Biomedical subjects

K Hamada

Publications and source records attributed to K Hamada.

At least 253 records · Page 14Linked to original sources

PubMed1996

Cloning and expression of the canine interferon-beta gene.

We isolated the canine interferon-beta (IFN-beta) gene from dog liver chromosomal DNA by the polymerase chain reaction (PCR). The coding region encodes a predicted protein of 197 amino acids, consisting of a signal sequence of 32 amino acids and a mature IFN-beta of 165 amino acids. In the IFN-beta sequence, there are five potential N-glycosylation sites and four cysteine residues. Canine IFN-beta has 44% and 60% amino acid sequence homology with murine and human IFN-beta, respectively, whereas it has only 28% homology with canine IFN-alpha. The canine IFN-beta gene was expressed in insect cells under the control of the polyhedrin promoter in a recombinant baculovirus. After infecting Sf21 cells with the recombinant baculovirus, IFN activity was detected in the culture medium, indicating that it is secreted from the cells. This activity was stable from pH 2 to 12 for 18 h at 4 degrees C. Southern blot analysis indicated that the gene for canine IFN-beta is a single gene in the dog haploid chromosome.

Amino Acid Sequence↗

PubMed1996

Crystal structure of Serratia protease, a zinc-dependent proteinase from Serratia sp. E-15, containing a beta-sheet coil motif at 2.0 A resolution.

The crystal structure of Serratia protease from Serratia sp. E-15 was solved by the single isomorphous replacement method supplemented with anomalous scattering effects from both the Zn atom in the native crystal and the Sm atom in the derivative crystal, and refined at 2.0 A resolution to a crystallographic R-factor of 0.194. The enzyme consists of N-terminal catalytic and C-terminal beta-sandwich domains, as observed in alkaline protease from Pseudomonas aeruginosa IFO3080. The catalytic domain with a five-stranded antiparallel beta-sheet and five alpha-helices shares a basically common folding topology with those of other zinc metalloendoproteases. The catalytic zinc ion at the bottom of the active site cleft is ligated by His176, His180, His186, Tyr216, and a water molecule in a distorted trigonalbipyramidal manner. The C-terminal domain is a beta-strand-rich domain containing eighteen beta-strands and a short alpha-helix, and has seven Ca2+ ions bound to calcium binding loops. An unusual beta-sheet coil motif is observed in this domain, where the beta-strands and calcium binding loops are alternately incorporated into an elliptical right-handed spiral so as to form a two-layer untwisted beta-sandwich structure. The Ca2+ ions in the C-terminal domain seem to be very important for the folding and stability of the beta-sheet coil structure.

Binding Sites↗

PubMed1996

Physiologic responses to exchange transfusion with hemoglobin vesicles as an artificial oxygen carrier in anesthetized rats: changes in mean arterial pressure and renal cortical tissue oxygen tension.

OBJECTIVES: To evaluate the oxygen transporting capabilities of hemoglobin vesicles by studying the physiologic responses to exchange transfusion with hemoglobin vesicles in anesthetized rats. Exchange transfusions with phosphate buffered saline, hemoglobin vesicles containing methemoglobin (and therefore, deprived of oxygen transporting capabilities), and washed rat red blood cells were used as controls. DESIGN: Prospective, randomized, controlled trial. SETTING: Department of Surgery, School of Medicine, Keio University. SUBJECTS: Twenty-seven male Wistar rats. INTERVENTIONS: The rats were anesthetized with an intraperitoneal injection of sodium pentobarbital (50 mg/kg). Catheters (PE-20 tubing, outer diameter 0.8 mm, inner diameter 0.5 mm) were introduced into the right jugular vein for infusion and the right common carotid artery for blood withdrawal and mean arterial pressure measurements. The left kidney was exposed by median abdominal incision, and a needle-type polarographic oxygen electrode was placed in the left renal cortex for renal cortical tissue oxygen tension measurements. MEASUREMENTS AND MAIN RESULTS: Phosphate buffered saline and methemoglobin vesicles were administered as nonoxygen-carrying fluids, and rat red blood cells as oxygen-carrying fluid. Measurements included mean arterial pressure, arterial blood gas analysis, and renal cortical tissue oxygen tension as an indicator of systemic oxygen transport. In the rat red blood cell and hemoglobin vesicles groups, mean arterial pressure was sustained at the end of the exchange transfusion (82.3 +/- 27.5% and 73.5 +/- 11.5%, respectively, from the basal values). However, in the phosphate buffered saline and methemoglobin vesicles groups, mean arterial pressure decreased significantly (p < .05) (33.9 +/- 13.8% and 35.7 +/- 8.2%, respectively). Renal cortical tissue oxygen tension in the rat red blood cell and hemoglobin vesicles groups was sustained at a significantly higher level (p < .05) (83.5 +/- 9.3% and 75.0 +/- 11.9%, respectively) compared with the phosphate buffered saline and methemoglobin vesicles groups (44.9 +/- 12.8% and 58.3 +/- 6.2%, respectively) at the end of the exchange transfusion. Metabolic acidosis was more progressive in the phosphate buffered saline and methemoglobin vesicles groups, manifested as lower pH and base excess values. Platelet counts tended to decrease slightly in the hemoglobin vesicles and methemoglobin vesicles groups, but the changes were not significant. CONCLUSIONS: Hemoglobin vesicles have an oxygen transporting capability almost equivalent to rat red blood cells and can be considered as a potential artificial oxygen carrier.

