Human midkine (hMK), a novel heparin-binding neurotrophic factor consisting of 121 amino acid residues with five intramolecular disulphide bonds, was synthesized by solution procedure in order to demonstrate the usefulness of our newly developed solvent system, a mixture of dichloromethane or chloroform and trifluoroethanol. The final protected 121-residue peptide was assembled from two large fully protected intermediates, Boc-(1-59)-OH and H-(60-121)-OBzl, in CHL/TFE(3:1, v/v) using water-soluble carbodiimide in the presence of HOOBt as coupling reagents. After removal of the protecting groups by HF followed by treatment with Hg(OAc)2 in 50% acetic acid, the fully deprotected peptide was subjected to the oxidative folding reaction. The final product was confirmed to have the correct disulphide structure from its tryptic peptide mapping and to possess the same biological activities as those of the natural product. In order to clarify the active region of the hMK molecule, the N-terminal half domains [(1-59) and (60-121)] were also synthesized by the same procedure used for the hMK synthesis. The C-half domain was confirmed to show the full pattern of bioactivities except for the neuronal cell survival activity, while the N-half one showed much less activity in general.

Amino Acid Sequence↗

PubMedDate not supplied

Fluorescence digital image analysis of serotonin-induced calcium oscillations in single blood platelets.

The intracellular concentration of Ca2+ [( Ca2+]i) was monitored continuously in single rabbit blood platelets by digital imaging microscopy in conjunction with Fura-2, a specific Ca(2+)-indicator dye. Ionomycin as well as aluminium fluoride caused sustained increase in [Ca2+]i in the platelet, but oscillations of [Ca2+]i were not observed. Serotonin (5-HT) induced oscillatory increases in [Ca2+]i in the presence of 1 mM CaCl2; these had not been detectable in cell populations because the oscillations were not in synchrony. This effect of 5-HT was diminished when CaCl2 was omitted from the medium, and was antagonized by 1 microM ketanserin, a specific 5-HT2 receptor antagonist. Furthermore, DOI, a specific 5-HT2 agonist, had the same effect as 5-HT at lower concentration. A specific effector mechanism, not fully understood at present, therefore appears to mediate 5-HT2 receptors thereby allowing rabbit platelets to generate [Ca2+]i oscillations. It is suggested that protein kinase C in platelets might play a key role in the regulation of [Ca2+]i, and possibly in [Ca2+]i oscillations.

Aluminum↗

PubMedDate not supplied

Severe neonatal nemaline myopathy with delayed maturation of muscle.

A Japanese infant with a severe neonatal form of nemaline myopathy showed features of muscle immaturity as well as rods in the biopsied quadriceps femoris muscle, and involvement of diaphragm was first confirmed at autopsy. The biopsied muscle showed numerous rods in 80% of muscle fibers, and an increased number of type 2C fibers (23.2%). Electron microscopic findings were characterized by the presence of many small, immature round fibers with central nuclei and sparse myofibrils, and an increased number of satellite cells in close association with the small muscle fibers, as well as numerous rod structures. These microscopic and electron microscopic findings are interpreted as immature muscle fibers. Maturational delay or arrest implies deprivation of a development promoting factor such as a neuronal signal. Severe involvement with numerous rods was demonstrated in both diaphragm and intercostal muscles at the time of the postmortem examination, compatible with the patient's respiratory failure.

Fatal Outcome↗

PubMedDate not supplied

Vascular involvement in benign infantile mitochondrial myopathy caused by reversible cytochrome c oxidase deficiency.

A 1-month-old Japanese girl had profound generalized weakness, hypotonia, and severe lactic acidosis. The infant improved gradually: she held her head at 9 months, learned to walk by 15 months. At the first muscle biopsy at 11 weeks of age, the specimen was characterized by numerous ragged-red fibers and decreased enzyme activity on cytochrome c oxidase (COX) staining. Electron microscopic findings were characterized by the presence of excessive abnormal mitochondria not only in skeletal muscle fibers but also in blood vessels. Vascular abnormalities consisted of an increased number of enlarged mitochondria in endothelial and smooth muscle cells of small arteries. Biochemical analysis showed an isolated defect of COX activity, which was only 16% of the mean control level. At the second biopsy at 44 months of age, the COX activity had increased to normal in the entire specimen. On electron microscopy, the abnormal mitochondria present on the first biopsy specimen had disappeared both in muscle fibers and blood vessels; nearly all mitochondria were morphologically normal at the second biopsy. Now at 5 years of age she can run and does not show muscle weakness. We report reversibility of abnormal mitochondria with age not only in skeletal muscle fibers but also in blood vessels in a patient, who had reversible COX deficiency with a benign clinical course.

