calcimycin and zaprinast

Congenital heart disease associated with increased pulmonary blood flow produces pulmonary hypertension. To characterize vascular alterations in the nitric oxide (NO)-cGMP cascade induced by increased pulmonary blood flow and pulmonary hypertension, 10 fetal lambs underwent in utero placement of an aortopulmonary vascular graft (shunt). When the lambs were 4-6 wk of age, we assessed responses of pulmonary arteries (PAs) and pulmonary veins (PVs) isolated from lungs of control and shunted lambs. PVs from control and shunted lambs relaxed similarly to exogenous NO (S-nitrosyl-acetyl-penicillamine), to NO produced endogenously (zaprinast and A-23187), and to cGMP (atrial natriuretic peptide). In contrast, relaxations to A-23187 and zaprinast were blunted in PAs isolated from shunted lambs relative to controls. Inhibitors of NO synthase (NOS) and soluble guanylate cyclase constricted control but not shunt PAs, indicating reduced basal NOS activity in shunt PAs. Pretreatment of shunt PAs with the substrates L-arginine and sepiapterin, a precursor for tetrahydrobiopterin synthesis, did not augment A-23187 relaxations. However, pretreatment with superoxide dismutase and catalase significantly enhanced A-23187 relaxations in shunt PAs. We conclude that increased pulmonary blood flow induces an impairment of endothelium-dependent relaxation that is selective to PAs. The impaired relaxation may be mediated in part by excess superoxide production.

Topics: 3',5'-Cyclic-GMP Phosphodiesterases; Animals; Atrial Natriuretic Factor; Calcimycin; Disease Models, Animal; Endothelium, Vascular; Enzyme Inhibitors; Fetus; Guanylate Cyclase; Hypertension, Pulmonary; In Vitro Techniques; Muscle Relaxation; Muscle, Smooth, Vascular; Nitric Oxide Donors; Nitric Oxide Synthase; Nitric Oxide Synthase Type III; Phosphodiesterase Inhibitors; Pulmonary Artery; Pulmonary Circulation; Pulmonary Veins; Purinones; Sheep

N-acetyl-L-cysteine, a thiol compound, has been shown to potentiate the inhibition of platelet aggregation exerted by organic nitrates and to increase the anti-aggregating effect of L-arginine, which promotes endogenous synthesis of nitric oxide (NO) acting as substrate of platelet constitutive nitric oxide synthase (NOS). It is not known whether this thiol can exert direct effects on platelet aggregability.. 14 healthy male volunteers provided platelet samples to investigate whether N-acetyl-L-cysteine directly influences platelet function and intraplatelet levels of 3',5' cyclic guanosine monophosphate (cGMP), which represents the second messenger involved in NO-induced antiaggregation. Some experiments were repeated in the presence of NOS inhibitor NG-monomethyl-L-arginine (L-NMMA), of nitric oxide-sensitive guanylyl cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ), of the selective cGMP phosphodiesterase inhibitor zaprinast and of calcium ionophores (A23187, ionomycin).. N-acetyl-L-cysteine at 3000-6000 micromol L-1 decreases the responses of human platelets both in platelet-rich plasma (aggregation induced by adenosine 5-diphosphate) and in whole blood (aggregation induced by collagen). The anti-aggregating effect was prevented by preincubation with L-NMMA and guanylyl cyclase inhibitor ODQ. In resting platelets, N-acetyl-L-cysteine increased the levels of cGMP starting from a concentration of 3000 micromol L-1. Permeabilized platelets exhibited an increased sensitivity to the anti-aggregating effect of N-acetyl-L-cysteine. Also, cGMP phosphodiesterase inhibition or the increase in calcium availability, enhanced N-acetyl-L-cysteine effects on platelets.. N-acetyl-L-cysteine exerts direct anti-aggregating effects through an increased bioavailability of platelet nitric oxide.

