A 94-kDa protein, identified as neutral endopeptidase-24.11, can inactivate atrial natriuretic peptide in the vascular endothelium.

Neutral endopeptidase 24.11 (EC 3.4.24.11) inactivates atrial natriuretic peptide by cleaving the hormone between Cys7 and Phe8, and inhibitors of the enzyme have consequent natriuretic and diuretic properties. The in vivo sites of degradation of this peptide by the zinc-metallopeptidase, however, remain to be established. Because an endopeptidase-24.11-like activity has recently been reported in the rat mesenteric artery, we have further investigated the degradation of atrial natriuretic peptide in vascular tissue. Endopeptidase-24.11 activity was detected in solubilized membrane preparations from rat and rabbit vascular tissue, using [3H]D-Ala2-leucine enkephalin as substrate, and both rabbit and rat aorta preparations were also found to cleave atrial natriuretic peptide between Cys7 and Phe8. In both cases, hydrolysis was inhibited by neutral endopeptidase inhibitors, with Ki values close to their Ki values for the pure enzyme. In preparations of rabbit aorta denuded of endothelium by saponin treatment, the hydrolysis of the Gly3-Phe4 bond of [3H]D-Ala2-leucine enkephalin and the Cys7-Phe8 bond of atrial natriuretic peptide was reduced by greater than 90%. The high performance liquid chromatography method used to follow the degradation of atrial natriuretic peptide differed from previously published procedures, in that samples to be injected were first treated with excess dithiothreitol to reduce the Cys7-Cys23 disulfide bridge. This facilitated the separation of the intact peptide and its metabolites. The presence of the 94-kDa neutral endopeptidase in rabbit aortic tissue was definitively established using a new potent 125I-labeled inhibitor, [125I]RB104 [2-[(3-[125I]iodo-4-hydroxy)phenylmethyl]-4-N-[3- hydroxyamino-3-oxo-1-phenylmethyl propyl]amino-4-oxobutanoic acid] (Ki, 30 pM), which selectively labeled the enzyme after sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the membrane preparations. Therefore, despite its low concentrations in the vasculature, the presence of endopeptidase-24.11 almost exclusively in endothelial tissue suggests that the enzyme is ideally localized to inactivate circulating atrial natriuretic peptide.     

The interaction of parafluorohexahydrosiladiphenidol at muscarinic receptors in vitro

1. The antagonistic actions of parafluorohexahydrosiladiphenidol (pFHHSiD) at muscarinic receptors has been studied in cardiac muscle, smooth muscle and cell culture preparations. In this paper, the classification scheme of Doods et al. (1987) is employed. This scheme is based upon differential affinities of muscarinic antagonists. pFHHSiD exhibited high pA2 values at M3 receptors mediating contractions of guinea-pig ileum and oesophageal muscularis mucosae (7.8 and 8.2 respectively) whereas low values were determined at M2 receptors mediating negative inotropic responses in guinea-pig atria (6.0). Intermediate pA2 values were determined at M1 receptors mediating contractions of the canine femoral and saphenous veins. 2. The pA2 values of pFHHSiD at receptors mediating endothelial-dependent relaxation of rat aortic rings, rabbit jugular vein and canine femoral artery (7.6-7.9) were similar to those determined on the ileum. However, the pA2 values of pFHHSiD at receptors mediating contractions of the guinea-pig trachea (7.1), which has been previously shown to possess M3 receptors, were different from those determined in the ileum. 3. The similarity in pA2 values of pFHHSiD between the M3 receptors in guinea-pig ileum and the receptors mediating endothelial-dependent relaxations provide further evidence for the role of M3 receptors in this vascular response. Taken together, pA2 values for pFHHSiD range from 7.1 to 8.2, depending upon the M3 preparation used. The selectivity of the compound therefore for the M3 versus the M2 muscarinic receptor ranged from 13 to 163 fold.(ABSTRACT TRUNCATED AT 250 WORDS)     

Inhibition of Human Mammary Cell Line Proliferation by Membrane Lectin-Mediated Uptake of Ha-ras Antisense Oligodeoxynucleotide

