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M Yoshida

Publications and source records attributed to M Yoshida.

At least 361 records · Page 20Linked to original sources
PubMed2000

Effect of electrostatic interactions on the binding of charged substrate to GroEL studied by highly sensitive fluorescence correlation spectroscopy.

The binding processes of GroEL with apo cytochrome c (apo-cyt c) and disulfide-reduced apo alpha-lactalbumin (rLA) in homogeneous solution at low concentration were analyzed by fluorescence correlation spectroscopy (FCS) with extremely high sensitivity. Although apo-cyt c, a positively charged substrate, was tightly bound to GroEL in both the absence and the presence of 200 mM KCl, the strength of the binding was changed with varying salt concentration. Results from experiments when two different salts (KCl or MgCl(2)) were titrated into a sample solution containing GroEL and apo-cyt c clearly showed that the binding strength decreased with increasing salt concentration. On the other hand, the binding affinity of GroEL for rLA, a negatively charged substrate, increased by adding of 200 mM KCl. These results indicate that electrostatic interactions substantially contribute to the binding interactions by manipulating the binding affinity of charged substrates.

Animals↗
PubMed2000

In vitro synthesis of hyaluronan by a single protein derived from mouse HAS1 gene and characterization of amino acid residues essential for the activity.

HAS1 was expressed as a FLAG-tagged HAS1 fusion protein in COS-1 cells. This recombinant protein was extracted with CHAPS (3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acid) from the membrane fraction and purified by anti-FLAG affinity chromatography and subsequent SDS-polyacrylamide gel electrophoresis. A protein solubilized from the one single band on the gel was able to synthesize hyaluronan when incubated with UDP-GlcNAc and UDP-GlcA as donor substrates without any further additions. The detergent-solubilized and purified HAS1 protein, however, exhibited quite different kinetic properties from the membrane-bound protein. When assayed under the reconstitutive conditions where the reaction mixture was layered onto the buffer containing high concentration of CHAPS, the activity was enhanced and the kinetic properties became similar to those of the membrane-bound protein. In addition, a HAS1 gene product by an in vitro transcription/translation system also showed HAS1 activity under the reconstitutive conditions. To our surprise, when incubated with UDP-GlcNAc alone, the protein was found to synthesize chito-oligosaccharide. Taking advantage of these enzyme reaction properties, active sites on the protein involved in for hyaluronan and chito-oligosaccharide synthesis were characterized. Site-directed mutagenesis induced in the cytoplasmic central loop domain of the protein revealed that several amino acid residues conserved among those domains of various proteins of a HAS family were essential for both hyaluronan and chito-oligosaccharide syntheses but one of them was not for chito-oligosaccharide synthesis. The substitutions that caused partial or severe loss of the activity gave no significant changes of the K(m) values of the mutated proteins, suggesting that no conformational or other indirect changes were involved in the effect. Taken together, the results suggest that the HAS1 protein alone is able to synthesize hyaluronan and different amino acid residues on the cytoplasmic central loop domain are involved in transferring GlcNAc and GlcA residues, respectively.

Amino Acid Motifs↗
PubMed2000

Frequent genetic heterogeneity in the clonal evolution of gynecological carcinosarcoma and its influence on phenotypic diversity.

Carcinosarcomas of the uterus, ovaries, and fallopian tubes are highly aggressive neoplasms with incompletely understood histogenesis. Although recent immunohistochemical, cell culture, and molecular genetic studies all favor these cancers to be monoclonal in origin, the extent of intratumoral genetic heterogeneity in these tumors with divergent histology has not been reported previously. For this study, we microdissected a total of 172 carcinomatous or sarcomatous foci from 17 gynecological carcinosarcomas and analyzed allelic status with 41 microsatellite markers on chromosomal arms 1p, 1q, 3p, 4q, 5q, 6q, 8p, 9p, 10q, 11p, 11q, 13q, 16q, 17p, 17q, 18q, and 22q. With the exception of a single case with microsatellite instability, we found shared allelic losses and retentions among multiple individually dissected foci of each case, strongly supportive of the concept of a monoclonal origin for these neoplasms. In eight of these cases, we also found heterogeneous patterns of allelic loss at limited numbers of chromosomal loci in either the carcinomatous or sarcomatous components of the neoplasms. These heterogeneous patterns of allelic losses were consistent with either genetic progression or genetic diversion occurring during the clonal evolution of these neoplasms. In two cases, we found the specific patterns of genetic progression to be consistent with sarcomatous components of the neoplasms arising from carcinomatous components. We conclude that most of the gynecological carcinosarcomas have a monoclonal origin, and that genetic progression and diversion parallel the development of divergent phenotypes in these tumors. Because phenotypically divergent areas of the tumors share numerous genetic alterations, this divergence most likely occurs relatively late in the evolution of these tumors.