Topics: 3',5'-Cyclic-GMP Phosphodiesterases; Acetylcysteine; Adenosine Diphosphate; Adult; Calcimycin; Cell Membrane Permeability; Collagen; Cyclic GMP; Dose-Response Relationship, Drug; Enzyme Inhibitors; Guanylate Cyclase; Humans; Ionomycin; Male; omega-N-Methylarginine; Oxadiazoles; Platelet Aggregation; Platelet Aggregation Inhibitors; Purinones; Quinoxalines

The management of sick newborn infants who have sustained a hypoxic insult is a common clinical problem but relatively little is known about the recovery process. The aim of this study was to investigate this process in newborn piglets.. Thirty five newborn piglets were exposed to chronic hypobaric hypoxia for three days, either from birth, three or 14 days of age, and were allowed to recover for one, three, or six days. Control animals of relevant age were also studied. The heart weight ratio and pulmonary arterial muscularity were measured. Endothelial dependent and independent relaxation of the isolated intrapulmonary conduit arteries was determined in classical organ chamber studies, together with measurement of basal and stimulated cGMP accumulation.. After six days of recovery the hypoxia induced right ventricular hypertrophy and pulmonary arterial medial hypertrophy had decreased in all animals but values were still abnormal in the two younger age groups. Relaxation was still impaired during the first three days of recovery in all groups, had normalised by six days in the two youngest groups, but relaxation (both endothelium dependent and independent) remained impaired in older animals. In these older animals basal nitric oxide (NO) production and basal and stimulated cGMP accumulation was normal.. The recovery of the smooth muscle cells lags behind that of the endothelial cells. A normal stimulated increase in cGMP with reduced relaxation suggests an altered threshold for cGMP effected relaxation. These findings help to explain why some hypoxic infants require protracted NO therapy.

Topics: Acetylcholine; Animals; Animals, Newborn; Calcimycin; Cyclic GMP; Enzyme Inhibitors; Hypertension, Pulmonary; Hypertrophy; Hypertrophy, Right Ventricular; Hypoxia; Nitric Oxide; omega-N-Methylarginine; Phosphodiesterase Inhibitors; Pulmonary Artery; Purinones; Swine; Tunica Intima; Vasoconstriction; Vasodilator Agents

Pulmonary hypertension plays a significant role in the pathophysiology of congenital diaphragmatic hernia (CDH). Although there has been an intensive research effort directed at mediators that may cause pulmonary vasoconstriction, no single agent has been identified. The authors hypothesize that there may be an alteration in the cGMP-nitric oxide (NO) pathway of vasodilatation contributing to the pulmonary hypertension observed in CDH. The purpose of these studies is to begin to elucidate vasoactive properties of pulmonary vessels with particular attention to the cGMP-NO pathway of vasodilatation in fetal lambs with CDH.. Fourth-generation pulmonary arteries and pulmonary veins were dissected from both right and left lungs of eight, 139-day gestational fetuses with surgically created CDH. Vessels were studied with standard isolated tissue bath techniques. Experiments examined basal release of NO in endothelium-intact PVs and PAs of both right and left lungs by measuring the contractile force of vessels constricted with norepinephrine (NE) in the presence and absence of the nitric oxide synthase (NOS) inhibitor N(omega)-nitro-L-arginine (L-NA). Concentration-response curves to the vasodilating agents zaprinast and A23187 were also obtained in vessels contracted by NE.. Left and right pulmonary artery responses to NE are enhanced over those of historic controls. Pretreatment of left pulmonary arteries with L-NA enhances the vasoconstrictor response to NE, whereas right PAs show no increased response. Relaxation responses to A23187 and zaprinast, in both left and right pulmonary arteries were not different from control lambs. Relaxation responses of both left and right pulmonary veins to A23187 and zaprinast are blunted compared with controls. This blunting is significantly more in left pulmonary veins than right. Further, right but not left pulmonary veins display enhanced vasoconstrictive response to NE after L-NA pretreatment.. The NO-cGMP pathway of vasodilatation is abnormal in the near term, fetal lamb with CDH. These abnormalities were most apparent in pulmonary veins and may reflect abnormal NOS activity or content between left and right lungs of the fetal lamb with CDH. Pulmonary arteries from CDH lambs have basal and stimulated NO release equal to that of historic controls but appear to be hypersensitive to exogenous vasoconstrictors.

