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Biomedical subjects

H Ohtani

Publications and source records attributed to H Ohtani.

At least 235 records · Page 13Linked to original sources
PubMed1995

Nucleic acid hybridization accompanied with excimer formation from two pyrene-labeled probes.

We developed a novel nucleic acid hybridization method based on excimer formation. We used two different 16-mer oligonucleotide probes that had a combined continuous-sequence run that was complementary to a target 32-mer. Prior to hybridization, the adjacent terminal ends (i.e. the 3'-terminal of one probe and the 5'-terminal of the other probe) were each labeled with one pyrene residue. When these probes simultaneously hybridized to the target, a 495 nm broad fluorescence band was produced. The intensity of this band increased as the intensity of the pyrene monomer bands decreased, indicating that the 495 nm band was attributed to the pyrene excimer. The excimer fluorescence, easily differentiated from the monomer bands for emission wavelength, opens up a new way to perform homogeneous hybridization assays and in vivo imaging of nucleic acids.

Base Sequence↗
PubMed1995

In situ expression of the cell adhesion molecules in bronchial tissues from asthmatics with air flow limitation: in vivo evidence of VCAM-1/VLA-4 interaction in selective eosinophil infiltration.

Eosinophils play a critical role in the pathogenesis of bronchial asthma by releasing various mediators. To understand the mechanisms of eosinophil migration to the site of inflammation, we examined the expression of adhesion molecules in the bronchial tissues of asthmatic subjects with air flow limitation. By immunohistochemical analysis, Mac-1, LFA-1, and VLA-4 were strongly positive in eosinophils and mononuclear cells infiltrated in the bronchial mucosa and submucosa. Their number was significantly increased compared with those in control tissue. Immunolocalization for ICAM-1, the ligand of Mac-1 and LFA-1, was detected in the endothelial cells of capillaries and venules, in the mononuclear cells in submucosa, and in the basal layer of the epithelium. Endothelial cells in capillaries and venules were also strongly positive for VCAM-1, the ligand of VLA-4. Immunolocalization for E-selectin was detected in some endothelial cells in capillaries and venules in bronchial submucosa, whereas there were very few positive cells in the bronchial tissues from control subjects. In situ hybridization demonstrated ICAM-1 mRNA expression in the endothelial cells and mononuclear cells in bronchial submucosa. Immunoelectron microscopy for ICAM-1, VCAM-1, and E-selectin demonstrated de novo synthesis of these molecules and their expression along the luminal cell membrane of endothelial cells. These results suggested that ICAM-1, VCAM-1, and E-selectin were newly synthesized prior to spontaneous asthma attacks, and that their expression, particularly that of VCAM-1, may play a key role in eosinophil infiltration into the airway.

Adult↗
PubMed1995

[Inhibitory effects of sepimostat mesilate (FUT-187) on the activities of trypsin-like serine proteases in vitro].

Inhibitory activities of FUT-187 on trypsin-like serine proteases were compared using camostat mesilate (camostat), and 4-(4-guanidino benzoyloxy)-phenyl acetic acid methanesulfonate (GBPA) known as an active metabolite of camostat in the blood. Ki values of FUT-187 on the competitive inhibition mechanism were 0.097 microM for trypsin, 0.029 microM for pancreatic kallikrein, 0.61 microM for plasma kallikrein, 0.57 microM for plasmin, 2.5 microM for thrombin, 20.4 microM for factor Xa and 6.4 microM for C1r. However, FUT-187 acted as a noncompetitive inhibitor for factor XIIa and an uncompetitive inhibitor for C1s, and Ki values for these proteases were 0.021 and 0.18 microM, respectively. Ki values of camostat for these proteases were in the range of 0.037 to 96.4 microM, and those of GBPA for the above proteases except trypsin and plasma kallikrein were higher than those of FUT-187. The inhibitory activity of FUT-187 on trypsin was not reduced by the addition of the serum at 10%, whereas, that of GBPA was reduced (4.3 fold) in terms of IC50 values. The concentration of FUT-187 required to double APTT (activated partial thromboplastin time) was 1.09 microM, while GBPA, by concentrations up to 1 mM failed to double APTT. The kinin formation by glandular kallikrein in the rat plasma was inhibited by FUT-187 with IC50 value of 0.024 microM, while camostat revealed no inhibition by concentrations up to 1 microM. The complement-mediated hemolyses in the classical and alternative pathways were also inhibited by FUT-187 with IC50 values of 0.17 and 3.5 microM, respectively, the corresponding values for camostat being 350 and 150 microM, respectively. It is concluded that FUT-187 is a potent and selective inhibitor of trypsin-like serine proteases, and its inhibitory activities are stronger than those of camostat on glandular kallikrein, factor XIIa and C1s in complement pathway.

