Application of an alpha-sidechain length-specific monoclonal antibody to immunoaffinity purification and enzyme immunoassay of 2,3-dinor-6-keto-prostaglandin F1 alpha from human urine.

2,3-Dinor-6-keto-prostaglandin F1 alpha (2,3-dinor-6-keto-PGF1 alpha) is a major urinary metabolite of PGI2 (prostacyclin) and one of the most reliable parameters of PGI2 production in vivo. A mouse was immunized with 2,3-dinor-6-keto-20-carboxy-PGF1 alpha conjugated to bovine serum albumin for preparation of a monoclonal antibody which recognized the difference in the alpha-sidechain length of 6-keto-PGF1 alpha and its 2,3-dinor-metabolite. A sensitive and specific enzyme immunoassay was developed by the solid-phase competition method with 2,3-dinor-6-keto-20-carboxy-PGF1 alpha labeled with peroxidase protein. The detection range of the assay was 14-1200 fmol (IC50 = 120 fmol). The cross-reactivities of the antibody with 6-keto-PGF1 alpha, 6,15-diketo-13,14-dihydro-PGF1 alpha, and other arachidonate metabolites were less than 0.01%. An immunoaffinity column was prepared by coupling the anti-2,3-dinor-6-keto-PGF1 alpha antibody to BrCN-activated Sepharose 4B. Human urine was applied to an octadecylsilyl silica cartridge, and the extract was applied to the immunoaffinity column. This procedure allowed an efficient separation of 2,3-dinor-6-keto-PGF1 alpha from unidentified urinary substances which interfered with immunoassay. Validity of the results obtained by the enzyme immunoassay was confirmed by GC/MS employing selected ion monitoring for quantification.

Further studies on the effects of epimers of thromboxane A2 antagonists on platelets and veins.

The C-15 hydroxy epimers of three TXA2/PGH2 antagonists were resolved by HPLC and their pharmacology evaluated in canine saphenous veins and human platelets. For all three compounds, the epimers were equipotent in their ability to antagonize U46619 (1 microM) induced platelet aggregation. In contrast, one of the epimers for all three of the antagonists was significantly more potent than the other in antagonizing U46619 (10 nM) induced saphenous vein contractions. The data support the notion that the TXA2/PGH2 receptors in veins may be different from those in platelets.

Congestive heart failure caused by aortocaval fistula after nephrectomy.

We treated a 66-year-old man and a 69-year-old woman with high output cardiac failure due to aortocaval fistula. They underwent nephrectomy 40 and 36 years ago, respectively. They suffered from heart failure for a very long time. However, the etiology was not elucidated until recently. The aortogram revealed massive shunt from the aorta to the inferior vena cava via the right renal artery. Three-dimensional computerized tomography clearly delineated the aortocaval fistula. Surgical closure of the fistula promptly improved heart failure. We stress the importance of history taking concerning nephrectomy and the importance of abdominal auscultation in high output cardiac failure.

Complete obstruction of right proximal pulmonary artery: a case report.

A fifty-five-year-old woman had worried about dyspnea for three years. A chest x-ray appeared normal except for hypovascularity of the upper lung field. On angiograms, a wedge-shaped obstruction was observed at the left proximal pulmonary artery, and well-developed collateral circulation from the right thoracic artery to the internal costal arteries was observed. On the exercise test, exertional dyspnea developed with tachycardia and a decreased saturation of arterial oxygen.

A case of an intraocular foreign body due to graphite pencil lead complicated by endophthalmitis.

We report a case of an 8-year-old boy who presented with an intraocular foreign body composed of graphite pencil lead. The patient had been accidentally poked in the right eye with a graphite pencil. Primary care consisted of corneal suturing and lens extraction. Two pieces of the pencil lead remained in the vitreous cavity following surgery, and 2 days later the patient developed endophthalmitis. Pars plana vitrectomy was performed immediately and the intraocular foreign bodies were removed through the scleral wound. Cultures of the vitreous fluid revealed no bacterial organisms. X-ray fluoroscopic analysis of the vitreous detected 1 ppm of aluminum (a constituent of the pencil lead). Although the clinical presentation indicated probable bacterial endophthalmitis, the detection of elemental aluminum within the vitreous cavity also suggested the possibility of further retinal toxicity due to some dissolving of the pencil lead.

[Bacterial intracranial aneurysm associated with infective endocarditis: a case showing enlargement of aneurysm size].

