Conclusions: These results suggest that AA‐induced arteriolar dilation is constrained in ApoE and LDLR via an increased production of TxA₂. While partially due to elevated oxidant stress, additional mechanisms contribute that are independent of acute alterations in oxidant tone.     

The in vitro response of the trout interrenal to various fragments of ACTH

The ability of various fragments of ACTH to stimulate cortisol secretion was tested in vitro using diced interrenal tissue from the rainbow trout Salmo gairdneri. 1–24ACTH induced maximum steroidogenesis at a concentration of 5 × 10^?8M, and a half-maximal response at 0.8 × 10^?9M. Des-acetyl-αMSH (1–13NH₂ACTH) had full intrinsic activity; its molar potency was 0.9 × 10^?2-fold less than 1–24ACTH but nearly 100-fold greater than acetylated αMSH. 1–10ACTH had an even lower potency, while 4–10ACTH was without effect at the highest concentration tested (5 × 10^?5M). The fragment 1–16ACTH was unusual in eliciting a higher maximum cortisol concentration than 1–24ACTH. These findings suggest that the increase in plasma cortisol, previously observed in trout adapted to a black background, are unlikely to be directly attributable to adrenal stimulation by the raised MSH titres.     

Simultaneous measurements of macromolecular leakage and arteriolar blood flow as altered by PGE1 and beta 2-receptor stimulant in the hamster cheek pouch.

Alterations in arteriolar blood flow and macromolecular leakage after prostaglandin (PGE₁) administration in the presence and absence of a β-receptor stimulant (terbutaline) were investigated in the hamster cheek pouch preparation by direct observation. After iv injection of FITC-dextran ($\text{M}{}_{\mathrm{<mtext>w</mtext>}}\text{= 150,000}$; 25 mg 100 g⁻¹ body wt), macromolecular leakage was quantified as the number of leakage sites per square centimeter appearing in the cheek pouch upon topical application of PGE₁ (0.1, 1.0, 10.0, and 100.0 ng ml⁻¹) in the presence and in the absence of terbutaline (1.0 μg ml⁻¹) in the superfusion solution. Concomitantly, arteriolar blood flow was calculated from measurements of red blood cell velocity and vessel diameter. PGE₁ produced a dose-dependent increase in macromolecular leakage during normal superfusion while this effect was reduced significantly when terbutaline was present. PGE₁ produced an increase in arteriolar blood flow which was not affected by terbutaline. Terbutaline, alone, reduced macromolecular leakage and increased arteriolar blood flow. Thus, it is concluded that terbutaline counteracts PGE₁-induced macromolecular leakage. This effect is not mediated through a reduction of the PGE₁-induced increase in arteriolar blood flow.     

A Role for Dietary Copper in Nitric Oxide-Mediated Vasodilation

Objective : This study was designed to investigate the role of dietary copper in nitric oxide-mediated arteriolar dilation. Methods : Male weanling Sprague—Dawley rats were fed a purified diet that was either copper-adequate (6.0 μg Cu per g diet) or copper-deficient (0.3 μg Cu per g diet) for a period of 4 weeks. Each rat was anesthetized with pentobarbital and its cremaster muscle was positioned in a Krebs'-fdled bath to which graded concentrations of vasoactive agents were added. In the first series, responses to norepineph-rine (NE 10^?9-10^?6 M) and acetylcholine (ACH 10^?7-10^?4 M) were compared in third-order arterioles. Second, the dilator response to 10^?5 M ACH in the absence and presence of 240 U/ml Cu, Zn-superoxide dismutase (SOD) was determined. Third, arteriolar dilation was determined in response to NO-independent stimulation of soluble guanylate cyclase with hydrogen peroxide (10^?7-10^?5 M) and to dibutyryl cGMP (10^?6-10^?4 M), dibutyryl cAMP (10^?6-10^?4 M), and papaverine (10^?4M). Results : The arteriole constrictor response to NE and the dilator response to hydrogen peroxide, dibutyryl cGMP and cAMP, and papaverine were not different between the dietary groups. Copper deficiency attenuated the ACH-induced dilation, but the response was restored in the presence of SOD. Conclusions : The inactivation of cytosolic Cu, Zn-SOD by restriction of dietary copper results in the depression of NO-mediated vascular smooth-muscle relaxation probably by interaction of NO with superoxide.     

Characterization of pancreatic islet monoamine oxidase

Monoamine oxidase (MAO) is present in isolated islets of Langerhans of rabbits, golden hamsters, and rats. Tryptamine, tyramine, serotonin, and dopamine can serve as substrates for this enzyme. We compared the properties of islet and liver MAO in the rabbit. The Michaelis constant (K_m) for tryptamine of islet MAO (6.5 × 10^?5M) is greater than the K_m of liver MAO (3 × 10^?5M). The K_m for tyramine of islet MAO (1.5 × 10^?4M) is similar to the K_m of liver MAO (1.8 × 10^?4M). Islet MAO appeared to be more susceptible to heat inactivation (50°C) than did liver MAO. This may be an artifact produced by the collagenase technique used in the preparation of the islets, as collagenase treatment of liver increased the thermal lability of the MAO in this tissue. Liver and islet MAO have a comparable sensitivity to MAO inhibitors such as clorgyline, deprenyl, tranylcypromine, pargyline, and harmine. The present report, along with previous reports that MAO inhibitors alter insulin secretion, suggests that islet MAO may modify insulin secretion.     