The Ha-ras oncogene promotes cell proliferation. Antisense oligonucleotides complementary to the ras gene sequence encompassing a mutated codon 12 selectively induce a cell proliferation inhibition. However, the concentration required to reach an effective inhibition is high due to the low efficiency of the oligonucleotide crossing through cell membranes, leading to a low concentration in the cytosol and/or the nucleoplasm. In the present paper, we show that anti-ras oligonucleotides linked to a glycosylated carrier, serum albumin bearing mannose 6-phosphate residues, are more efficient than free oligonucleotides or oligonucleotides bound to an unglycosylated carrier at inhibiting proliferation of a human tumor mammary cell line expressing the mutated Ha-ras. Using fluorescein-labeled neoglycoproteins and fluorescein-labeled oligonucleotides bound to neoglycoproteins, flow cytometry and confocal microscopy revealed that (i) these tumor cells express a membrane lectin specific for mannose 6-phosphate-bearing proteins, (ii) the membrane lectin actively mediates the uptake of macromolecules substituted with mannose 6-phosphate, and (iii) the fluorescein-labeled oligonucleotides bound to the neoglycoprotein accumulate in intracellular vesicles. Furthermore, with antisense oligonucleotides carried by the neoglycoproteins, the concentration required to inhibit cell proliferation is lower than that of the carrier-free antisense oligonucleotides.     

Laparoscopic hysterectomy

Summary Thirty-three patients were selected for laparoscopic hysterectomy and operated on in the Department of Obstetrics, Gynecology and Reproductive Medicine of Clermont-Ferrand University Hospital. Surgical techniques included blunt dissection with scissors and bipolar coagulation to achieve hemostasis. A case was considered successful when all the uterine vessels were treated by laparoscopy. Twenty-four cases were completed laparoscopically (72.7%). None of these patients had postoperative bleeding; 22 had an uneventful postoperative recovery. Nine procedures were converted to laparotomy (27.3%), five because of a difficult or unsatisfactory hemostasis. We conclude that in selected cases, a total hysterectomy can be performed safely by experienced laparoscopists. Further technological progress is necessary to make this procedure more acceptable. Its value as compared to the others will have to be demonstrated.     

Nonadrenergic [3H]idazoxan binding sites are physically distinct from alpha 2-adrenergic receptors

We have recently demonstrated that the alpha 2-adrenergic radioligand [3H]idazoxan also labels additional sites that do not recognize catecholamines but bind with high affinity several chemically distinct drugs previously assumed to be highly selective for alpha 2-adrenergic receptors [Mol. Pharmacol. 35:324-330 (1989)]. We now have used three approaches to distinguish the nonadrenergic [3H]idazoxan sites from alpha 2-adrenergic receptors. (a) No nonadrenergic [3H]idazoxan binding sites were found in COS-7 cells transfected with the genes for the two known alpha 2-adrenergic receptor subtypes. (b) The ratio of alpha 2-adrenergic and nonadrenergic [3H]idazoxan sites in human platelet membranes varied considerably between various donors. (c) Highly purified platelet plasma membranes were enriched for alpha 2-adrenergic receptors but did not contain any nonadrenergic [3H]idazoxan binding sites. We conclude that the nonadrenergic [3H]idazoxan binding sites are not co-expressed with alpha 2-adrenergic receptors and at least in human platelets may be located in an intracellular compartment.     

Pharmacokinetics of prednisolone in children with the nephrotic syndrome

The aim of this study was to establish whether the criteria for the clinical effectiveness of steroids are correlated with the pharmacokinetics of prednisolone in children treated with prednisone during an attack of idiopathic nephrotic syndrome (INS). Thirteen patients with nephrosis were included. Prednisolone, prednisone and cortisol levels were measured using a specific high-performance liquid chromatography assay after an oral dose of 1 mg/kg body weight of prednisone taken at the onset of the disease. All the pharmacokinetic parameters, including the conversion of prednisone to prednisolone were similar to the data already published in children with INS. No correlation was found between the values of pharmacokinetic parameters and criteria of clinical effectiveness. Hypo-albuminaemia was significantly correlated with the area under the plasma-concentration curve but not with the elimination half-life of prednisolone. Moreover, the prednisolone elimination half-life correlated with the urinary exretion of 17-hydroxycorticosteroids achieved in the first 6h. The present study suggests that routine measurements of prednisolone kinetics do not help when assessing the treatment of children with INS.     

