Azotobacter vinelandii gene clusters for two types of peptidic and catechol siderophores produced in response to molybdenum.

AIM: To characterize the complementary production of two types of siderophores in Azotobacter vinelandii. METHODS AND RESULTS: In an iron-insufficient environment, nitrogen-fixing A. vinelandii produces peptidic (azotobactin) and catechol siderophores for iron uptake to be used as a nitrogenase cofactor. Molybdenum, another nitrogenase cofactor, was also found to affect the production level of siderophores. Wild-type cells excreted azotobactin into molybdenum-supplemented and iron-insufficient medium, although catechol siderophores predominate in molybdenum-free environments. Two gene clusters were identified to be involved in the production of azotobactin and catechol siderophores through gene annotation and disruption. Azotobactin-deficient mutant cells produced catechol siderophores under the molybdenum-supplemented and iron-insufficient conditions, whereas catechol siderophore-deficient mutant cells extracellularly secreted excess azotobactin under iron-deficient condition independent of the concentration of molybdenum. This evidence suggests that a complementary siderophore production system exists in A. vinelandii. CONCLUSIONS: Molybdenum was found to regulate the production level of two types of siderophores. Azotobacter vinelandii cells are equipped with a complementary production system for nitrogen fixation in response to a limited quantity of metals. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first study identifying A. vinelandii gene clusters for the biosynthesis of two types of siderophores and clarifying the relationship between them.

Biosynthetic characterization and biochemical features of the third natural nisin variant, nisin Q, produced by Lactococcus lactis 61-14.

AIMS: To characterize the genetic and biochemical features of nisin Q. METHODS AND RESULTS: The nisin Q gene cluster was sequenced, and 11 putative orfs having 82% homology with the nisin A biosynthesis gene cluster were identified. Nisin Q production was confirmed from the nisQ-introduced nisin Z producer. In the reporter assay, nisin Q exhibited an induction level that was threefold lower than that of nisin A. Nisin Q demonstrated an antimicrobial spectrum similar to those of the other nisins. Under oxidizing conditions, nisin Q retained a higher level of activity than nisin A. This higher oxidative tolerance could be attributed to the presence of only one methionine residue in nisin Q, in contrast to other nisins that contain two. CONCLUSIONS: The 11 orfs of the nisin producers were identical with regard to their functions. The antimicrobial spectra of the three natural nisins were similar. Nisin Q demonstrated higher oxidative tolerance than nisin A. SIGNIFICANCE AND IMPACT OF THE STUDY: Genetic and biochemical features of nisin Q are similar to those of other variants. Moreover, owing to its higher oxidative tolerance, nisin Q is a potential alternative for nisin A.

Specific but differential antagonism by glibenclamide of the vasodepressor effects of cromakalim and nicorandil in spinally-anaesthetized dogs.

1. Whether the vasodepressor effects of cromakalim and nicorandil, potassium channel openers, were differentially antagonized by glibenclamide, a supposed specific antagonist of potassium channel openers, was investigated in spinally-anaesthetized dogs in which arterial pressure was maintained elevated by i.v. infusion of noradrenaline. 2. Cumulative administration of cromakalim (3-100 micrograms kg-1, i.v.), nicorandil (30-1000 micrograms kg-1, i.v.), diltiazem (3-300 micrograms kg-1, i.v.) and nitroglycerin (0.3-100 micrograms kg-1, i.v.) caused dose-dependent decreases in mean arterial pressure. In dogs which received glibenclamide (3 mg kg-1, i.v.), the dose-vasodepressor response curves for cromakalim and nicorandil were located on the right in parallel to the respective curves determined in control dogs, but those for diltiazem and nitroglycerin were not different. ED50% values increased about 6.7 fold for cromakalim and 2.2 fold for nicorandil. 3. The depression of LV dP/dtmax produced by a high dose of cromakalim (100 micrograms kg-1, i.v.) was abolished by glibenclamide and that produced by nicorandil was not only antagonized but converted to an increase. 4. These results suggest that the vasodepressor action of cromakalim is due predominantly to potassium channel opening, but that of nicorandil involves not only potassium channel opening but its action as a nitrate.

Comparison of the effects of the novel vasodilator FK409 with those of nitroglycerin in isolated coronary artery of the dog.