Animals↗
PubMed1995

Induction of heat-shock proteins HSP73 and HSP90 in rat kidneys after ischemia.

We examined rat kidneys for serial expressions of two major heat-shock proteins (HSPs), HSP73 and HSP90, after 60 min of unilateral renal ischemia up to day 28. Immunohistochemical studies showed that HSP73 and HSP90 were rapidly induced in the cytoplasm of injured epithelial cells of the S3 segment of proximal tubules and were again induced in the cytoplasm of regenerative cells in this segment from day 3. In epithelial cells of the Henle's loops, HSP90 was also induced in the cytoplasm of both injured and regenerative cells, but HSP73 was not induced in this portion. Furthermore, a transient accumulation of HSP73 into the nucleus was observed in epithelial cells of papillary collecting ducts shortly after ischemia. Serial immunoblot analysis of isotonic buffer extractable fractions from ischemic kidneys revealed the induction of both HSP73 and HSP90 in the degenerative and regenerative phases: the maximal inductions in the two phases were at 3-6 and on days 5-7, respectively. These results demonstrate that HSP73 and HSP90 are induced in injured tubular epithelial cells with a regional heterogeneity during the degenerative and regenerative phases after renal ischemia and suggest that these HSPs are involved in the process of postischemic cellular recovery.

Animals↗
PubMed1995

[An autopsy case of segmental mediolytic arteritis (SMA) accompanied with microscopic polyarteritis nodosa].

Segmental Mediolytic Arteritis (SMA) is a very rare arterial lesion which is limited in adults to the involvement of the intra-abdominal muscular arteries. The pathology is characterized by segmental disruption of the arterial media which leads segmental mediolysis, with subsequent dissecting aneurysm or rupture. A 73-year-old man was admitted to a hospital because of high fever, general fatigue and weight loss. These symptoms were resistant to antibiotic therapy, and soon after, renal insufficiency developed. Three months after the onset of symptoms, he died suddenly of hemorrhagic shock. Autopsy revealed rupture of the splenic artery and systemic necrotizing arteritis in the small-sized arteries of liver, pancreas and kidneys, as well as in the connective tissues around the adrenal glands. Histopathology of the splenic artery was consistent with SMA, and that of the systemic vascular lesions, with microscopic polyarteritis nodosa. Focal glomerular lesions characteristic of crescentic and/or granulomatous glomerulonephritis were present. A developmental mechanism for SMA is discussed with respect to this case together with a review of previous reports of this disease.

Adult↗
PubMed1995

[Molecular diagnosis of a kindred with novel mutation of methylmalonyl-CoA mutase gene using non-RI SSCP].

A deficiency of methylmalonyl-CoA mutase (MCM) results in methylmalonic acidemia, which is inherited as an autosomal recessive disease and is characterized by accumulation of precursors and abnormal derivatives of methylmalonyl-CoA in body fluids. Abnormal splicing with 13 base pairs (bp) insertion at MCM exons 2 and 3 junction in MCM transcripts and a homozygous point mutation, g to a transition, on 5 bp downstream exon 2 were detected in a proband with methylmalonic acidemia. The parents in the kindred were heterozygous carriers of the g to a transition in MCM intron 2. Non-RI single strand conformation polymorphisms (SSCP) was conducted to devise for analysis of this MCM mutation. This non-RI SSCP is considered to be useful diagnostic means with high potential for extended clinical application.

Adult↗
PubMed1995

[Effective management and its counter plan in clinical laboratories].

The effective management for clinical laboratories at Kitasato University Hospital in these seven years was reported. To improve the economical efficiency, I pointed out that it is important to improve the efficiency of the laboratory work and to reorganize the clinical laboratory. In addition, we may have to take advantage of contract tests in consideration of profit-and-loss. On the other hand, it is desirable among the clinicians to improve emergency tests, to support their research, to consult on the test results and to ask for any additional interpretations of the obtained data. A system that can satisfy their needs should be established.