Metabolic myocardial viability assessment with iodine 123-16-iodo-3-methylhexadecanoic acid in recent myocardial infarction: Comparison with thallium-201 and fluorine-18 fluorodeoxyglucose

The best test presently available to ascertain residual viability within an infarct-related area involves the use of fluorine-18 fluorodeoxyglucose (FDG) to detect the persistence of some cellular metabolism. Rest reinjection of thallium-201 is a less accurate alternative but is easy to perform. Iodinated fatty acids, which are used with standard gamma cameras, are proposed as markers of cellular metabolism. This study was performed to assess the value of 16-iodo-3-methyl-hexadecanoic acid (MIHA) as a marker of the residual cellular metabolism by comparison with FDG in patients with a recent myocardial infarction, and to evaluate its contribution compared with the²⁰¹Tl stress-redistribution-reinjection technique. Stress-redistribution-reinjection²⁰¹T1 imaging, rest MIHA imaging and glucoseloaded FDG imaging were performed in 22 patients with recent myocardial infarction. Out of the 628 myocardial segments obtained from the left ventricular analysis, 400 were hypoperfused (relative uptake <0.75 of maximum uptake on stress²⁰¹T1 imaging), 177 of which were severely hypoperfused (relative uptake <0.50). Receiver operating characteristic (ROC) curves for predicting metabolic myocardial viability with FDG were derived from the results in respect of (a)²⁰¹T1 activity during exercise, redistribution and reinjection and (b) MIHA up-take, using the two FDG thresholds most commonly considered to define metabolic viability (0.50 and 0.60). Analysis of the 400 hypoperfused segments demonstrated that²⁰¹T1 reinjection was the most accurate test in predicting the presence of myocardial viability (area under the ROI curves=0.85 and 0.86 at the 0.50 and 0.60 FDG thresholds, respectively;P<0.05 vs other tests). The global predictive values of MIHA and²⁰¹T1 reinjection were, respectively, 0.87 and 0.89 at the 0.50 FDG threshold (NS), and 0.82 and 0.87 at the 0.60 FDG threshold (NS). When only the 177 severely hypoperfused segments were considered,²⁰¹T1 reinjection remained the most accurate test (accuracy 0.84 at the 0.50 FDG threshold and 0.82 at the 0.60 FDG threshold), while the accuracy of MIHA decreased significantly (0.78 at the 0.50 FDG threshold and 0.73 at the 0.60 FDG threshold,P<0.05 vs²⁰¹T1 reinjection). In all circumstances, MIHA was less specific than²⁰¹T1 reinjection for the detection of metabolic viability. In conclusion, in patients with recent myocardial infarction, MIHA accurately detects the persistence of metabolic viability, but is not superior to²⁰¹T1.     

Brain glucose utilisation in acquired childhood aphasia associated with a sylvian arachnoid cyst: recovery after shunting as demonstrated by PET.

Regional brain glucose utilisation was investigated with PET and fluorodeoxyglucose (FDG) in a case of epileptic aphasia (Landau-Kleffner syndrome) associated with a left sylvian arachnoid cyst. CT and MRI had failed to disclose any mass effect of the cyst on surrounding brain structures. Sequential metabolic measurements showed a comparable pronounced hypometabolism in cortical regions around the cyst, involving speech areas, and suggested mild but chronic compression of the developing brain. After placement of a cyst-peritoneal shunt system, significant metabolic improvement occurred in all cortical regions, especially the inferior frontal gyrus and the perisylvian area, with predominant residual deficit in the left superior temporal gyrus. These findings were correlated with a pronounced increase in word fluency and slower progress in verbal auditory comprehension. This report suggests that PET is able to evaluate the functional disturbances associated with expanding arachnoid cysts, and to follow the neurological improvement after drainage.     

Life-threatening respiratory failure due to cranial dystonia after dental procedure in a patient with multiple system atrophy.

We report on a woman with a an 8-year history of multiple system atrophy with predominance of parkinsonism who developed jaw-locking oromandibular dystonia within hours after insertion of ill-fitting dentures. Dystonia spread rapidly to involve other facial muscles and the larynx causing stridor with respiratory failure necessitating crush intubation.