1. The vasorelaxant effects of FK409, a new nitrovasodilator synthesized from a microbial product, were compared with those of nitroglycerin in isolated coronary artery rings of the dog contracted with U46619 (10(-7) M). 2. FK409 (10(-11)-10(-5) M) and nitroglycerin (10(-9)-10(-4) M) each produced a concentration-dependent relaxation. Comparison of EC50 values showed that FK409 was about 25 times more potent than nitroglycerin. 3. Submaximum concentrations of nitroglycerin (10(-6) M) and FK409 (3 x 10(-8) M) elevated guanosine 3':5'-cyclic monophosphate (cyclic GMP) levels, effects associated with vasorelaxation. Adenosine 3':5'-cyclic monophosphate (cyclic AMP) levels were unaffected. 4. The concentration-relaxation curves for nitroglycerin and FK409 were shifted to the right by methylene blue (3 x 10(-6) - 3 x 10(-5) M), an inhibitor of soluble guanylate cyclase, and to the left by M&B22,948 (3 x 10(-6) - 3 x 10(-5) M), an inhibitor of cyclic GMP phosphodiesterase. 5. After exposure of coronary arteries to the maximally-effective concentration of nitroglycerin (10(-4) M), the mean EC50 value of FK409 did not change significantly, although that of nitroglycerin increased about 60 fold. After exposure to the maximally-effective concentration of FK409 (10(-5) M), the mean EC50 value of FK409 increased about 6 fold and that of nitroglycerin about 11 fold. 6. These results suggest that the vasorelaxant effect of FK409, like that of nitroglycerin, is due to activation of soluble guanylate cyclase and a resultant increase in intracellular cyclic GMP. However, compared with nitroglycerin, there was less self-tolerance to the relaxant effects of FK409 and relatively little cross-tolerance between the two agents.

Cardiohemodynamic effect of FK409, a novel highly potent nitrovasodilator, in anesthetized dogs.

The cardiohemodynamic effect of (+/-)-(E)-4-ethyl-2-[(E)-hydroxyimino]-5-nitro-3-hexenamide (FK409), a novel potent vasodilator, was studied in anesthetized open-chest dogs. FK409 (1 to 10 micrograms/kg, i.v.) decreased mean blood pressure, cardiac output and venous return (sum of the flow through the inferior and the superior vena cava). These changes accompanied decreases in left ventricular pressure, in its maximum rate of rise and in right atrial pressure. This cardiovascular profile of FK409 is very similar to those of classical nitrates.

Vasodepressor mechanisms of 2-(1-octynyl)-adenosine (YT-146), a selective adenosine A2 receptor agonist, involve the opening of glibenclamide-sensitive K+ channels.

The vasodepressor mechanism of 2-(1-octynyl)-adenosine (YT-146), a selective adenosine A2 receptor agonist, was compared with that of adenosine in spinally anaesthetized dogs whose blood pressure was kept elevated with i.v. infusion of noradrenaline. Cumulative i.v. administration of YT-146 (1-1000 nmol/kg) caused a slowly developing and long-lasting decrease in mean blood pressure (MBP) and a small decrease in heart rate (HR) at high doses, whereas single i.v. administration of adenosine caused short-lived decreases in MBP and HR. In dogs given glibenclamide (6 mumol/kg i.v.) or theophylline (30 mumol/kg i.v.) dose-response curves for decrease in MBP due to YT-146 underwent parallel rightward shifts. The ED50 values for YT-146 were increased about 5.5-fold with glibenclamide and about 11.9-fold with theophylline. However, no further rightward shifts of the curves were obtainable in dogs given glibenclamide plus theophylline; the increase in ED50 values for YT-146 was about 6.8-fold, being close to the increase obtained with theophylline alone. The ED50 values for adenosine to lower MBP increased about 1.6-fold with theophylline but not with glibenclamide. Glibenclamide and theophylline failed to antagonize the negative chronotropic effects of both drugs. These results suggest that the vasodepressor effect of YT-146 involves two mechanisms following stimulation of adenosine A2 receptors; one is probably a cyclic AMP-dependent mechanism and the other is the opening of glibenclamide-sensitive K+ channels.

TY-12533, a novel Na(+)/H(+) exchange inhibitor, prevents myocardial stunning in dogs.

Anesthetized open-chest dogs were subjected to 15-min myocardial ischemia followed by 2-h reperfusion to induce myocardial stunning. A novel Na(+)/H(+) exchange inhibitor 6,7,8,9-tetrahydro-2-methyl-5H-cyclohepta[b]pyridine-3-carbonylguanidine maleate (TY-12533), administered 10 min before or 10 min after start of ischemia (3 mg/kg/10 min, i.v.), did not affect reductions in regional myocardial wall thickening, blood flow and pH during ischemia, but it significantly improved recovery of the wall thickening and blood flow after reperfusion. These results indicate that TY-12533, even when administered during ischemia, could prevent myocardial stunning without affecting myocardial dysfunction or acidosis induced by brief ischemia.