The authors report a case of bacterial intracranial aneurysm associated with infective endocarditis. A 48-year-old male was admitted on March 26, 1994, with complaints of difficulty in speaking and mild swelling of the right leg following mild fever. On examination he showed motor aphasia and mild weakness of the right upper and lower limbs. Cardiac auscultation revealed a grade 3/6 holosystolic murmur. Laboratory data revealed signs of infection through white blood cell count and CRP. Enterococcus faecalis was isolated from the blood culture at the time of admission. A computerized tomographic (CT) scan and magnetic resonance (MR) imaging showed a round mass with perifocal edema. Angiography revealed an aneurysm from the precentral artery of the left middle cerebral artery. A mycotic aneurysm due to bacterial endocarditis was diagnosed. The patient was treated with high doses of antibiotics. However, angiography 2 weeks after the initial study demonstrated the enlargement of the aneurysm and severe narrowing of the angular artery. On April 19, excision of the aneurysm was performed. Operative findings showed degeneration and thickening of the walls of the aneurysm. After the operation, antibiotic therapy was continued. The patient was asymptomatic upon discharge and has continued to do well. Repeated angiography on September 12 showed no further aneurysm. There is a danger of rupture in mycotic aneurysm due to bacterial endocarditis. It is important to repeat angiography and to manage the primary disease. If an aneurysm enlarges with serial angiography, it should be treated surgically without further delay.

[Sevoflurane inhalation reduced bronchospasm after extubation].

A 73-year-old female was scheduled for left upper lobectomy. She had no history of asthma or chronic obstructive pulmonary disease. During the operation, respiratory sound was clear. As spontaneous breathing was regular and stable after the reversal of neuromuscular blockade, an endotracheal tube was extubated. But soon after the extubation, wheezing and gasping respiration occurred and she complained of dyspnea. Therefore, we administered aminophylline and steroid intravenously, and the patient's lungs were ventilated manually with 100% oxygen. But after 10 minutes, there was no improvement in symptoms, and sevoflurane inhalation was started immediately. Inspiratory sevoflurane concentration was 4% at first, and was decreased to 2%. About 20 minutes after starting sevoflurane inhalation, wheezing was reduced. Sevoflurane may be useful in the treatment of bronchospasm after extubation.

[A case report of primary pulmonary hypertension: congestive heart failure induced by pregnancy and delivery].

A thirty-year-old woman had complaints of general fatigue and gain in body weight, which had developed since the third trimester of pregnancy. On admission, she had lip cyanosis, general edema, and tachycardia. A chest rentogenogram showed cardiomegaly and dilatation of the pulmonary arteries. On cardiac catheterization, pulmonary artery pressure and pulmonary vascular resistance were elevated moderately. A pulmonary perfusion scintigram revealed a defect in the left lower lung field with no evidence of thrombus in a pulmonary artery angiogram. In this case, an underlying disease which caused pulmonary hypertension might have been the underfined cause. For this reason, primary pulmonary hypertension was the diagnosis. In the present case there is a danger that congestive heart failure may be induced by pregnancy and delivery and, furthermore, there is a possibility that during the clinical course of the illness pulmonary infarction might occur.

[Prostacyclin mimetics with non-prostanoid structures].

In searching for new drugs, we have developed receptor binding assays for prostanoids. Among the various compounds that we have tested, we have found that hydronaphthalene derivatives can interact with some prostanoid receptors. Modification of such compounds produced several pure prostacyclin agonists (ONO-AP-227 and ONO-AP-437) and a unique prostacyclin agonist with inhibitory activity against thromboxane synthase (ONO-AP-500-02). These compounds showed specific binding to the IP receptor with Ki values less than 0.2 microM, without binding to EP and TP receptors. These compounds also inhibited human platelet aggregation with IC50 values of 0.03-0.24 microM. These compounds inhibited ex vivo platelet aggregation in dogs or rats in the dose range of 1 to 30 mg/kg. Furthermore, ONO-AP-500-02 inhibited ex vivo thromboxane formation at doses similar to those inhibiting platelet aggregation in rats. These results, taken together, suggest that chemical syntheses of compounds targeting prostanoid receptors can produce unique prostanoid agonists or antagonists, and rational syntheses might be possible for compounds with different pharmacological actions such as inhibition of thromboxane synthase.

Purification and some properties of an inhibitor for a yeast pleiotropic drug resistant pump from Kitasatospora sp. E-420.

A strain producing an inhibitor for pleiotropic drug resistant 5 (Pdr5) was isolated using our original screening system in yeast. The strain was classified and named as Kitasatospora sp. E-420. The purified inhibitor (molecular weight = 1,193 by FAB-MS) inhibited a Pdr5-mediated efflux of cycloheximide and cerulenin. The intracellular accumulation of a fluorescent dye, rhodamine 123, by the inhibitor was also confirmed. Some physicochemical data suggested that the Pdr5-specific inhibitor was different from an immunosuppressant, FK506, reported as the only inhibitor of Pdr5 in vivo.