Clinical Laboratory Techniques↗
PubMed1995

[Molecular differential diagnosis of herpes virus using common primer pairs: detection of HSV-1, HSV-2, VZV and CMV by the PCR].

Differential diagnosis of herpes virus species is often required in various clinical courses, such as transplantation and immunocompromised patients. Pairs of common primers which enabled sufficient amplifying and differential diagnosis of herpes virus species were designed and nested PCR using the common primer pairs was conducted to evaluate for its clinical application. The PCR was shown to provide amplification and differential diagnosis of HSV-1, HSV-2, VZV and CMV. The results of the PCR were consistent with those from fluorescent antibody of herpes virus culture and isolation. From clinical vesicular specimens, the PCR enabled the direct detection of HSV-1, HSV-2, and VZV. The PCR using the common primer pairs offers speed in the differential diagnosing of herpes virus.

Base Sequence↗
PubMed1995

Induction and intracellular localization of 90-kilodalton heat-shock protein in rat kidneys with acute gentamicin nephropathy.

BACKGROUND: We previously reported the induction of 73-kilodalton heat-shock protein (HSP73) in injured tubular epithelial cells in rat kidneys with gentamicin-induced acute renal failure. In the present study, we examined serial expression of 90-kilodalton heat-shock protein (HSP90), another major HSP, in this animal model. EXPERIMENTAL DESIGN: Sprague-Dawley rats received gentamicin (80 mg/kg/day) for 14 days and developed acute proximal tubular injury. Serial immunohistochemical localization of HSP90 was observed at both light microscopic and electron microscopic levels, using a specific antibody against HSP90. In addition, serial renal extracts were analyzed by immunoblot. RESULTS: On light microscopy, HSP90 was induced in injured proximal tubular epithelial cells and accumulated in fine granules 36 hours after the gentamicin exposure. The size and number of these granules gradually increased to Day 12 and decreased from Day 18, and the granules disappeared on Day 27. Electron microscopy showed the accumulation of HSP90 in the swollen lysosomes and the nucleoli of the injured proximal tubular epithelial cells. On serial immunoblot analysis of renal extracts, increased amounts of HSP90 were found in association with the induction of HSP90 in injured cells. Furthermore, on immunoblot of nuclear fractions from kidneys at Days 0 and 6, HSP90 was detected in the fraction at Day 6. CONCLUSIONS: These results demonstrate that HSP90 is induced in the lysosomes and the nucleoli of damaged cells during the course of gentamicin-induced acute tubular injury. HSP90 may have roles in the disposition of degenerated proteins and in the new protein synthesis for the protection and repair of target cells from gentamicin nephrotoxicity.

Acute Kidney Injury↗
PubMed1995

Immunohistochemical localization of transforming growth factor-beta receptors I and II in inflammatory bowel disease.

Transforming growth factor (TGF)-beta inhibits cell proliferation and stimulates cell differentiation. For the signal transduction of TGF-beta, receptors I and II are required. The present study discloses the immunohistochemical localization of TGF-beta receptors I and II in inflammatory bowel disease. In the normal colon and small intestinal tissue, TGF-beta receptors were expressed in the epithelial cells in the upper crypts. In inflammatory bowel disease, a disordered expression pattern of receptors I and II was observed in the epithelial cells. Fibroblasts in the area of early stage fibrosis were positive for both receptors, while fibroblasts in area of advanced fibrosis lacked immunoreactivity for both receptors. Our study suggests that TGF-beta receptors I and II are important for the initial step of fibrosis.

Animals↗
PubMed1995

Excimer formation by hybridization using two pyrene-labeled oligonucleotide probes.

Pyrene excimer formation was achieved in an aqueous solution by nucleic acid hybridization. We used two different 16-mer oligonucleotide probes which had a combined continuous-sequence run complimentary to a target 32-mer. Prior to hybridization, the adjacent terminal ends (i.e., the 3'-terminal of one probe and the 5'-terminal of the other probe) were each labeled with one pyrene residue. The simultaneous hybridization of the probes to the target induced a 495-nm broad fluorescence band characteristic for pyrene excimer. Temperature and molecular configurations of the hybrid influenced the excimer formation.

Base Sequence↗
PubMed1994

Effects of tetracyclines on neutrophil, gingival, and salivary collagenases. A functional and western-blot assessment with special reference to their cellular sources in periodontal diseases.