Anesthesia↗

PubMed1996

Expression and alternative splicing of Pit-1 messenger ribonucleic acid in pituitary adenomas.

Twenty-eight human pituitary adenomas were analyzed for the expression of Pit-1 messenger ribonucleic acid (mRNA) by using reverse transcriptase-polymerase chain reaction analysis of frozen-section mRNA. Pit-1 mRNA was detected in all functioning tumors and in 9 of 11 nonfunctioning tumors. Pit-1 beta, which is a more active isoform of transcriptional factor for growth hormone than Pit- alpha and which arises from an alternative splicing mechanism, was detected in 14 of 17 functioning tumors and in 5 of 11 nonfunctioning tumors. The transcript that corresponds to Pit-1T, which increases thyroid-stimulating hormone beta promoter activity in rat thyrotropic tumor cells, was not found. There was no significant difference in the total Pit-1 (alpha+beta) mRNA expression level between functioning tumors and nonfunctioning tumors. Growth hormone-producing tumors and other pituitary adenomas also showed no significant difference in the Pit-1 beta/Pit-1 alpha expression ratio. Our data suggest that the major role of Pit-1 gene in pituitary adenoma might not be involved in the regulation of hormone production.

Adenoma↗

PubMed1996

The metabolism of testosterone by dermal papilla cells cultured from human pubic and axillary hair follicles concurs with hair growth in 5 alpha-reductase deficiency.

Androgens regulate the growth of many human hair follicles, but only pubic, axillary, and scalp hair growth occur in men with 5 alpha-reductase deficiency. This suggests that 5 alpha-dihydrotestosterone is the active intracellular androgen in androgen-dependent follicles, except in the axilla and pubis. Since the dermal papilla plays a major regulatory role in hair follicles and may be the site of androgen action, we have investigated androgen metabolism in six primary lines of cultured dermal papilla cells from pubic and axillary hair follicles; previous studies have shown that beard cells take up and metabolize testosterone, retaining and secreting 5 alpha-dihydrotestosterone. After 24 h preincubation in serum-free Eagle's medium 199, 100-mm dishes of confluent cells were incubated for 2 h with 5 nM [1,2,6,7-3H]testosterone. Media were collected and the cells washed with phosphate-buffered saline and extracted with chloroform: methanol (2:1). After the addition of unlabeled and 14C-labeled marker steroids, the extracts were analyzed by a two-step thin-layer chromatography system; steroid identity was confirmed by recrystallization to a constant 3H/14C ratio. Beard and pubic dermal papilla cells were also incubated for 24 h, and the medium was analyzed at various times. The results from pubic and axillary primary cell lines were similar. In both cells and media the major steroid identified was testosterone, but significant amounts of androstenedione were present, indicating 17 beta-hydroxysteroid dehydrogenase activity; androstenedione was also identified within the cells, but a small amount of 5 alpha-dihydrotestosterone was only identified in one pubic cell line. Beard dermal papilla cells secreted large amounts of 5 alpha-dihydrotestosterone into the medium over 24 h in contrast to pubic cells, which produced only very small amounts. The pubic and axillary cell results contrasts with the observations of pronounced 5 alpha-dihydrotestosterone in beard cells and confirm that androgen metabolism in cultured dermal papilla cells reflects the parent follicle's ability to respond to androgen in the absence of 5 alpha-reductase type II in vivo. This supports our hypothesis that androgen acts on hair follicles via the dermal papilla and suggests that cultured dermal papilla cells may offer an important model system for studies of androgen action.