Carcinosarcoma↗
PubMed2000

Macular degeneration associated with aberrant expansion of trinucleotide repeat of the SCA7 gene in 2 Japanese families.

OBJECTIVE: To evaluate the macular function of Japanese patients with a trinucleotide repeat expansion in the spinocerebellar ataxia type 7 (SCA7) gene. METHODS: Ophthalmic findings in patients whose DNA analysis revealed expanded alleles of the trinucleotide repeat in the SCA7 gene were evaluated. RESULTS: Trinucleotide repeat was expanded from 40 to 48 in affected patients (control subjects, 12 repeats). Affected patients were characterized by different degrees of visual acuity decrease (0.09-0.9), a tritan axis color vision, a coarse granular appearance of the macular region on scanning laser ophthalmoscopy, depression of multifocal electroretinograms, and macular degeneration. However, pigmentary changes were not observed in the retina. The trinucleotide repeat was longer and the onset of macular dysfunction was earlier in the younger generation. One patient in a family manifested decreased visual acuity 10 years preceding other neurologic signs. CONCLUSIONS AND CLINICAL RELEVANCE: Patients with SCA7 mutations showed macular dysfunction or degeneration with expansion of CAG repeat in the SCA7 gene. However, the lesions were less pigmented than those previously reported. Patients also showed ophthalmologic anticipation, which has not been reported for the ocular changes in other patients who have trinucleotide repeat expansion of the responsible genes.

Adult↗
PubMed2000

Monitoring of human herpesvirus-6 and -7 genomes in saliva samples of healthy adults by competitive quantitative PCR.

Human herpesviruses-6 and -7 (HHV-6 and HHV-7) are thought to be transmitted during early infancy through saliva. However, the kinetics of the virus shedding in saliva of healthy adults, from whom children are assumed to acquire the viruses, is mostly unknown. This study was conducted to determine how many copies of the genome are secreted in saliva of healthy adults and to clarify the relationship between viral DNA load and virus isolation of HHV-6 and HHV-7. Competitive PCR was performed using primer sets in the U42 gene of each viral genome. In saliva samples from 29 healthy adults, HHV-6 and HHV-7 DNA was detected in 41.4% and 89.7%, respectively. The average copy number of the HHV-7 genome in the positive samples was higher than that of the HHV-6 genome. Follow-up studies of six seropositive individuals for 3 months showed that the amount of HHV-7 DNA was constant in each individual and that "high producers" and "low producers" could be distinguished. By contrast, the amount of HHV-6 DNA varied drastically over time in each individual. Although HHV-6 was never isolated from the saliva of any of the six individuals during the follow-up period, HHV-7 was isolated from each individual several times. The amount of HHV-7 DNA tended to be higher at the times when the virus was isolated than at the times when the virus was not isolated. These data demonstrate a striking contrast between HHV-6 and HHV-7 in the kinetics of genome and virus shedding.

Adult↗
PubMed2000

CD44 is a potential target of amplification within the 11p13 amplicon detected in gastric cancer cell lines.