Topics: Animals; Calcimycin; Cyclic GMP; Disease Models, Animal; Dose-Response Relationship, Drug; Fetus; Hernia, Diaphragmatic; Ionophores; Nitric Oxide; Phosphodiesterase Inhibitors; Pulmonary Artery; Pulmonary Veins; Purinones; Sheep; Vasoconstriction; Vasodilation

Neonatal pulmonary hypertension is associated with increased pulmonary vascular reactivity. We studied the responses of isolated porcine intrapulmonary arteries after exposure of piglets to chronic hypobaric hypoxia (CHH) from 0 to 2.5, 3 to 6, or 14 to 17 days of age. CHH inhibited the postnatal development of endothelium-dependent vasorelaxation to acetylcholine (ACh) and the calcium ionophore A-23187. Basal accumulation of guanosine 3', 5'-cyclic monophosphate (cGMP) was unaffected, but cGMP response to ACh was inhibited. Endothelium-independent relaxation to nitric oxide (NO) and zaprinast (a phosphodiesterase inhibitor) was also inhibited, but cGMP accumulation in response to these agonists was normal. The ability of sodium nitroprusside (SNP) to cause vasorelaxation and increase cGMP accumulation was unaffected. Contractile responses to potassium chloride and prostaglandin F2 alpha (PGF2 alpha) were similar to normal after exposure from birth and 3 days and were decreased in the older group, but the ability of NG-monomethyl-L-arginine acetate to increase PGF2 alpha-induced contractions decreased. Thus exposure of newborn piglets to CHH causes 1) no increase in contractile responses and 2) impairment of endothelium-dependent and -independent relaxation by impairing signal transduction mechanisms involved in the release of NO and the effectiveness of cGMP.

Topics: Acetylcholine; Animals; Animals, Newborn; Atmospheric Pressure; Calcimycin; Chronic Disease; Cyclic GMP; Dinoprost; Endothelium, Vascular; Heart; Hypertension, Pulmonary; Hypoxia; In Vitro Techniques; Nitric Oxide; omega-N-Methylarginine; Potassium Chloride; Pulmonary Artery; Purinones; Swine; Vasodilation; Vasomotor System

Guanylate cyclase activity in human erythrocytes is investigated by evaluating the intracellular guanosine 3',5'-cyclic monophosphate (cGMP) content in the presence of various agents that exert specific effects on soluble or particulate guanylate cyclase. The increase in the intraerythrocyte cGMP content by the soluble guanylate cyclase activators nitroprusside and NaNO2 suggests the presence of this enzyme in human erythrocytes. The effects of four different atrial natriuretic peptide (ANP) fragments on the intraerythrocyte cGMP content is also studied. ANP II and ANP III increase the intraerythrocyte cGMP content, whereas ANP I and des-Ser5,des-Ser6-ANP III are ineffective. Thus our data show that human erythrocytes possess particulate guanylate cyclase together with the soluble enzyme. The ANP fragments ANP II and ANP III also activate the erythrocyte Na+/H+ exchange. Nitroprusside, M & B 22948 (an inhibitor of cGMP phosphodiesterase), and the cGMP analogues dibutyryl cGMP and 8-bromoguanosine 3',5'-cyclic monophosphate also increase the erythrocyte Na+/H+ exchange rate. The latter data also suggest that the erythrocyte Na+/H+ exchange is regulated by cGMP.

Topics: 3',5'-Cyclic-GMP Phosphodiesterases; 8-Bromo Cyclic Adenosine Monophosphate; Adenosine Triphosphate; Atrial Natriuretic Factor; Calcimycin; Cyclic GMP; Enzyme Activation; Erythrocytes; Guanylate Cyclase; Hemoglobins; Homeostasis; Humans; Kinetics; Manganese; Methemoglobin; Nitroprusside; Peptide Fragments; Phosphodiesterase Inhibitors; Purinones; Sodium Nitrite; Sodium-Hydrogen Exchangers