Cells, Cultured↗

PubMed1996

Allelic loss of chromosome 16q in endometrial cancer: correlation with poor prognosis of patients and less differentiated histology.

Deletion of certain chromosomal regions can be demonstrated in malignant cells. Chromosome 16q is one of the regions where allelic loss is frequently detected in carcinoma of the breast and many other tumors, suggesting that gene(s) which retard tumor growth may exist here. To elucidate the clinicopathological significance of chromosome 16q, loss of heterozygosity (LOH) was investigated using microsatellite polymorphism analysis in 58 patients with endometrial lesions (50 with endometrial carcinoma and 8 who had hyperplasia with or without atypia). When 11 regions of chromosome 16q were examined, LOH was found in 20 patients with carcinoma (40%) and none of the patients with hyperplasia. The tumors of 9 of the 20 patients (45%) showed total loss of 16q, while the others (55%) showed partial deletion. Tumors with LOH were histologically less differentiated than those without LOH (P = 0.038, chi2 test). Patients with tumors showing LOH of 16q had a worse prognosis than those without LOH according to Kaplan-Meier survival analysis (P=0.0158, log-rank test). In addition, LOH of 16q showed a significant relationship to prognosis by Cox regression analysis. Deletion mapping of 16q demonstrated that two regions (16q22.1 and 16q22.2-23.1) were frequently involved. Patients with 16q22.1 LOH had a poorer prognosis than those with intact 16q22.1 (P=0.0003, log-rank test). These findings suggest that gene(s) of which defect is possibly related to the aggressiveness of endometrial cancer are localized on a limited region of 16q that includes 16q22.1.

Alleles↗

PubMed1996

Mechanisms of the suppression of the bladder activity by flavoxate.

BACKGROUND: This study was designed to clarify the primary site of action of flavoxate, clinically used for the treatment of urinary frequency. METHODS: In rats, the effect of flavoxate on contractile responses in isolated detrusor strips, bladder contraction induced by pelvic nerve stimulation, isovolumetric rhythmic bladder contractions, and pelvic nerve activity were examined. In decerebrated cats, flavoxate was microinjected into the nuclei in the pons, and its effect on reflex micturition was observed. RESULTS: Flavoxate suppressed carbachol- and calcium ion (Ca2+)-induced contractions of isolated detrusor strips in a noncompetitive and a competitive manner, respectively. Intravenous flavoxate suppressed both initial phasic, and later tonic, bladder contractions induced by electrical stimulation of the distal end of the pelvic nerve. It abolished isovolumetric rhythmic bladder contractions and the associated efferent pelvic nerve activity, without affecting baseline vesical pressure and afferent pelvic nerve activity. When administered intracerebroventricularly or intrathecally, it abolished isovolumetric rhythmic bladder contractions. Flavoxate microinjected into the nucleus reticularis pontis oralis (PoO; pontine micturition inhibitory region) of decerebrated cats inhibited the reflex micturition, but had no effect when microinjected into the locus coeruleus alpha (pontine micturition center) or locus subcoeruleus (pontine urine storage center). CONCLUSIONS: Flavoxate suppressed the micturition reflex primarily by facilitating the inhibitory action of the PoO on the descending pathways from the pontine micturition center to the sacral parasympathetic intermediolateral nuclei.

Animals↗

PubMed1996

[A case of the greater omentum abscess due to Peptostreptococcus ssp].