Classical cytogenetic approaches have revealed many of the chromosomal aberrations that may occur in gastric cancers (GC), although few alterations of specific genes have been identified so far. Genes that affect progression of this disease need to be identified if clinicians are to achieve optimal management of patients with GC. As the first step toward the cloning of gene(s) that may be involved in gastric carcinogenesis, we examined 25 GC cell lines for aberrations in DNA copy number to detect chromosomal gains and losses, as well as gene amplifications, by comparative genomic hybridization (CGH). Our CGH study revealed high-level amplifications in chromosomal regions that had been well defined in GC but also in sites that had not, including 3p24, 5p15, 11p11.2-14, 13q34, 15q26, Xp24, and Xq26-28. The minimal common region at 11p13, within the 11p11.2-14 amplicon, harbors the CD44 gene. Northern and Western blot analyses showed that an alternatively spliced form of CD44, with variant exons 8-10 (CD44E), was overexpressed in all cell lines bearing the 11p13 amplicon. However, cell adhesion activity was no greater in these lines than in cell lines without amplifications at the CD44 locus, suggesting that the major property of upregulated CD44 in these cases might be to transduce signals critically associated with growth and proliferation of the tumor cells.

Blotting, Northern↗
PubMed2000

Chiral discrimination of N-carbazole-carbonyl derivatives of alpha-amino acids with a short linear alkyl side chain by bovine serum albumin.

Chiral discrimination of racemic carbazole carbonyl (CC)-amino acids with linear alkyl sidechain (C(1)-C(4)) by bovine serum albumin (BSA) was investigated by competitive replacement experiments using dansyl-L-proline and dansyl-D-norvaline as fluorescent probes. It was found that the CC derivatives of the D-forms of alanine (C(1)), amino butyric acid (C(2)), norvaline (C(3)), and norleucine (C(4)) bound to the dansyl-L-proline site much more strongly than their L-forms, whereas the interactions between both enantiomers of these amino acids with dansyl-D-norvaline site were slight.

Alanine↗
PubMed2000

Brainstem-type Lewy body disease presenting with progressive autonomic failure and lethargy.

The authors report an autopsy case characterized by progressive lethargy and autonomic failure with a distinctive pattern of occurrence of Lewy bodies. Autonomic dysfunction such as sleep apnea, orthostatic hypotension, dysuria, and hypohidrosis predominated with lethargy, whereas parkinsonism was not apparent. Numerous Lewy bodies were widely evident microscopically in brainstem nuclei and the intermediolateral cell columns of the spinal cord, as well as in the sympathetic ganglia, but were rare or absent in the cerebral cortex and other supratentorial structures. Marked neuronal loss was seen in the locus ceruleus, raphe nuclei, dorsal vagal nuclei, and intermediolateral cell columns, but neurons in the substantia nigra, other brain regions, and sympathetic ganglia appeared undiminished. This case represents a specific clinicopathologic form of Lewy body disease occurring predominantly in the brainstem, spinal cord, and sympathetic ganglia.

Autonomic Nervous System↗
PubMed2000

Mitral valve plasty in systemic lupus erythematosus in the setting of antiphospholipid syndrome.

The first case was of a 27-year-old female, who was diagnosed as having mitral valve stenosis with regurgitation, systemic lupus erythematosus and antiphospholipid syndrome at her previous pregnancy. We performed mitral valve plasty, which included open mitral commissurotomy and Kay's annulo plasty. The second case was of a 53-year-old female, who was diagnosed as old myocardial infarction, mitral regurgitation, systemic lupus erythematosus and antiphospholipid syndrome. She underwent mitral valve plasty and coronary artery bypass grafting. Both cases were treated by administration of methylpredonisolone and heparin perioperatively to avoid thrombosis and aggravation of systemic lupus erythematosus. Both patients showed good postoperative outcome without complications. We consider that it is important to perform the plasty as far as possible, and to administer effective anticoagulation treatment to prevent complications for patients in the setting of systemic lupus erythematosus and antiphospholipid syndrome.

Adult↗
PubMed2000

MELAS with the mitochondrial DNA 3243 point mutation: a neuropathological study.

We performed a neuropathological examination of the central nervous system from seven autopsied patients with mitochondrial myopathy, encephalopathy, lactic acidosis and stroke-like episodes (MELAS). Five of the seven cases were confirmed to have the mitochondrial DNA (mtDNA) 3243 point mutation. In addition to the changes reported previously, diffuse atrophy of the cerebral and cerebellar cortices, diffuse gliosis of cerebral and cerebellar white matter, and cactus formation of Purkinje cells were observed. Electron microscopy revealed accumulation of mitochondria in the cactus formations. These lesions are common in MELAS with the mtDNA 3243 point mutation, but cannot be explained solely by mitochondrial angiopathy, and suggest that intrinsic mitochondrial malfunction contributes to neuronal damage in MELAS pathology. Moreover, the pathological changes observed in the cerebellum suggest that cerebellar function should be evaluated more carefully at the clinical level.