Arteries↗

PubMedDate not supplied

Childhood form of adrenoleukodystrophy in the sucking infant.

The childhood form of adrenoleukodystrophy (ALD) in a sucking infant was discovered before the appearance of clinical symptoms. C24:0/C22:0 and C26:0/C22:0 ratios in umbilical cord plasma were significantly higher so that very long-chain saturated fatty acid-restricted milk was given. Ultrastructural study on biopsied rectal mucosa revealed rectilinear and/or curved electron-lucent clefts in the histiocytes at 3 months of age, which were also recognized in the elder brother and another boy with symptoms of ALD. However, magnetic resonance imaging and T1 and T2 values were within the normal range in cerebral gray matter, cerebral white matter, caudate nucleus, internal capsule, and thalamus.

Adolescent↗

PubMedDate not supplied

Essential role of newly synthesized ATP for cyclic GMP-induced relaxation in alpha-toxin permeabilized smooth muscle of rat proximal colon.

The role of newly synthesized ATP in cyclic GMP-induced relaxation was studied in membrane permeabilized longitudinal muscle preparations of the rat proximal colon. Cyclic GMP and 8 bromo cGMP induced concentration-dependent relaxation of alpha-toxin permeabilized preparations which were precontracted by 3 microM Ca2+ in the presence of 4 mM ATP and 5 mM phosphocreatine (PC). The relaxation by 8 bromo cGMP was inhibited by Rp-8-pCPT cGMPS, an inhibitor of cyclic GMP dependent protein kinase. The relaxation was inhibited by removal of PC from the bathing solution, in spite of the presence of ATP. The relaxation was also inhibited by dinitrofluorobenzene (DNFB), a selective inhibitor of creatine kinase. The removal of PC or treatment with DNFB is known to produce accumulation of ADP within smooth muscle cell, however, ADPbetaS did not affect the relaxation. After irreversible inhibition of endogenous creatine kinase by DNFB in beta-escin permeabilized preparations, treatment of the preparations with exogenous creatine kinase restored the relaxation. In the presence of ADP and PC but without ATP, 8-bromo cGMP induced the relaxation to the similar extent to that in the presence of ATP and PC. These results suggest that ATP newly synthesized from ADP and PC by creatine kinase is essential for cyclic GMP-induced relaxation of the smooth muscle preparations obtained from the proximal colon of rats.

Adenosine Triphosphate↗

PubMedDate not supplied

Advanced adenosquamous carcinoma of the gallbladder with bilio-biliary fistula: an uncommon case treated by hepatopancreatoduodenectomy.

A 70 year-old female, who presented with jaundice and abdominal pain, was found to have an advanced gallbladder cancer involving the liver parenchyma, duodenum, and transverse colon. This was complicated by a bilio-biliary fistula between the gallbladder and both the right and left hepatic ducts. After obtaining an accurate pre-operative diagnosis, the patient underwent hepatopancreatoduodenectomy (HPD) with lymph node dissection around the hepatic pedicle, celiac trunk, aorta, and inferior vena cava. Histologic examination revealed adenosquamous carcinoma. This rare variant accounts for 3.5% of gallbladder cancers, and is associated with a worse prognosis than adenocarcinoma. The patient is in good condition without any signs of recurrence 42 months after the HPD. In this case report, we discuss the histological type and internal biliary fistula with regard to the literature, and the usefulness of an aggressive surgical procedure such as HPD with extended lymph node dissection which can improve survival and quality of life in selected patients.

Aged↗

PubMedDate not supplied

Solution synthesis of Na+,K(+)-ATPase inhibitor-1 (SPAI-1).

