Biomedical subjects
Publications and source records attributed to M Maeda.
A patient with erythema on the dorsum of the left foot after the removal of gypsum fixation applied for the treatment of bone fracture is reported. Both the clinical and histopathologic features led to the diagnosis of mucor infection.
The unstable hemoglobin disease result from the presence of a structurally abnormal hemoglobin variant with substitution or deletion of amino acid in the red cell. Characteristic of the unstable hemoglobin disease is the presence of inclusions of precipitated denatured hemoglobin called Heinz bodies in the red cells. Almost 200 unstable hemoglobins have now been identified. Hundred of these hemoglobins arise hemolysis or abnormal oxygen affinity, and another 100 hemoglobins have no hematological abnormalities but with instability on in vitro test. The inheritance pattern of the unstable hemoglobin is autosomal dominant. The most common causes of instability re amino acid substitutions that disrupt the alpha-helix of globin or alter the steric configuration of the hem pocket by the insertion of polar amino acid in the interior of the hemoglobin molecule, or deletion of amino acid in the helix. The unstable hemoglobins have a greater tendency to be spontaneously oxidized to methemoglobin with subsequent formation of hemichromes and precipitations.
Perioperative airway management for sleeve pneumonectomy in a 66-year-old female with tuberculous tracheal stenosis which was 2 cm above the carina to the right main bronchial orifice is reported. Endotracheal intubation was required, because she complained of dyspnea due to airway stenosis in preoperative period for anti-tuberculous chemotherapy. A special tracheal tube developed for laryngomicrosurgery (MLT tube, Mallinckrodt Co. Ltd., 21 Fr size) was used in this patient. However, 13 days after intubation with an MLT tube, sleeve pneumonectomy was done because because of right lung atelectasis and progressive hypoxemia. Left one lung ventilation was successful by an MLT tube during right thoracotomy and a spiral tube (28 Fr) was inserted to the left main bronchus in the operative field during the resection of the carina. For this type of patient with tracheo-bronchial stenosis, it is essential to select an endotracheal tube with appropriate length and diameter.
The discriminant function (Z) for predicting postoperative performance status in patients with giant bulla was addressed in our previous paper. In the present study, patients with dyspnea were classified into Group 1 or Group 2 based on preoperative function, Group 1 showing continuous improvement in dyspnea and Group 2 unchanged or worsened condition after bullectomy. Of the 47 patients in this study, 28 had dyspnea of grade 2 or more, 19 revealing no symptoms prebullectomy. The group predictions for the 28 patients were compared with the postoperative status in dyspnea for over four years following surgery. The predicted grouping in 26 of the 28 (93%) agreed with the postoperative status but in two it did not: one was familial bullous emphysema the other had repeated episodes of pneumonia, both of which were predicted for Group 1. All 19 patients without preoperative dyspnea were studied their symptoms and lung functions before and after bullectomy. After surgery, none showed dyspnea and significant changes of functions. As to preoperative pulmonary function, patients with FEV1.0% of more than 60% and delta N2 of less than 2% were improved, the prediction agreeing with the actual results. Fourteen patients with FEV1.0% of less than 55% showed high delta N2. When delta N2 exceeded 3.5%, deterioration of dyspnea was observed following surgery. Bullectomy is indicated in patients with low pulmonary function, by preoperative FEV1.0% and delta N2.
Thirty-two (1994) and 55 (1995) patients in the terminal stage of many kinds of cancer were cared for satisfactorily at home by a visiting team of nurses from Niigata Cancer Center Hospital. A prostate cancer patient with multiple bone metastasis was suffering from morphine-resistant intractable pain, and highly dependent on medical care. We continued to take care of him by frequent visitation and psychological assist until he eventually died at home according to his wishes. It is important to create a suitable support team for the continual difficult home care of each terminal patient.
We previously isolated a novel protein having two C2-like domains known to interact with Ca2+ and phospholipid, and named Doc2 (Double C2). Doc2 is predominantly expressed in brain and is implicated in Ca(2+)-dependent neurotransmitter release. We have isolated here an isoform of Doc2 and named the original one Doc2 alpha and the new one Doc2 beta. Doc2 beta alsp has two C2-like domains and is 61% identical to Doc2 alpha at the amino acid level. In contrast to Doc2 alpha, the Doc2 beta mRNA is expressed ubiquitously. These results indicate that there are at least two isoforms of Doc2, and suggest that Doc2 beta is involved in Ca(2+)-dependent intracellular vesicle trafficking in various types of cells.