The characterization and regulation of matrix metalloproteinases (MMPs) have been studied to determine their role(s) in periodontal tissue destruction. Progress in elucidating the roles of MMPs in periodontal tissue destruction has led to a new concept involving the chemotherapeutic inhibition on MMPs, a therapeutic strategy which less than a decade ago was considered "a difficult and perhaps impossible task." Tetracyclines/doxycycline (DOXY) and their chemically modified nonantimicrobial derivatives (CMTs) are known to inhibit the matrix metalloproteinases, especially preferring human neutrophil collagenase (MMP-8), and prevent the oxidative activation of procollagenases. We characterized by Western blotting the molecular forms and cellular sources of gingival tissue, dental plaque, gingival crevicular fluid (GCF), and salivary MMPs associated with periodontitis. Also the molecular forms of tissue inhibitors of matrix metalloproteinases (TIMP-1 and TIMP-2) in periodontitis were studied by Western blot. Neutrophil (PMN)-derived MMPs were found to predominate in periodontitis, and phospholipase C present in increased amounts in periodontitis sites was found to be a potential inducer of PMN degranulation. We further studied the effects of DOXY on molecular forms of different latent and active MMPs purified from different cellular sources (PMNs, fibroblasts, keratinocytes) and present in vivo in oral exudates (gingival extracts, GCF, and saliva). DOXY inhibition of activated (oxidatively or proteolytically) MMPs were not associated with MMP fragmentation. Michaelis-Menten plots of initial rates of degradation of soluble type I collagen revealed an apparent Km value of 0.3-0.6 microM for MMP-8, and 75 microM DOXY inhibited MMP-8 in a manner which did not result in changes in apparent Km value but did prevent the initial degradation reaching Vmax providing evidence for noncompetitive inhibition. Treatment of patients with long-term DOXY medication results in decreased MMP-8 activities/levels in gingival tissue, crevicular fluid, and saliva, but not fragmentation of MMP-8 in vivo. These data further support and extend the key role of PMN-MMPs in periodontitis, and the activities of these PMN MMPs can be inhibited directly by therapeutic levels of DOXY.

Blotting, Western↗
PubMed1994

Genomic instability of microsatellite repeats and mutations of H-, K-, and N-ras, and p53 genes in renal cell carcinoma.

Thirty-six primary renal cell carcinoma samples and one metastatic lymph node DNA sample were examined for mutations of H-, K-, and N-ras and p53 genes, and genomic instability at (AC)n, (CA)n.(GT)n, and (TA)n.(GT)n repeats. No mutations were noted for H-, K-, and N-ras genes and only 2 of all the samples (5.6%) showed mutations at exon 8 of the p53 gene. Differences in unrelated microsatellites for tumor and normal DNA were detected in 9 (25.0%) of the cases examined. Somatic alterations in seven microsatellites, D3S1228, D3S643, D5S107, LPL5GT, D9S63, D17S261, and DCC, were found in 1 (2.8%), 3 (8.3%), 2 (5.7%), 5 (14.7%), 3 (8.3%), 3 (8.3%), and 3 (8.3%) cases, respectively. Five of 26 (19.2%) clear cell type and 4 of 10 (40.0%) non-clear cell type patients showed DNA instability. Two of 11 (18.2%) grade 1, 5 of 20 (25.0%) grade 2, and 2 of 5 (40.0%) grade 3 patients showed abnormal patterns. One of 2 (50.0%) stage pT1, 4 of 24 (16.7%) stage pT2, and 4 of 10 (40.0%) stage pT3 patients were shown to have microsatellite instability. In 4 of 9 alteration-positive cases (44.4%), mutations in multiple microsatellites were observed. Alterations in microsatellite instability may be more common in non-clear cell type, high-grade, and high-stage renal cell carcinoma patients.

Base Sequence↗
PubMed1994

Genomic instability of microsatellite repeats and its association with the evolution of chronic myelogenous leukemia.