Cardioprotective effect of TY-12533, a novel Na(+)/H(+) exchange inhibitor, on ischemia/reperfusion injury.

The effects of 6,7,8, 9-tetrahydro-2-methyl-5H-cyclohepta[b]pyridine-3-carbonylguanidine maleate (TY-12533) on myocardial ischemia/reperfusion injury were evaluated in rats. Inhibitory effects of TY-12533, TY-50893 (the 9-chloro derivative of TY-12533) and cariporide on the platelet Na(+)/H(+) exchanger in vitro were almost equal at pH 6.2 and decreased at pH 6.7; but TY-12533 was four times more potent than TY-50893 and cariporide at pH 6.7. TY-12533, TY-50893 and cariporide administered before ischemia (0.01-1 mg/kg, i.v.) suppressed the ischemia/reperfusion-induced arrhythmias to the same extent in vivo; but TY-12533 was more effective than cariporide and TY-50893 when they were administered during ischemia (0.1-1 mg/kg). Similar results were obtained for the inhibitory effects of these drugs administered before ischemia (0.03-0.1 mg/kg, i.v.) and during ischemia (0.1-1 mg/kg) on the ischemia/reperfusion-induced myocardial infarction. These differences between TY-12533 and the other drugs in vitro and in vivo may be ascribed to the pK(a) values of the guanidinium moiety of TY-12533 (6.93), TY-50893 (6.35) and cariporide (6.28).

The group at C2 of N-ethylnicotinamide determines the vasodilator potencies and mechanisms of action of nicorandil and its congeners in canine coronary arteries.

The relaxant mechanisms of action of nicorandil and its congeners (SG-86, SG-103, SG-209 and SG-212) on large coronary arteries were investigated in isolated canine circumflex arteries contracted with 25 mmol/l KCl or 10(-7) mol/l U46619, a thromboxane A2 analogue. SG-212, SG-86, SG-209 and SG-103 were obtained by replacement of the nitroxy group of nicorandil by bromine, the hydroxy, acetoxy and nicotinoyloxy groups, respectively. Nicorandil (10(-6)-10(-3) mol/l), SG-212 (3 x 10(-4)-10(-2) mol/l), SG-209 (10(-4)-10(-2) mol/l), SG-103 (3 x 10(-4)-10(-2) mol/l), and SG-86 (10(-3)-10(-2) mol/l) all produced a concentration-dependent relaxation in KCl- or U46619-contracted arteries. The order of relaxant potency was as follows: Nicorandil much greater than SG-209 greater than SG-212 = SG-103 greater than SG-86. The relaxant effect of nicorandil was not affected by glibenclamide but antagonized by methylene blue. In the presence of glibenclamide, the concentration-relaxation curves for SG-209 underwent rightward parallel shifts. The relaxant effect of SG-209, however, was not affected by methylene blue. The concentration-relaxation curves for SG-212 underwent rightward parallel shifts only to a limited extent in the presence of glibenclamide, but they were not affected by methylene blue. The relaxant effect of SG-103 was affected by neither glibenclamide or methylene blue. The relaxant effect of SG-86 was not affected by glibenclamide. The relaxant effect of nicorandil accompanied an increase in cyclic-GMP levels but that of SG-209 did not.(ABSTRACT TRUNCATED AT 250 WORDS)

Nicorandil increases coronary blood flow predominantly by K-channel opening mechanism.

Nicorandil has a hybrid property between nitrates and potassium (K)-channel openers. In order to clarify which mechanism of action is responsible for its effect in increasing coronary blood flow, we investigated how this effect was antagonized by glibenclamide, which was recently found to behave as a pharmacologic antagonist of K-channel openers. Cromakalim, one of the most specific K-channel openers currently available, and nitroglycerin were used as reference drugs. In isolated, blood-perfused papillary muscle preparations of dogs, intraarterial injections of nicorandil and cromakalim increased (coronary) blood flow, and at high doses a negative inotropic effect and ventricular fibrillation occurred. Dose-response curves for the increase in coronary blood flow produced by nicorandil or cromakalim were shifted to the right in a parallel manner and to similar extents by glibenclamide given intravenously to support dogs. Schild analysis yielded pA2 values of 6.08 and 6.34 for glibenclamide versus nicorandil and cromakalim, respectively. Nitroglycerin injected intraarterially produced only an increase in coronary blood flow. This effect was not affected by glibenclamide. These results indicate that the effect of nicorandil in increasing coronary blood flow, like that of cromakalim, is predominantly due to its mechanism of action as a K-channel opener. The negative inotropy and ventricular fibrillation seen with high doses of nicroandil and cromakalim were also antagonized by glibenclamide, indicating that these effects are also due to K-channel opening.