Glu-beta 185 of the Escherichia coli H(+)-ATPase (ATP synthase) beta subunit was replaced by 19 different amino acid residues. The rates of multisite (steady state) catalysis of all the mutant membrane ATPases except Asp- beta 185 were less than 0.2% of the wild type one; the Asp- beta 185 enzyme exhibited 15% (purified) and 16% (membrane-bound) ATPase activity. The purified inactive Cys- beta 185 F1-ATPase recovered substantial activity after treatment with iodoacetate in the presence of MgCl2; maximal activity was obtained upon the introduction of about 3 mol of carboxymethyl residues/mol of F1. The divalent cation dependences of the S-carboxymethyl- beta 185 and Asp- beta 185 ATPase activities were altered from that of the wild type. The Asp- beta 185, Cys- beta 185, S-carboxymethyl-beta 185, and Gln- beta 185 enzymes showed about 130, 60, 20, and 50% of the wild type unisite catalysis rates, respectively. The S-carboxymethyl- beta 185 and Asp- beta 185 enzymes showed altered divalent cation sensitivities, and the S-carboxymethyl- beta 185 enzyme showed no Mg2+ inhibition. Unlike the wild type, the two mutant enzymes showed low sensitivities to azide, which stabilizes the enzyme Mg-ADP complex. These results suggest that Glu- beta 185 may form a Mg2+ binding site, and its carboxyl moiety is essential for catalytic cooperativity. Consistent with this model, the bovine glutamate residue corresponding to Glu- beta 185 is located close to the catalytic site in the higher order structure (Abrahams, J.P., Leslie, A.G.W., Lutter, R ., and Walker, J.E. (1994) Nature 370, 621-628)
In Xenopus laevis, the spermatogenic cells derived from a primary spermatogonium divide synchronously and form a cyst surrounded by somatic Sertoli cells. To clarify how many times the spermatogonia divide mitotically before differentiating to primary spermatocytes, the number of spermatogenic cells in the cysts was counted. Each cyst which consisted of cells morphologically identical to secondary spermatogonia contained discrete numbers of cells such as 2(6), 2(7), or 2(8) cells. On the other hand, the number of the germ cells in a cyst of zygotene/pachytene primary spermatocytes was approximately 2(8). These results indicate that spermatogenic cells which had finished eight mitotic divisions and showed a similar morphology to secondary spermatogonia were preleptotene spermatocytes. The nuclei of the preleptotene spermatocytes increased in volume concomitantly with premeiotic DNA replication and differentiation into zygotene/pachytene primary spermatocytes. Therefore the cells morphologically indistinguishable from secondary spermatogonia but with larger nuclei than typical secondary spermatogonia were identified as preleptotene spermatocytes. Electron microscopic observations of the preleptotene spermatocytes identified as above showed that flattened vesicles, 0.1-0.5 micron in length, about 0.05 micron in width, and with more electron-dense membrane than typical endoplasmic reticulum, first appeared in the cytoplasmic area of preleptotene spermatocytes.
The mechanisms of energy coupling and catalytic co-operativity are not yet understood for H(+)-ATPase (ATP synthase). An Escherichia coli gamma subunit frameshift mutant (downstream of Thr-gamma 277) could not grow by oxidative phosphorylation because both mechanisms were defective (Iwamoto, A., Miki, J., Maeda, M., and Futai, M. (1990) J. Biol. Chem. 265, 5043-5048). The defect(s) of the gamma frameshift was obvious, because the mutant subunit had a carboxyl terminus comprising 16 residues different from those in the wild type. However, in this study, we surprisingly found that an Arg-beta 52-->Cys or Gly-beta 150-->Asp replacement could suppress the deleterious effects of the gamma frameshift. The membranes of the two mutants (gamma frameshift/Cys-beta 52 with or without a third mutation, Val-beta 77-->Ala) exhibited increased oxidative phosphorylation, together with 70-100% of the wild type ATPase activity. Similarly, the gamma frameshift/Asp-beta 150 mutant could grow by oxidative phosphorylation, although this mutant had low membrane ATPase activity. These results suggest that the beta subunit mutation suppressed the defects of catalytic cooperativity and/or energy coupling in the gamma mutant, consistent with the notion that conformational transmission between the two subunits is pertinent for this enzyme.
The protein product of the myotonic dystrophy (DM) gene is a putative serine-threonine protein kinase (DM kinase). Previous reports have characterized the DM gene product as various 50-62-kDa proteins. The predicted protein size from DM cDNA sequence is 69 kDa. We therefore expressed a full-length recombinant human DM kinase protein and compared its size and expression to heart, cardiac Purkinje fibers, and skeletal muscle from normal and DM subjects. Recombinantly expressed DM kinase and endogenous DM kinase in human heart, displayed two immunoreactive DM kinase proteins with apparent molecular sizes of 71 and 80 kDa, suggesting that these prior reports are incorrect. In cardiac Purkinje fibers the 71-kDa protein was the major form, and in skeletal muscle the 80-kDa protein was the major form. Immunostaining showed DM kinase localized to neuromuscular junctions in skeletal muscle and intercalated discs in heart and Purkinje fibers. DM subjects showed low abundance of DM kinase in heart and skeletal muscle, suggesting haplotype insufficiency as a potential mechanism for disease expression. These studies describe differential expression of two protein forms of DM kinase, which are localized to specialized cellular structures associated with impulse transmission.