Ultrasonographic detection of early and curable carcinoma of the gallbladder.

Five patients with early carcinoma of the gallbladder detected by ultrasonography were studied. Three complained of non-specific upper abdominal symptoms or were asymptomatic and two had severe biliary colic. Gallstones were present in the two patients without biliary symptoms. Early carcinoma of the gallbladder was demonstrated as a polypoid tumour by ultrasound in four patients and as thickening of the gallbladder wall in one. The tumour was over 2 cm in diameter in all but one patient in whom the tumour enlarged rapidly from 5 to 10 mm. Size of the tumour and extent of spread were closely related. All but one patient underwent curative resection. Ultrasonography enhances the detection of early carcinoma of the gallbladder especially in old patients with non-specific abdominal symptoms and the operative cure rate is thereby improved.

Coronary vasomotor response to acetylcholine late after angioplasty.

To elucidate the changes in coronary vasomotion in a previously balloon-dilated segment, we examined the vasoconstricting response of previously balloon-dilated and non-dilated segments to acetylcholine. Acetylcholine was administered into coronary arteries cumulatively (left and right coronary artery: 10-100 micrograms) in 15 patients (age: 60 +/- 3 years, 12 males and 3 females) at 7.4 +/- 1.5 months after successful percutaneous transluminal coronary angioplasty (PTCA). In PTCA segments with no restenosis, does-dependent constriction in response to acetylcholine was observed in only 1 patient. In non-PTCA segments of PTCA and non-PTCA arteries, 12 patients showed dose-dependent vasoconstriction in response to acetylcholine. Coronary spasm, which was defined as a more than 75% reduction in coronary diameter compared with that after isosorbide dinitrate, was provoked in one PTCA segment (7%). In non-PTCA segments of PTCA and non-PTCA arteries, 15 of 44 arteries (34%) demonstrated coronary spasm in 9 of 15 patients (60%, p < 0.005 vs PTCA segment). In conclusion, PTCA segments free of restenosis showed no hyper-reactivity to acetylcholine, while non-PTCA segments showed hypercontractility in response to acetylcholine. Coronary balloon angioplasty may alter the coronary vasomotor reaction to acetylcholine several months after angioplasty.

Synthesis of structural analogues of leukotriene B4 and their receptor binding activity.

Structural analogues of leukotriene B4 (LTB4) were designed based on the plausible conformation of LTB4 (1). Joining C-7-C-9 of the conformer A or B into an aromatic ring system led to the discovery of benzene analogues 2, 4 and 6a. Joining C-4-C-9 of the conformer C or D into an aromatic ring system led to the discovery of analogues 3, 5 and 7. The compounds examined in this study were evaluated as to their inhibition of [3H] LTB4 binding to human neutrophils, and by a secondary intact human neutrophil functional assay for agonist/antagonist activity. The first analogues prepared, compounds 2-7, demonstrated moderate potency in the LTB4 receptor binding assay. The modification of these compounds by the introduction of another substituent into the aromatic ring produced a marked increase in receptor binding (28c, IC50 = 0.020 microM; 38c, IC50 = 0.020 microM; 52a, IC50 = 0.020 microM; 52b, IC50 = 0.018 microM). Most of these structural analogues of LTB4 demonstrated agonist activity. Of the analogues prepared in this study, only compound 57 demonstrated weak LTB4 receptor antagonist activity, at 10 microM.

Trisubstituted benzene leukotriene B4 receptor antagonists: synthesis and structure-activity relationships.

A series of trisubstituted benzenes which demonstrate leukotriene B4 (LTB4, 1) receptor affinity was prepared. Previous trisubstituted benzenes from our laboratory showed high affinity to the LTB4 receptor but demonstrated agonist activity in functional assays. Compound 3a, the initial lead compound of this new series, showed only modest affinity (IC50 = 0.20 microM). However, 3a was a receptor antagonist with no demonstrable agonist activity up to 30 microM. Further modification of the lipid tail and aryl head groups region led to the discovery of 3b (ONO-4057). This compound, free of agonist activity, possesses high affinity to the LTB4 receptor (Ki = 3.7 +/- 0.9 nM).

Interaction of mutagenic tryptophan pyrolysate with d(CGATCG)2: intercalation model based on NMR experiments.

The solution structure of the complex of 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1), a potent mutacarcinogen isolated from tryptophan pyrolysate, with the hexamer duplex d(CGATCG)2, was analysed by 1H-NMR spectroscopy and molecular dynamic calculation. Trp-P-1 was intercalated between the CpG base pairings in a suitable manner to form the guanosine-Trp-P-1 adduct which corresponds to the major reactant of Trp-P-1 with DNA.