Manometric findings of the upper esophageal sphincter in esophageal achalasia.

Pharyngeal and upper esophageal sphincter (UES) manometry was performed in 15 patients with esophageal achalasia and compared with that in 10 healthy controls. Neither the pharyngeal contraction pressure nor the UES resting pressure were significantly different between the two groups, although the UES residual pressure in patients with achalasia was significantly increased compared with that in controls. Pneumatic dilatation of the lower esophageal sphincter (LES) was performed in these patients. After successful LES dilatation, the increased UES residual pressure in patients with esophageal achalasia decreased significantly. Our results suggest that UES relaxation in patients with esophageal achalasia is incomplete compared with that in healthy adults. This UES abnormality is not a primary defect but a secondary phenomenon.

A further study of the vasodilator and negative inotropic mechanisms of action of nicorandil and its congeners in the canine heart.

The vasodilator and negative inotropic mechanisms of action of nicorandil and its congeners (SG-209, SG-103, and SG-86) were investigated in isolated canine papillary muscle preparations cross-circulated through the anterior septal artery with support dogs. SG-209, SG-103, and SG-86 were obtained by replacement of the nitroxyl group of nicorandil by acetoxyl, nicotinoyloxyl, and hydroxyl groups, respectively. Nicorandil (0.03-10 mumol), SG-209 (0.1-10 mumol), SG-103 (1-30 mumol), and SG-86 (3-100 mumol) all produced an increase in coronary blood flow through the anterior septal artery. Both nicorandil and SG-209 produced a near-maximal increase in coronary blood flow, the latter being about 5.5 times less potent than the former. SG-103 and SG-86 were far less potent than SG-209 in that order. The vasodilator actions of nicorandil and SG-209 were antagonized by glibenclamide given IV to support dogs, but those of SG-103 and SG-86 were not. The pKB values of glibenclamide were 6.34 toward nicorandil and 6.49 toward SG-209. Developed tension of the papillary muscle was reduced by nicorandil, SG-209, and SG-103, but not by SG-86. In this respect SG-209 was about 4.7 times less potent than nicorandil, and SG-103 was much less potent than SG-209. The negative inotropic effects of nicorandil and SG-209 were antagonized by glibenclamide, but that of SG-103 was not.(ABSTRACT TRUNCATED AT 250 WORDS)

Cardiac versus coronary dilator effects of SD-3211, a new nondihydropyridine calcium antagonist, in isolated, blood-perfused dog hearts.

Coronary and cardiac effects of SD-3211 were compared in isolated, blood-perfused sinoatrial (SA) node, atrioventricular (AV) node, and papillary muscle preparations of dogs. SD-3211 was administered intraarterially. In all preparations SD-3211 produced an increase in coronary blood flow. In SA node preparations, the drug produced a decrease in sinus rate, and in high doses atrial standstill occurred. The dose that produced a 15% decrease in sinus rate was about 5.6 times the dose that doubled coronary blood flow. In AV node preparations, when injected into the artery supplying the AV node, the drug produced an increase in AV conduction time, and in high doses second- or third-degree AV block occurred. The dose that produced a 15% increase in AV conduction time was about 1.6 times the dose that doubled coronary blood flow. In the same preparations the drug slightly increased AV conduction time only at high doses when injected into the artery supplying the His-Purkinje-ventricular system. In paced papillary muscle preparations, the drug produced a decrease in the force of contraction. The dose that produced a 50% decrease in the force of contraction of the paced papillary muscle was about 50 times the dose that doubled coronary blood flow. The drug was virtually ineffective on ventricular automaticity. In short, in doses that doubled coronary blood flow, SD-3211 depressed only AV nodal conduction. This cardiovascular profile differs from those of diltiazem and verapamil, which do not discriminate between coronary vasculature and the SA and the AV node.

Similarity and dissimilarity in mode and mechanism of action between YT-146, a selective adenosine receptor A2 agonist, and adenosine in isolated canine hearts.