BACKGROUND: Fabry's disease is considered very rare. Left ventricular hypertrophy is one of the common manifestations in adults with classic hemizygous disease. Recently, several cases of an atypical variant of hemizygous Fabry's disease, with manifestations limited to the heart, have been reported. Therefore, we assessed the incidence of hemizygosity for Fabry's disease among male patients with left ventricular hypertrophy. METHODS: We measured plasma alpha-galactosidase activity in 230 consecutive male patients with left ventricular hypertrophy. Clinical manifestations were assessed, endomyocardial biopsies were performed, and the patients were screened for mutations in the alpha-galactosidase gene. RESULTS: Seven of the 230 patients with left ventricular hypertrophy (3 percent) had low plasma alpha-galactosidase activity (0.4 to 1.2 nmol per hour per milliliter; 4 to 14 percent of the mean value in normal controls). These seven unrelated patients, ranging in age from 55 to 72 years, did not have angiokeratoma, acroparesthesias, hypohidrosis, or corneal opacities, which are typical manifestations of Fabry's disease. Endomyocardial biopsy was performed in five patients and revealed marked sarcoplasmic vacuolization in all five. Samples from four patients were examined by electron microscopy and revealed typical lysosomal inclusions with a concentric lamellar configuration in all four. Two patients had novel missense mutations in exon 1 (Ala20Pro) and exon 6 (Met296lle). The remaining five had no mutations in the coding region of the alpha-galactosidase gene, but the amounts of the alpha-galactosidase messenger RNA were markedly lower than normal. CONCLUSIONS: Seven unrelated patients with atypical variants of hemizygous Fabry's disease were found among 230 men with left ventricular hypertrophy (3 percent). Fabry's disease should be considered as a cause of unexplained left ventricular hypertrophy.
A new animal model of cerebral infarction was developed by magnetic embolization with carbonyl iron particles. An electromagnet was placed upon the lateral portion outside the orbit of Mongolian gerbils and charged with a current intensity of 2 A, 3 V (400 G) for 10 min. By intracardiac injection of iron particles at the beginning of the electric charge, infarction was selectively produced in the cerebral cortex. Two days after the operation, microhemorrhaging was seen in the necrotic region. Multiple scattered infarctions accompanied with microhemorrhagic signs closely resembled embolic infarction encountered in the human brain.
The human histamine H2 receptor gene (a 13 kb 5'-upstream sequence, the 1,077 bp coding region and a 245 bp 3'-downstream sequence) was cloned and its 5'-upstream 1.8 kb nucleotide sequence was determined. Northern blot hybridization showed that the message for the receptor was transcribed in human gastric adenocarcinoma MKN45 cells. When the upstream sequence was ligated in front of the luciferase gene and the construct was introduced into this cell line, the reporter gene was effectively transcribed, as demonstrated by the expression of enzyme activity. The minimum promoter was located in the region between -610 and -278. This sequence contained AP2 sites and GATA motifs, but not a TATA-box like sequence. Further upstream regions (-1,202 approximately -611 bp and -1,773 approximately -1,203 bp) stimulated and inhibited luciferase gene expression, respectively. These regions may be important for modulation of the mRNA level in cells expressing the H2 receptor.
Histochemical study indicated that the posterior pituitary possesses numerous microvesicles (MVs) containing synaptophysin, a marker protein specific for brain synaptic vesicles (Navone, F., Di Gioia, G., Jahn, R., Browning, M., Greengard, P., and De Camilli, P. (1989) J. Cell Biol. 109, 3425-2433). By monitoring cross-reactivity with anti-synaptophysin antibody, the MVs were highly purified from bovine posterior pituitaries by a combination of differential and sucrose density gradient centrifugations. The purified MVs had an average diameter of about 60 nm and were associated with synaptophysin as revealed by immunoelectron microscopy. The vesicles contained ATPase activity partially sensitive to bafilomycin A1 and to vanadate. The membrane fraction immunoisolated with anti-synaptophysin antibody also exhibited similar ATPase activity. The two ATPases could be purified separately; the vandate-sensitive enzyme was identified as a 115-kDa polypeptide immunochemically similar to chromaffin granule P-ATPase (forming phosphoenzyme intermediate), and the bafilomycin A1-sensitive ATPase showed essentially the same properties as those of vacuolar type H(+)-ATPases. Upon addition of ATP, the MVs formed an electrochemical gradient of protons and took up norepinephrine in a reserpine-sensitive manner, indicating the presence of secondary monoamine transporter coupled with vacuolar type H(+)-ATPase. No uptake of L-glutamate, gamma-aminobutyrate, glycine, or acetylcholine was observed. The identification of MVs as organelles responsible for storage of monoamines is important for understanding the physiological function of the posterior pituitary.