The antiinflammatory activity of rolipram, a selective inhibitor of the cyclic AMP-specific phosphodiesterase (PDE IV), was studied. Rolipram did not inhibit 5-lipoxygenase activity but did inhibit human monocyte production of leukotriene B4 (LTB4, IC50 3.5 microM). Likewise, murine mast cell release of leukotriene C4 and histamine was inhibited. In vivo, rolipram inhibited arachidonic acid-induced inflammation in the mouse, while the low Km-cyclic-GMP PDE inhibitor, zaprinast, did not inhibit. Rolipram had a modest effect on LTB4 production in the mouse, but markedly reduced LTB4-induced PMN infiltration. Beta-adrenergic receptor activation of adenylate cyclase was important for rolipram antiinflammatory activity since beta blockade abrogated arachidonic acid-induced inflammation. Thus, the antiinflammatory profile of rolipram is novel and may result from inhibition of PMN function and perhaps vasoactive amine release and leukotriene biosynthesis. These actions may be dependent upon endogenous beta-adrenergic activity and are likely mediated through inhibition of PDE IV.

Topics: 3',5'-Cyclic-AMP Phosphodiesterases; Animals; Anti-Inflammatory Agents, Non-Steroidal; Arachidonic Acid; Calcimycin; Cells, Cultured; Colforsin; Cyclic Nucleotide Phosphodiesterases, Type 4; Ear, External; Eicosanoids; Histamine Release; Humans; Imidazoles; Inflammation; Leukotriene B4; Male; Mast Cells; Mice; Mice, Inbred BALB C; Monocytes; Nadolol; Naproxen; Neutrophils; Phosphodiesterase Inhibitors; Phosphoric Diester Hydrolases; Purinones; Pyrazoles; Pyrrolidinones; Receptors, Adrenergic, beta; Rolipram; SRS-A; Thiazoles

The present study was designed to investigate whether cyclic GMP or cyclic AMP modulates the release of endothelium-derived relaxing factor/nitric oxide (EDRF/NO) in cultured bovine aortic endothelial cells (BAE cells). BAE cell-conditioned medium was transferred onto rat fetal lung fibroblasts (RFL-6 cells) and the increase in cyclic GMP in these cells was used as a sensitive bioassay of EDRF/NO activity. BAE cells released a material that markedly enhanced cyclic GMP in RFL-6 cells. The synthesis of this substance could be stimulated with bradykinin (10 nM) or Ca++ ionophore A23187 (1 microM) and was completely prevented by treatment of the BAE cells with the EDRF/NO synthesis inhibitors NG-nitro-L-arginine (100 microM) or NG-methyl-L-arginine (1 mM). Addition of hemoglobin (10 microM) or incubation of the RFL-6 detector cells with methylene blue (10 microM) also abolished the cyclic GMP increase in the RFL-6 cells. The release of EDRF/NO by bradykinin and A23187 was accompanied by an approximately 2-fold increase in the cyclic GMP content in the producing BAE cells (in the presence of the cyclic GMP phosphodiesterase inhibitor M&B 22,948, 0.1 mM). Incubation of BAE cells with atrial natriuretic peptide (0.1 microM) or sodium nitroprusside (10 microM) enhanced cyclic GMP content of BAE cells 6.5-fold and 4.1-fold, respectively (in the presence of M&B 22,948, 0.1 mM). These increases in the cyclic GMP levels in BAE cells had no effect on basal or bradykinin- and A23187-stimulated release of EDRF/NO. Bradykinin (10 nM) and A23187 (1 microM) also stimulated prostacyclin production in BAE cells 2.4-fold and 5.6-fold, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Aorta; Arginine; Calcimycin; Cattle; Cells, Cultured; Cyclic AMP; Cyclic GMP; Endothelium, Vascular; Epoprostenol; Nitric Oxide; Nitroarginine; Purinones