Tumorigenesis has been shown to proceed through a series of genetic alterations involving protooncogenes and tumor-suppressor genes. Investigation of genomic instability of microsatellites has indicated a new mechanism for human carcinogenesis in hereditary nonpolyposis colorectal cancer and sporadic cancer and this instability has been shown to be related to inherited predisposition to cancer. This study was conducted to determine whether such microsatellite instability is associated with the evolution of chronic myelogenous leukemia (CML) to the blast crisis. Nineteen CML patients clinically progressing from the chronic phase to accelerated phase or blast crisis and 20 other patients in the CML chronic phase were studied. By polymerase chain reaction assay, DNAs for genomic instability in five separate microsatellites in chromosome arms 5q (Mfd27), 17p (Mfd41), 18q (DCC), 3p (CI3-9), and 8p (LPL) were examined. Differences in unrelated microsatellites of chronic and blastic phase DNAs in 14 of 19 patients (73.7%) were demonstrated. Somatic instability in five microsatellites, Mfd27, Mfd41, DCC, CI3-9, and LPL, was detected in 2 of 19 (10.5%), 8 of 19 (42.1%), 11 of 19 (57.9%), 4 of 17 (23.5%), and 4 of 17 (23.5%) cases. In 10 of 19 cases (52.6%), genetic instability in at least two of five microsatellites was observed and was categorized as replication error (RER+) phenotype. CML evolution cases with myeloid, lymphoid, and mixed phenotypes and the blast crisis and accelerated phase showed somatic instability in a number of microsatellites. No alterations in leukemic cells at the chronic phase could be detected in any microsatellites. These data indicate instability of microsatellites (RER+) but not familial predisposition to possibly be a late genetic event in the evolution of CML to blast crisis. In the microsatellite of the DCC gene, complicated alterations in band patterns caused by instability as well as loss of heterozygosity (LOH) were observed in 13 of 19 cases (68.4%): instability in 9 cases, instability plus LOH in 2 cases, and only LOH in 2 cases. These highly frequent alterations in microsatellites, including instability and LOH, suggesting that secondary events due possibly to loss of fidelity in replication and repair machinery may be significantly associated with CML evolution.

Adult↗
PubMed1994

Remodelling of microvessels by coronary hypertension or cardiac hypertrophy in rats.

We attempted to clarify the individual effects of coronary hypertension and ventricular pressure-overload on the remodelling of coronary arterial microvessels. We examined structural vascular changes in the right ventricular free walls using two experimental rat models: 12-week ascending aortic banding, i.e. a model of coronary hypertension without ventricular pressure-overload and 12-week pulmonary artery banding, i.e. a model of ventricular pressure-overload without coronary hypertension. In vivo peak systolic left ventricular pressure and left ventricular weight to body weight ratio in the aortic banding model increased to 198 +/- 10 mmHg and by 39% above the controls, respectively. In vivo peak right ventricular pressure and right ventricular weight to body weight ratio in the pulmonary artery banding model increased to 87 +/- 7 mmHg and by 69% above the controls, respectively. The right coronary reserve measured by the Doppler method decreased in both experimental models. Aortic banding increased the medial area and perivascular collagen to lumen area ratios in the microvessels > 30 microns in lumen diameter but not in those < 30 microns. Pulmonary artery banding increased the perivascular collagen to lumen area ratio in the microvessels < 60 microns, but not in those > 60 microns. Medial thickening was not observed over the whole range of lumen diameter. These results suggest that the mechanisms responsible for the remodelling of coronary microvessels differ between non-hypertrophied hearts with coronary hypertension and hypertrophied hearts without coronary hypertension.

Animals↗
PubMed1994

Bilateral malignant lymphoma of the breast in patients with a familial history of cancer: a report of three cases.

We report three cases of bilateral malignant lymphoma of the breast in patients with a familial history of cancers. All the patients had B-cell malignant lymphoma and were treated by a radical mastectomy and polychemotherapy. Bilateral tumors were detected synchronously in one case, whereas the second tumors in the other cases occurred 4 years and 8 years after the initial treatment. Although the patients had no relevant past history, these cases were notable for the fact that all were associated with a familial history of cancers. One patient had a definite familial history of breast cancer, that is to say, her mother, sister and brother all suffered from this disease. The brother of another patient died of gastric cancer while two of her uncles each suffered from sarcoma of the lower extremities and a brain tumor, respectively. As for the remaining patient, one of her maternal uncles and an aunt also died of prostate cancer and leukemia, respectively. However, only one out of seven patients with unilateral malignant lymphoma of the breast used for comparison had a familial history of cancer. A possible relationship between bilateral primary malignant lymphoma of the breast and a familial history of cancer is suggested.

Adult↗