SPAI-1, a 49-amino acid peptide including eight Cys residues with Na+,K(+)-ATPase inhibitory activity, was synthesized by the solution procedure. Protecting groups, including the formyl group on the Trp residue, were cleaved simultaneously by HF treatment in the presence of a sufficient amount of thiol compound. After removal of the Acm group on the Cys residue, the resulting octa SH peptide was subjected to an oxidative folding reaction in the presence or absence of redox reagents and/or denaturant. Addition of redox reagents accelerated the reaction but was not crucial to the formation of the correct disulfide bonds in the molecule. The product was purified to homogeneity and found to have the same physicochemical properties and inhibitory potency as those reported for the natural product. Four intramolecularly linked disulfide bridges were assigned as connecting Cys8 to Cys37, Cys15 to Cys41, Cys24 to Cys36, and Cys30 to Cys45 based on results from a combination of enzymatic and synthetic approaches.

Amino Acid Sequence↗

PubMedDate not supplied

Interactions of carteolol and other beta-adrenoceptor blocking agents with serotonin receptor subtypes.

The importance of the anti-serotonergic activity of carteolol and other beta-adrenoceptor blockers in their efficacy as anti-migraine agents has been examined in the membrane fraction of rat brain frontal cortex and pig choroid plexus, using radioligand binding methods. Carteolol and l-propranolol, which are suggested to have anti-migraine activity in man, were found to be active inhibitors of the binding of [125I]ICYP to 5-HT1B recognition sites and of [3H]-8-OH-DPAT to 5-HT1A recognition sites. Carteolol is devoid of activity at 5-HT1C and 5-HT2 recognition sites, whereas l-propranolol shows substantial affinity for these receptor subtypes. Atenolol, another beta-adrenoceptor blocker with anti-migraine activity, is devoid of activity at any of the 5-HT receptor subtypes examined. The possibility that carteolol and other beta-adrenoceptor antagonists exert their pharmacological effects through central 5-HT receptor subtypes is discussed in relation to the potential mechanism of the anti-migraine activity of carteolol.

8-Hydroxy-2-(di-n-propylamino)tetralin↗

PubMedDate not supplied

Intra-arterial digital subtraction portography with a blood-isotonic, non-ionic, dimeric contrast medium.

Intra-arterial digital subtraction portography (IA-DSP) with a blood-isotonic, non-ionic, dimeric contrast medium was carried out in 27 patients with hepatocellular carcinoma. It was possible to obtain images of excellent or good quality of the portal vein and its bilateral main branches in all patients. The third-order branches of the portal vein in the right lobe could be identified in all patients, and images of excellent or good quality were obtained in a mean of 80.2% of patients. Images of third-order branches in the left lobe were of lower quality than those of third-order branches in the right lobe; in particular, images obtained were of poor quality for 27.3% of the medial branches of the left lobe. It was impossible to identify the caudal branches in almost all patients. The side effects of IA-DSP, pain and sensations of heat were very mild: only one patient complained of mild pain, while 18 patients (69.2%) complained of no sensations of heat whatsoever.

Aged↗

PubMedDate not supplied

Synthesis of an O-palmitoylated 44-residue peptide amide (PLTX II) blocking presynaptic calcium channels in Drosophila.

PLTX II, a presynaptic calcium channel blocker in Drosophila isolated from the plectreurys spider venom, is a 44-residue peptide containing ten Cys residues and an O-palmitoylated threonine amide at the carboxy-terminus. In this study, the palmitoylated peptide was synthesized in solution by applying our maximum protection strategy using the HF method at the final deprotecting step. Before designing the synthesis, we examined the stability of the palmitoyl moiety under the conditions for the synthesis of the peptide using several model peptides. The O-palmitoyl group was confirmed to be stable during elongation of the peptide bonds, but was partially removable during the deprotection reaction in HF. The depalmitoylation reaction in HF was temperature- and time-dependent. Therefore, the decision was made to protect the Asp residues with benzyl ester, since it is more susceptible to HF than cyclohexyl ester, which is now commonly used in the Boc-based, solid-phase synthesis. Thus, the HF reaction was carried out at -10 degrees or -15 degrees C for 1 h in order to reduce the extent of the depalmitoylation reaction. The resulting palmitoylated and depalmitoylated products were separated, the remaining Acm groups were removed using Hg(OAc)2, and then the completely deprotected peptides were folded to their native forms. The final palmitoylated peptide was proven to be identical with the natural one using various HPLC systems and by bioassay.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