A case of the greater omentum abscess due to Peptostreptococcus ssp. is reported. A 63-year-old male was admitted to Heisei Memorial Hospital with a complaint of abdominal pain. An intraperitoneal mass measuring 6 cm in diameter was detected by CT and ultrasonogram. He was diagnosed as having an intraperitoneal abscess. After antibiotics therapy by cefotiam, laparotomy was performed and an abscess about 7 cm in diameter at the edge of the greater omentum was found. There was no evidence of perforation in the intestine wall. The abscess was removed and Peptostreptoccocus spp. was isolated from the dark red pus drained from the abscess.

Abdominal Abscess↗

PubMed1996

[A clinical features of acute respiratory infection with Haemophilus influenzae by transtracheal aspiration].

We performed a clinical study of 26 cases (27 episodes) of acute respiratory infection with H. influenzae by trans tracheal aspiration (TTA) from May 1987 to April 1995. 15 episodes (14 cases) were bronchitis and 12 episodes (12 cases) were pneumonia. 8 episodes were monomicrobial infection and 19 episodes were polymicrobial infection. Compared to the group of patients of monomicrobial infection, the number of elderly patients and the levels of WBC and CRP were higher in the group of patients of monomicroibal infection. In bronchitis cases, monomicrobial infections of H. influenzae were 7 episodes and polymicrobial infection containing H. influenzae were 8 episodes. In the latter group, PaO2 level was lower and CRP was higher on average. All patients recovered, but the period for treatment was longer in the latter group. In the pneumonia group, only one episode was monomicrobal infection and 11 episodes were polymicrobial infection. Inspite of treatment, one patient died. It was considered that polymicrobial infection was an important factor of acute respiratory infection with H. influenzae.

Acute Disease↗

PubMed1996

[A clinical study of chronic lower respiratory infection with Haemophilus influenza by transtracheal aspiration].

We have performed a clinical study on chronic lower respiratory tract infection (CLRTI) with Haemophilus influenzae (H. influenzae) by transtracheal aspiration (TTA) and analyzed clinical factors of the acute exacerbation. In 40 episodes (38 cases) of H. influenzae isolated from CLRTI, monobacterial infection with H. influenzae were 21 episodes and polymicrobial infection were 19 episodes. We classified the disease into acute exacerbated (27 episodes) and stable (13 episodes) phase and the former episodes were divided into bronchitis type (7 episodes) and pneumonia type (20 episodes). Polymicrobial infections were seen more in the pneumonia type (13 episodes) than in the bronchitis type (2 episodes). The principal organism detected with H. influenzae were alpha-Streptococcus and Neisseria sp. in the bronchitis type and S. pneumoniae in the pneumonia type. The acute exacerbated cases were divided into the following 4 patterns; 1. polymicrobial infection with continuous infection of P. aeruginosa, 2. monomicrobial infection after acute upper respiratory tract infection, 3. polymicrobial infection with S. pneumoniae after continuous infection of H. influenzae, 4. bacterial replacement by P. aeruginosa after acute exacerbation. The results of the study suggests that polymicrobial infection is an important chronic lower respiratory tract infection when caused H. influenzae.

Acute Disease↗

PubMed1996

Schizosaccharomyces pombe is more sensitive to pressure stress than Saccharomyces cerevisiae.

The effects of hydrostatic pressure on ultrastructure, microtubules and microfilaments of Schizosaccharomyces pombe were investigated by fluorescence microscopy, conventional electron microscopy and immunoelectron microscopy. Cells were treated with hydrostatic pressure from 0.1 to 400 MPa for 10 min at room temperature. The nuclear membrane was disrupted at above 100 MPa. At 150 MPa the matrixes of mitochondria had an electron dense area. At 250 MPa the cytoplasmic substances changed dramatically, the cellular organelles could hardly be detected and the fragmented nuclear membrane was barely visible. The fluorescence in alpha-tubulin was lost in most of the cells at 100 MPa. The gold particles for anti alpha-tubulin were not visible in the cells at the same level. Cell cycle specific actin distribution was lost even at 50 MPa, although actin dots localized at the central region remained unchanged. Thick actin cables appeared at 100 MPa. Complete depolymerization of F-actin was observed at 150 MPa. These results suggest that S. pombe cells were more sensitive than Saccharomyces cerevisiae cells. The damage to microtubules and nuclear membrane caused by hydrostatic pressure was though to be followed by breakdown of nuclear division apparatus and the inhibition of nuclear division. This damage might contribute to the frequent formation of polyploidy in S. pombe.