Adult↗
PubMed2000

Effects of ovarian hormones on beta-adrenergic receptor-mediated relaxation in the female rabbit bladder.

The effects of ovarian hormones on beta-adrenergic receptor-mediated responses in female rabbit detrusor smooth muscles were investigated. Ovariectomized mature female New Zealand white rabbits were untreated or treated with estrogen and/or progesterone for 2 weeks. The contractile responses to carbachol and KCl in the detrusor strips were not significantly different in all groups. As compared with dobutamine and GS-332, isoproterenol and procaterol significantly relaxed the detrusor strips derived from all groups on KCl-induced tonic contractions. Combined with estrogen treatment, isoproterenol, procaterol and GS-332 caused a significant increase in this muscle relaxation. Furthermore, estrogen treatment caused a significant increase in relaxation as a result of forskolin and the cyclic AMP (cAMP) production that was induced by isoproterenol, procaterol and GS-332. However, estrogen treatment did not affect the relaxant response to dibutyryl cyclic AMP. Progesterone treatment did not affect beta-adrenergic receptor-mediated responses. These results suggest that estrogen treatment causes the increased relaxant responses mediated by beta2- and beta3-adrenergic receptor subtypes, which may be related to the increased cAMP content in female rabbit detrusor smooth muscles.

Adrenergic beta-Agonists↗
PubMed2000

Morphological Purkinje cell changes in spinocerebellar ataxia type 6.

Spinocerebellar ataxia type 6 (SCA6) was recently identified as a form of autosomal dominant spinocerebellar ataxia associated with a small CAG repeat expansion of the gene encoding an alpha 1 A-voltage-dependent calcium channel gene subunit on chromosome 19p13. In this study 50-microm-thick sections of cerebellar tissue from one patient with SCA6 were subjected to free-floating immunohistochemical staining with calbindin-D and parvalbumin antibodies. Severe loss of Purkinje cells was found, particularly in the vermis, and various morphological changes in Purkinje cells and their dendritic arborizations were demonstrated. Many of the remaining Purkinje cells were found to have heterotopic, irregularly shaped nuclei, an unclear cytoplasmic membrane outline, and somatic sprouts. Increased numbers of spine-like protrusions from swelling dendritic arborizations were found in the molecular layer. The axonal arrangement was disordered, and many torpedos were found in the granular layer and white matters. These morphological changes are completely different from those observed in paraneoplastic cerebellar degeneration (PCD) and multiple system atrophy (MSA) and are considered to be related to the genetic abnormality that causes abnormal development of Purkinje cells.

Aged↗
PubMed2000

Angiotensin-converting enzyme activity by canine pulmonary microvascular and central pulmonary artery endothelial cells exposed to hypoxia.

To compare the amount of angiotensin-converting enzyme (ACE) activity in pulmonary artery endothelial cells from different sites and to examine the effect of severe hypoxia (less than 1% of O(2) in 5% CO(2) and 95% N(2)) on the ACE activity expressed by these cells, endothelial cells were harvested and cultured from canine main pulmonary artery by scraping the luminal surface of the artery and from canine pulmonary artery microvessels by infusing chilled buffer with microcarrier beads and 0.02% ethylenediamine tetraacetic acid (EDTA). ACE activity in cell lysates and culture medium was evaluated by fluorometric assay with hippuryl-L-histidyl-L-leucine as a substrate. ACE activity in cell lysates and postculture medium of pulmonary microvascular endothelial cells (PMVEC) was higher than in cell lysates and culture medium of central pulmonary artery endothelial cells (PAEC). However, hypoxia suppressed cellular ACE activity in both PAEC and PMVEC. The degree of suppression of ACE activity by hypoxia, which was determined as (ACE activity in normoxia - ACE activity in hypoxia)/ACE activity in normoxia x 100(%), was larger in PMVEC than in PAEC. The pulmonary microvasculature may be a greater source of ACE than central pulmonary artery, and the ACE activity of pulmonary microvascular endothelial cells seem to be sensitive to hypoxia, although the small diameter of the vessels improves conditions for interaction of blood-borne substance with endothelial enzymes.