Microsomal prostaglandin E synthase 1 determines tumor growth in vivo of prostate and lung cancer cells

There is strong evidence for a role of prostaglandin E₂ (PGE₂) in cancer cell proliferation and tumor development. In PGE₂ biosynthesis, cyclooxygenases (COX-1/COX-2) convert arachidonic acid to PGH₂, which can be isomerized to PGE₂ by microsomal PGE-synthase-1 (MPGES-1). The human prostate cancer cell line DU145 expressed high amounts of MPGES-1 in a constitutive manner. MPGES-1 expression also was detectable in human prostate cancer tissues, where it appeared more abundant compared with benign hyperplasia. By using shRNA, we established stable and practically complete knockdown of MPGES-1, both in DU145 cells with high constitutive expression and in the non-small cell lung cancer cell line A549, where MPGES-1 is inducible. For microsomes prepared from knockdown clones, conversion of PGH₂ to PGE₂ was reduced by 85–90%. This resulted in clear phenotypic changes: MPGES-1 knockdown conferred decreased clonogenic capacity and slower growth of xenograft tumors (with disintegrated tissue structure) in nude mice. For DU145 cells, MPGES-1 knockdown gave increased apoptosis in response to genotoxic stress (adriamycin), which could be rescued by exogenous PGE₂. The results suggest that MPGES-1 is an alternative therapeutic target in cancer cells expressing this enzyme.     

A densitometric method for determining the filtration coefficients of single capillaries in the frog mesentery.

Single frog mesenteric capillaries were perfused with T1824-albumin solution via a micropipette and occluded some way downstream. A step change in pressure was applied through the micropipette to the closed-off section of vessel. The consequent change in filtration rate was measured from the change in optical density, caused by concentration of dye in the capillary. The slope of the relationship between filtration rate and pressure gave the filtration coefficient of the capillary. For capillaries which retained T1824-albumin during the course of the determination (45–60 sec), filtration coefficients at 20° ranged from 4 × 10^?3 to 12 × 10^?3 μm/sec cm H₂O; where T1824-albumin leaked across the capillary wall, filtration coefficients were three times higher (21–33 × 10^?3 μm/sec cm H₂O). When filtration occurred from a closed-off capillary at constant pressure, the difference between the intracapillary hydrostatic pressure and the intracapillary protein osmotic pressure declined exponentially with time. Values of L_p calculated from the time constant agreed with values determined in the same vessels from changes in intracapillary pressure.     

Early changes in the arachidonic acid metabolism of HeLa cells in response to the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) and related compounds

Summary In order to search for possible mediators involved in the transient radiomimetic effectiveness of TPA and related compounds early changes in the AA metabolism of HeLa cells prelabeled with 1-¹⁴C-AA have been analyzed. Maximum release of AA with different concentrations of TPA (3×10^-9 to 3×10^-5 M) was observed after 2–3 h treatment in the presence of 10% calf serum. Released AA was reincorporated by the cells after that period, a phenomenon which was largely abolished or delayed by cycloheximide. Reincorporation of released AA was observed in the presence of 10% fresh serum as well as with 0.5% BSA, and appears to be due to an induction of responsible enzyme(s) by the phorbol ester. The earliest metabolites of AA produced via the cyclooxygenase such as PGE₂ and PGF₂ and via lipoxygenases such as 12-, and 15-hydroxyeicosatetraenoic acids appear in small amounts and after later time points. AA release exhibited a pluriphasic dose response to TPA with maxima at 3×10^-8 M and 10^-5 M. Comparative dose response measurements with respect to AA release were established using various promoting skin mitogens which exhibited the following order of potency: TPA > teleocidin RPA > mezerein EPA>4-O-Me-TPA. For reasons discussed it appears unlikely that AA, Prostaglandins, or hydroxyeicosatetraenoic acid products play a significant role as mediators of the radiomimetic effects of TPA in G₂ of the cell cycle.Abbreviations AA arachidonic acid - BSA bovine serum albumin - EPA 12-O-ethacrynylphorbol-13-acetate - 4-O-Me-TPA 4-O-methyl-TPA - PGE prostaglandin E₂ - RPA 12-O-retinoylphorbol-13-acetate - TPA 12-O-tetradecanoylphorbol-13-acetate Dedicated to Professor Dr. E. Hecker on the occasion of his 60th birthday