Actin Cytoskeleton↗

PubMed1996

Cytopathic effect inhibition assay for canine interferon activity.

Conditions for cytopathic effect (CPE) inhibition assay of canine interferon (IFN) activity in Madin-Darby canine kidney (MDCK) cells and in canine tumor cell line A72 was investigated using the New Jersey strain of vesicular stomatitis virus (VSV). The culture supernatant from canine splenocytes stimulated with ultraviolet-irradiated Newcastle disease viruses was used as reference IFN. MDCK cells were resistant for growth of VSV when the cells were confluent. Full CPE was observed only in a sparsely growing culture. Canine IFN activity could be assayed on less than 10(4) MDCK cells/well of a 96-well microplate, and more than 10(5) TCID50/ml of VSV was required. In A72 cells, VSV growth was not as dependent on cell density as in MDCK cells, requiring 10(3) TCID50/ml of VSV. MDCK-VSV system showed a higher IFN sensitivity than A72-VSV, whereas reproducibility was higher for the latter than the former. Based on these findings, A72-VSV system for canine IFN assay is recommended for practical use due to its easy handling characteristics.

Animals↗

PubMed1996

Evaluation of biochemical indices as a hair cycle marker in C3H mice.

The hair follicles exhibit an intrinsic hair cycle that is divided into three phases; growth (anagen), transition (catagen) and quiescence (telogen). To make sure of the effects on hair growth by chemical substances, we should evaluate the induction of the anagen phase and/or elongation of the anagen period and delay in catagen separately, but the regulatory mechanism of the hair cycle is unclear. We have investigated the levels of biochemical markers in the third hair cycle period of C3H mouse (8 weeks, male) after depilation and compared them with those in a non-treated group. The dorsal areas (2 cm x 4 cm) were clipped and depilated with hair remover. The dorsal skin samples were collected 1, 8, 11, 15 and 18 days after depilation and the levels of biochemical markers, i.e. skin transglutaminase, skin sulfhydryl oxidase, cathepsin D, gamma-glutamyl transpeptidase, alkaline phosphatase, acid phosphatase, tyrosinase activities and histamine content in each skin sample were examined. The levels of gamma-glutamyl transpeptidase, tyrosinase and alkaline phosphatase were relevant to hair re-growth in the control group, but not skin transglutaminase, skin sulfhydryl oxidase, cathepsin D activities. The histamine content increased just after depilation treatment and returned to the normal level within two weeks, compared with the non-treated group. All these results suggest that the markers examined in this C3H mice model are useful for studying the distinctive process of hair re-growth caused by active substances.

Acyltransferases↗

PubMed1996

[Unclassified chronic myeloproliferative disorder (Ph negative/bcr rearrangement negative CML) with late appearing Philadelphia like chromosome].

A 48-year-old male was diagnosed to be unclassified chronic myeloproliferative disorder (UCMPD)/Ph negative bcr rearrangement negative (Ph-/bcr-) CML by hematological, cytogenetical and DNA analyses (Jpn. J. Clin. Hematol. 33(4): 525-531, 1992). Three years and a half after the diagnosis of UCMPD/Ph-bcr- CML, Ph chromosome was observed in 17 of 20 examined cells. Hematological findings showed a transformation into blast crisis. The late appearing of Ph in a case of UCMPD/Ph1-bcr- CML described here is rare. Southern blot analysis using 3' and 5' bcr probe showed no bcr rearrangement. Analyses of BCR/ABL chimeric RNA by RT-PCR method were negative in both of Major- and Minor BCR/ABL chimeric RNA. In the present case it is speculated that Ph is developed as the result of multistep progression and also speculated that the breakpoint at BCR gene differs from Major- and Minor-bcr in usual Ph+CML and de nove Ph+ ALL. Therefore, it may be reasonable that the present case is understood to be a case with late appearing Ph-like chromosome.