Acute Disease↗
PubMed2000

Differential somatic CAG repeat instability in variable brain cell lineage in dentatorubral pallidoluysian atrophy (DRPLA): a laser-captured microdissection (LCM)-based analysis.

Employing a laser-captured microdissection (LCM), we have investigated the somatic instability of CAG repeats in the variable brain cell lineage in three patients with dentatorubral pallidoluysian atrophy (DRPLA). LCM enables the isolation of single lineage brain cells for subsequent molecular analysis. We have found that CAG repeat size and the range of CAG repeats in the cerebellar granular cells is smaller than those in cerebellar glial cells. Similarly, those in the cerebral neuronal cells are significantly shorter than those in cerebral glial cells. These data directly indicate that the CAG repeat is relatively more stable in neuronal cells than in glial cells. Furthermore, cerebellar granular cells show significantly smaller main CAG repeat size and CAG repeat range than either Purkinje cells or cerebral neuronal cells, suggesting that somatic instability in the CAG repeat is markedly variable even among the different types of neuronal populations. The cell-specific CAG repeat instability may thus be more complex than has previously been considered. LCM is a powerful tool for elucidating the mechanism of the triplet repeat instability of each cell type.

Adult↗
PubMed2000

Limited pancreatectomy: significance of postoperative maintenance of pancreatic exocrine function.

This study attempted to clarify whether limited pancreatectomy (duodenum-preserving total pancreatic head resection [DPTPHR], or medial pancreatectomy [MP], maintain pancreatic exocrine function more than conventional pancreaticoduodenectomy (Whipple) or pylorus-preserving pancreaticoduodenectomy (PPPD). A total of 125 patients (18 with Whipple, 71 with PPPD, 13 with DPTPHR, and 23 with MP) were studied. Fecal chymotrypsin and p-type amylase, and pancreatic function diagnostant (PFD) tests were used for evaluation. There were no differences in preoperative background. Pancreatic function was seen to be significantly lower after surgery than before surgery in patients who underwent the Whipple procedure and PPPD (P < 0.05), but there was no difference between pre- and postoperative pancreatic function in patients who underwent DPTPHR and MP. Postoperative pancreatic function was shown to be significantly worse in Whipple procedure and PPPD patients than in those with DPTPHR and MP (P < 0.05). Patients who underwent the Whipple procedure and PPPD showed significantly lower pancreatic function than patients who underwent DPTPHR and MP (P < 0.05). There was no difference in pancreatic function between patients who underwent DPTPHR and those with MP. DPTPHR and MP, both of which preserve the entire duodenum, maintain pancreatic function more than the Whipple procedure and PPPD.

Aged↗
PubMed2000

Revision of failed metal-backed patellar component of Miller/Galante-I total knee prosthesis.

Of 38 Miller/Galante-I total knee prostheses implanted between 1987 and 1990, 20 (53%) were revised. Failed metal-backed patella (15 knees; 40%) was the most frequent reason for revision. In most patients, the components were found well fixed to bone. The purpose of this study was to describe the revision technique we used for failed metal-backed patella. Total revision of all components would be ideal. However, removal of all components results in large bone loss, large blood loss, and a longer operation time. Because of the patients' relatively old age (78.7 years), less demanding activities of daily life (ADL), and the lower body weight of the Japanese patient, we chose a less invasive approach. The revision technique had the following features: (1) the grooved femoral component was not replaced, (2) the failed patellar component was replaced by an all-polyethylene patella, (3) synovectomy was performed, and (4) realignment of patellar tracking was done by lateral release and medial imbrication of the quadriceps mechanism. The mean follow-up after revision was 2 years, 4 months, and good short-term results were obtained. This less invasive approach would be beneficial for elderly patients whose ADL are less demanding.

Aged↗