To elucidate the differences in mode and mechanism of action between YT-146, a highly selective adenosine A2 receptor agonist, and adenosine, we compared their effects on coronary circulation and myocardium and modifications of these effects by glibenclamide, a blocker of ATP-sensitive potassium (K) channels, in three kinds of isolated, blood-perfused canine heart preparations. YT-146 and adenosine were injected i.a. In all preparations both YT-146 and adenosine increased coronary blood flow and in this respect YT-146 was about 5 times as potent as adenosine. The increase in blood flow caused by adenosine was transient, whereas that produced by YT-146 was biphasic; the transient increase was followed by a sustained one. In isolated, blood-perfused sinoatrial (SA) node preparations, YT-146 failed to affect sinus rate, whereas adenosine reduced sinus rate by about 38% at its maximum effect. In isolated, blood-perfused atrioventricular (AV) node preparations, when injected into the artery supplying the AV node, YT-146 exerted no effect on AV conduction time, whereas adenosine prolonged AV conduction time by about 17% at the maximum effect. In isolated, blood-perfused papillary muscle preparations, the force of contraction was affected by neither YT-146 nor adenosine. In the same preparations the effect of YT-146 in increasing coronary blood flow was antagonized by glibenclamide in such a manner that the maximum increase was suppressed, but that of adenosine was not. Reactive hyperemia induced by ischemia for 30 seconds was not affected by glibenclamide. These results suggest that although both YT-146 and adenosine produce an increase in coronary blood flow via adenosine A2 receptors, the opening of ATP- or glibenclamide-sensitive K channels is involved in the action of the former, but scarcely in the action of the latter. The opening of ATP- or glibenclamide-sensitive K-channels is less likely involved in reactive hyperemia.

Aneurysmal rupture of the pancreaticoduodenal artery successfully treated by transcatheter arterial embolization.

We report a case of aneurysmal rupture of the pancreaticoduodenal artery successfully treated by transcatheter arterial embolization. A 61-year-old man with a history of hypertension underwent surgery at our hospital in November 1995 for local peritonitis caused by perforation of the sigmoid colon secondary to cancer. On the 9th postoperative day, he developed shock, with complaints of epigastric and back pain. Abdominal computed tomography showed an enhanced mass, thought to be a peripancreatic aneurysm. Emergency angiography demonstrated an aneurysm arising from the arcade of the anterior pancreaticoduodenal artery. After diagnostic angiography, transcatheter arterial embolization was performed. With steel coils, the anterior superior pancreaticoduodenal artery and anterior inferior pancreaticoduodenal artery were embolized near the origin of the aneurysm. Angiography 7 weeks later revealed no recanalization of the aneurysm and the absence of anomalous collateral vessels. The patient has been well for 19 months without re-bleeding or recurrence of sigmoid colon cancer. Transcatheter arterial embolization is an effective therapeutic approach for aneurysm of the pancreaticoduodenal artery and is the preferred initial treatment.

Clinicopathological features of superficial spreading and nonspreading squamous cell carcinoma of the esophagus.

OBJECTIVES: Superficially spreading carcinoma of the esophagus, consisting mainly of intraepithelial carcinoma, is not as rare as was previously thought. Despite the surgical significance of this entity, no general definition has been established, and the clinical features of this disease remain to be clarified. METHODS: A total of 54 patients with superficial carcinoma of the esophagus (defined as carcinoma limited to the epithelium or superficially invading the lamina propria or submucosa) were classified into two groups according to the longitudinal extent of the lesion. A total of 13 patients with superficially spreading carcinoma (defined as a superficial carcinoma measuring >5 cm and consisting mainly of intraepithelial carcinoma) were compared to 41 patients with nonspreading esophageal carcinoma. RESULTS: One patient with superficially spreading carcinoma had a positive resection margin because of multiple cancerous lesions. The only significant difference in the clinical and pathological features of the two groups was a higher prevalence of associated multiple cancerous lesions in patients with the superficially spreading type. CONCLUSIONS: Superficially spreading carcinoma of the esophagus is often associated with multiple cancerous lesions. For endoscopists and esophageal surgeons, it is important to define the proximal extent of intraepithelial cancer and the presence of multiple cancerous lesions to perform curative resection.

Primary lymphoma of the liver with bile duct invasion and tumoral occlusion of the portal vein: report of a case.

A 55-year-old woman presented to hospital with epigastric pain and jaundice. Diagnostic imaging studies revealed a biliary stricture of the hepatic confluence and a hepatic tumour of the left and caudate lobes with a portal tumour thrombus, which occupied the main portal trunk, the umbilical portion of the left portal vein, and the right anterior and posterior portal branches. Left hepatic trisegmentectomy, caudate lobectomy, portal tumour thrombectomy, bile duct resection and bilioenteric anastomosis were performed. There were no other lesions, and so it was diagnosed as a primary lymphoma of the liver (B-cell, diffuse, large cell type). The patient underwent postoperative chemotherapy and has remained well for 4.5 years after surgery. Primary lymphoma of the liver is very rare, and this is the first case report with bile duct invasion and tumoral occlusion of the portal vein.