Base Sequence↗

PubMed1996

A comparison of the culture and growth of dermal papilla cells from hair follicles from non-balding and balding (androgenetic alopecia) scalp.

Male pattern baldness is a common, androgen-dependent skin problem in adult men which is not well understood, although androgens are believed to act on the hair follicle via the mesenchyme-derived dermal papilla situated in the middle of the hair follicle bulb. Since dermal papilla cells retain specific characteristics in culture, such as hair-growth promoting ability and appropriate features of the mechanism of androgen action, dermal papilla cells from follicles undergoing androgen-stimulated miniaturization may provide a useful in vitro model system. Therefore, dermal papilla cells have been derived from intermediate follicles from balding and nearly clinically normal sites of men with androgenetic alopecia. Balding dermal papillae were much smaller than non-balding ones and grew much less well under normal growth conditions. Supplementing the medium with human serum, rather than fetal calf serum, increased both the yield of established cultures and the number and health of the dermal papilla cells produced. Non-balding cells also grew faster in human serum. Balding cells retained the normal fibroblastic shape and aggregative behaviour of dermal papilla cells, but always grew less well than non-balding cells. Nearly clinically normal dermal papillae were similar, or slightly smaller, in size to non-balding ones, but their growth resembled balding cells. Since balding dermal papilla cells can be cultured, though with much greater difficulty than nonbalding ones, and exhibit differing growth characteristics to non-balding cells, they merit further investigation which may increase our understanding of, and ability to control, androgenetic alopecia.

Adult↗

PubMed1996

Rhythmic discharges recorded from tail muscle nerves after injection of nialamide and L-DOPA solution in spinalized cats.

In 22 decapitated and high spinalized cats, rhythmic discharges were recorded from the nerves supplying the tail muscles, m. extensor caudae lateralis (ECL) and m. flexor caudae longus (FCL) after intravenous injection of Nialamide and L-DOPA solution. In 15 out of 22 cats, stable rhythmic discharges were recorded from tail muscle nerves. Two different discharge patterns were observed. The predominant pattern consisted of an alternating activation between left and right tail muscle nerves and a synchronous activation of ECL and FCL nerves on one side. The second pattern consisted of synchronous activity involving all four tail muscle nerves.

Animals↗

PubMed1996

[Atypical mycobacteriosis (Mycobacterium xenopi) with "initial aggravation"].

A 66-year-old man was admitted to Nara Medical University Hospital because of sputum production and fevre. A chest X-ray film obtained on admission revealed many cysts and an infiltrative shadow in the right upper lung field. The patient was treated with antimycobacterial drugs (isoniazid 400 mg, streptomycin 0.75 g, and rifampicin 450 mg) because acid-fast bacilli were detected in his sputum. Although the symptoms and laboratory data improved, a new infiltrative shadow developed in the right lower lung field two months after the start of treatment. Transbronchial biopsy specimens showed intraluminal organizing exudate and alveolitis. The new lesion resolved when treated with the same antimycobacterial drugs. Mycobacterium xenopi was cultured from the sputum 80 days later. This is the third reported case of atypical mycobacteriosis (non-tuberculous mycobacteriosis) due to M. xenopi in Japan with the "initial aggravation" seen in some patients with typical pulmonary tuberculosis.

Aged↗

PubMed1996

[MRI changes in spontaneous intracranial hypotension].

We report MRI changes in a spontaneous intracranial hypotension(SIH). The patient was 29-year-old woman, who developed headaches in upright position, nausea, and vomiting preceded by pressure feeling of ears. Neurological examination was unremarkable except for hyperreflexia in the lower extremities. Lumbar punctures revealed very low opening pressure, a mild elevated CSF protein and a mild pleocytosis. No evidence of underlying systemic or neoplastic diseases was noted. The brain and cervical MRI showed diffuse and continuous pachymeningeal enhancement with gadolinium. Her symptoms gradually improved within two months without any treatment, and follow-up MRI showed resolution of the abnormalities within five months. The dural enhancement with gadolinium seen in the SIH should be kept in mind in case of hypertrophic pachymeningitis of unknown etiology, and be differentiated from such diseases as hypertrophic pachymeningitis associated with infectious, neoplastic diseases or sarcoidosis.

Adult↗