The reaction mechanism of bacteriophage T4 endonuclease V was investigated using modified oligodeoxyribonucleotide duplexes containing a cis-syn thymine dimer. For the pyrimidine dimer glycosylase step, the formation of a covalent intermediate has been proposed. A fluorine atom was attached to the 2'-position of the 5'-component of the thymine dimer site, which could stabilize the covalent complex and prevent the ring opening of the sugar moiety. The strand cleavage of the 12 base pair substrate analog did not occur, although the glycosyl bond was cleaved by this enzyme. A covalent enzyme--substrate complex was separated by gel electrophoresis under denaturing conditions. It was shown that the enzyme molecules were completely converted to a stable complex in the reaction mixture. Two mechanisms have been proposed for the beta-elimination step. A 12-mer containing a phosphorothioate linkage between adjacent thymidines was prepared. The diastereomers were separated, and the absolute configurations were determined. After formation of the thymine dimer and 32P-labeling of the 5'-terminus, these oligonucleotides were annealed to the complementary 12-mer, and the reaction rates of the pyrimidine dimer glycosylase step and the overall reaction for each duplex were measured under the substrate-saturation conditions. The rate constants indicated that the chemical reaction at the beta-elimination step was rate-limiting. Since no difference was observed in the rate constants for the Rp- and Sp-phosphorothioate substrates, it is concluded that the beta-elimination reaction is catalyzed, not by the internucleotide phosphate, but by an amino acid residue of the enzyme.
L-Glutamate was microinjected into the nucleus tractus solitarii (NTS) in anesthetized (chloralose and urethane), paralyzed and artificially ventilated rats, and spinal cord blood flow (SCBF) was determined using a combination of labeled microspheres. Unilateral chemical stimulation of the NTS (n = 13) significantly decreased SCBF in the cervical cord from 43 +/- 6 (mean +/- SEM) to 28 +/- 4 (P < 0.05), in the thoracic cord from 35 +/- 3 to 24 +/- 4 (P < 0.01), and in the lumbar cord from 49 +/- 3 to 40 +/- 3 ml min-1 (100 g)-1 (P < 0.05). The decrease in SCBF was not due to the decrease in arterial blood pressure (ABP) because the SCBF during the chemical stimulation of the NTS was significantly smaller (P < 0.05) than the SCBF during controlled hemorrhagic hypotension (n = 11). Chemical stimulation of the NTS did not affect the reactivity of the spinal cord vessels to hypercapnia (n = 5). Microinjection of the vehicle solution into the NTS had no effects on spinal cord circulation (n = 9). These results suggest that the cell bodies within the NTS may play a role in the control of spinal cord circulation.
Using magnetic resonance (MR) diffusion-weighted method, we examined the optic and the trigeminal nerves of jimpy and twitcher mice, considered to be animal models of Pelizaeus-Merzbacher disease, hypomyelination disorder, and Krabbe disease, demyelination disorder, respectively. In jimpy mice, diffusional anisotropy of optic nerve did not show a significant difference compared to age-matched control mice, suggesting that diffusional anisotropy does exist in absence of multiple layers of myelin sheath. In twitcher mice, diffusional anisotropy was attenuated remarkably in the optic and trigeminal nerves. Loss of axonal straightness on longitudinal section confirmed by electron microscopy appeared to be the principal explanation for it. It is further suggested that this MR diffusion-weighted imaging method enables us to differentiate hypomyelination from demyelination in vivo.
Two repeated C2-like domains interacting with Ca2+ and phospholipid are found in synaptotagmin and Rabphilin-3A which are implicated in neurotransmitter release. Here we have isolated a cDNA encoding a novel protein having two repeated C2-like domains from a human brain cDNA library. The isolated cDNA encodes a protein with 400 amino acids and a M(r) of 44,071. The purified recombinant protein indeed interacts with Ca2+ and phospholipid. We have named this protein Doc2 (Double C2). Doc2 is exclusively expressed in brain and is highly concentrated in the synaptic vesicle fraction. These results suggest that Doc2 is a novel brain protein and serves as a Ca2+ sensor in neurotransmitter release.