Modulation of the human polymorphonuclear leukocyte (PMN) respiratory burst by selective cyclic 3',5' adenosine monophosphate (cAMP) phosphodiesterase (PDE) inhibitors was studied with respect to PDE isozyme characteristics. Zaprinast, an inhibitor of a cyclic guanosine monophosphate (cGMP)-specific PDE (PDE I), at concentrations up to 100 mumol/L, had no significant effect on the respiratory burst. Milrinone and imazodan, inhibitors of cAMP-metabolizing, cGMP-sensitive PDE (PDE III), reduced the respiratory burst to 60% of control magnitude but only had significant effects when they were introduced at high (100 mumol/L) concentrations. In contrast, rolipram and RO 20-1724, inhibitors of a cAMP-metabolizing, cGMP-insensitive PDE (PDE IV), had significant effects at low concentrations (0.1 mumol/L) and caused marked reduction of the respiratory burst at higher concentrations (25% of control at 10 mumol/L). The selective PDE IV inhibitors significantly potentiated PMN inhibition by isoproterenol. Diethylaminoethyl (DEAE)-Sepharose chromatography demonstrated a predominant PDE isozyme with high affinity and selectivity for cAMP that was insensitive to cGMP and was completely inhibited by rolipram, a PDE IV inhibitor. These results are consistent with the conclusion that the PMN respiratory burst is inhibited by an elevation of cAMP induced by PDE IV inhibition.

Topics: 4-(3-Butoxy-4-methoxybenzyl)-2-imidazolidinone; Adrenergic beta-Agonists; Adult; Calcimycin; Chromatography, Ion Exchange; Humans; Isoenzymes; Isoproterenol; Middle Aged; Milrinone; Neutrophils; Oxygen Consumption; Phosphodiesterase Inhibitors; Phosphoric Diester Hydrolases; Purinones; Pyridazines; Pyridones; Pyrrolidinones; Rolipram; Superoxides

1. Production of endothelium-derived relaxing factor (EDRF) by primary cultures of pig aortic endothelial cells was assessed indirectly by measuring endothelial cyclic GMP content, and prostacyclin production was measured by radioimmunoassay of 6-keto prostaglandin F1 alpha (6-keto PGF1 alpha). 2. The resting level of cyclic GMP fell significantly following removal of extracellular calcium (1 mM EGTA present), but elevations of cyclic GMP content induced by sodium azide (10 microM) or atriopeptin II (10 nM) were similar in the absence and presence of extracellular calcium. 3. Haemoglobin (10 microM) reduced the resting level of cyclic GMP in the presence, but not the absence of extracellular calcium. M&B 22,948 (100 microM), superoxide dismutase (30 u ml-1), bradykinin (0.1 microM), ATP (10 microM) and ionophore A23187 (0.1 microM) each induced an increase in endothelial cyclic GMP content that was reduced in the absence of extracellular calcium. 4. In cascade bioassay experiments using endothelial cells on microcarrier beads and perfused in columns, continuous infusion of bradykinin (0.1 microM) induced release of EDRF, assayed on rabbit aortic rings, that was maximal after 2 min and still detectable up to about 16 min. 5. In the presence of extracellular calcium, the time course of bradykinin (0.1 microM)-stimulated production of EDRF, assessed as endothelial cyclic GMP content was maximal within 1 min, declined thereafter, but was still significant after 30 min. Production of 6-keto PGF1 alpha, measured simultaneously rose rapidly but was complete within 3 min. 6. In the absence of extracellular calcium the resting endothelial content of cyclic GMP fell, but resting production of 6-keto PGF1 alpha was unaffected. 7. In the presence of TMB-8 (100 microM) resting endothelial content of cyclic GMP rose slightly, but production of 6-keto PGFg fell. The bradykinin (0.1 microM)-stimulated increase in cyclic GMP content was augmented, but the stimulation of 6-keto PGF1I production was blocked. Results from cascade bioassay experiments confirmed that TMB-8 (100 microM) did not inhibit bradykinin-induced production of EDRF. 8. The data suggest that resting production of EDRF but not prostacyclin is dependent upon the presence of extracellular calcium. Bradykinin-stimulated production of EDRF is sustained and requires the presence of extracellular calcium, but stimulated production of prostacyclin is transient and may result from discharge of an intracellular po

Topics: Adenosine Triphosphate; Animals; Atrial Natriuretic Factor; Azides; Biological Factors; Bradykinin; Calcimycin; Calcium; Cells, Cultured; Cyclic GMP; Endothelium, Vascular; Epoprostenol; Gallic Acid; Hemoglobins; Humans; Nitric Oxide; Purinones; Sodium Azide; Superoxide Dismutase; Swine