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T Ishihara

Publications and source records attributed to T Ishihara.

At least 55 records · Page 3Linked to original sources
PubMed2001

Purification, N-terminal amino acid sequence, and some properties of Cu, Zn-superoxide dismutase from Japanese flounder (Paralichthys olivaceus) hepato-pancreas.

Cu, Zn-superoxide dismutase (SOD) has been purified to homogeneity from Japanese flounder Paralichthys olivaceus hepato-pancreas. The purification of the enzyme was carried out by an ethanol/chloroform treatment and acetone precipitation, and then followed by column chromatographies on Q-Sepharose, S-Sepharose and Ultrogel AcA 54. On SDS-PAGE, the purified enzyme gave a single protein band with molecular mass of 17.8 kDa under reducing conditions, and showed approximately equal proportions of 17.8 and 36 kDa molecular mass under non-reducing conditions. Three bands were obtained when the purified enzyme was subjected to native-PAGE, both on protein and activity staining, but the electrophoretic mobility of the purified enzyme differed from that of bovine erythrocyte Cu, Zn-SOD. Isoelectric point values of 5.9, 6.0 and 6.2, respectively, were obtained for the three components. The N-terminal amino acid sequence of the purified enzyme was determined for 25 amino acid residues, and the sequence was compared with other Cu, Zn-SODs. The N-terminal alanine residue was unacetylated, as in the case of swordfish SOD. Above 60 degrees C, the thermostability of the enzyme was much lower than that of bovine Cu, Zn-SOD.

Amino Acid Sequence↗
PubMed2001

Improved diabetic syndrome in C57BL/KsJ-db/db mice by oral administration of the Na(+)-glucose cotransporter inhibitor T-1095.

1. The therapeutic effects of an orally active inhibitor of Na(+)-glucose cotransporter (SGLT), T-1095 (a derivative of phlorizin; 3-(benzo[b]furan-5-yl)-2',6'-dihydroxy-4'-methylpropiophenone 2'-O-(6-O-methoxycarbonyl-beta-D-glycopyranoside)) were examined in C57BL/KsJ-db/db (db/db) mice, a genetic animal model of obese type 2 diabetes. 2. The higher renal SGLT activity in db/db mice than normoglycaemic C57BL/KsJ-db/+m (db/+m) mice may support the rationale for using an SGLT inhibitor in the treatment regimen for type 2 diabetes. Both T-1095 and its metabolite, T-1095A, which had approximately 10 times more potency, effectively inhibited renal SGLT activity of these mice in vitro. 3. Single oral administration of T-1095 (10, 30, 100 mg kg(-1), p.o.) to db/db mice caused a dose-dependent reduction in blood glucose levels and a concomitant increase in glucose excretion into urine. In contrast, T-1095 only slightly affected blood glucose levels in db/+m mice. 4. Chronic administration of T-1095 (0.1% w w(-1) pellet chow, for 12 weeks) decreased blood glucose and haemoglobin A(1C) levels, and improved glucose intolerance in db/db mice. The age-related decrease in plasma insulin levels was markedly inhibited and there was a 2.5 fold increase of insulin content in the pancreas of T-1095-treated db/db mice. Food consumption was not changed, while impaired body weight gain was ameliorated by T-1095 treatment. 5. Both the development of albuminuria and the expansion of glomerular mesangial area in db/db mice were significantly suppressed by chronic T-1095 treatment, indicating the prevention of the progression of diabetic nephropathy. 6. These results demonstrate that the SGLT inhibitor T-1095 is able to improve the metabolic abnormalities and inhibit the development of diabetic complications in db/db mice. Thus, T-1095 can be used for therapy of type 2 diabetic patients.

Animals↗
PubMed2001

Loss of beta-catenin expression associated with disease progression in malignant melanoma.

BACKGROUND: beta-catenin plays a crucial role in the function of cell adhesion molecules and also participates in growth regulatory signalling pathways that may be involved in malignant transformation. OBJECTIVES: To examine beta-catenin expression in lesions of melanocytic origin for associations with clinicopathological markers of disease progression and for its significance as a predictor of disease recurrence and prognosis. METHODS: beta-catenin expression was examined by immunoperoxidase staining in 50 melanocytic naevi and 91 primary and 50 metastatic melanomas. RESULTS: beta-catenin was expressed in 96% of melanocytic naevi, in 94% and 65%, respectively, of radial and vertical growth phase primary melanomas, and in 38% of metastatic melanomas. Benign and malignant melanocytic lesions had distinct patterns of beta-catenin localization. Most lesions expressing beta-catenin exhibited cytoplasmic staining; however, over 40% of benign lesions also displayed nuclear staining, which was present only in 10% of primary and 15% of metastatic melanomas. Absent or weak expression of beta-catenin in primary melanomas was associated with several markers of disease progression, including tumour thickness and presence of lymph node metastases. A similar but not statistically significant trend was observed for the association of beta-catenin expression with disease recurrence and prognosis. CONCLUSIONS: These results suggest that loss or downregulation of beta-catenin expression in melanoma cells plays a significant role in progression of the disease.

Adolescent↗
PubMed2001

Use of a monoclonal antibody against Lafora bodies for the immunocytochemical study of ground-glass inclusions in hepatocytes due to cyanamide.

AIMS: Ground-glass inclusions (GGIs) in hepatocytes are known to be associated with cyanamide treatment in patients with alcohol dependency. The purpose of this study was to assess the reactivity of a monoclonal antibody (MAb) raised against polyglucosan and to detect early events in GGI formation. METHODS AND RESULTS: Formalin-fixed paraffin-embedded liver tissues from four patients treated with cyanamide were used. Sections were stained with haematoxylin and eosin and periodic acid-Schiff with and without diastase digestion, and were immunohistochemically stained with the MAb. For electron microscopic study, routinely processed liver tissue from one patient was examined with conventional and immunoelectron microscopy with use of the MAb. All specimens from the four cyanamide-treated patients contained GGIs in the cytoplasm of hepatocytes, and these GGIs reacted intensely with the MAb. Fully developed GGIs contained various organelles, whereas early ones consisted primarily of glycogen granules and dilated smooth endoplasmic reticulum. In immunoelectron microscopic preparations, gold particles were located within GGIs, and the immunolabelled organelles appeared to be glycogen granules. CONCLUSIONS: This novel MAb is useful for the detection of GGIs caused by cyanamide. Our results support the idea that GGI formation may result from specific abnormalities in glucose metabolism.

Adult↗
PubMed2001

Role of UGT1A1 mutation in fasting hyperbilirubinemia.

BACKGROUND AND AIM: Low-grade fasting hyperbilirubinemia is a common observation in healthy subjects (HS), whereas high-grade fasting hyperbilirubinemia is believed to be a characteristic finding of Gilbert's syndrome. This study was undertaken to assess the role of mutation in bilirubin UDP- glycosyltransferase gene (UGT1A1) on fasting hyperbilirubinemia. METHODS: Analysis of UGT1A1 and a caloric restriction test (400 kcal for 24 h) were performed in 56 healthy subjects (25 males, 31 females), and 28 patients with Gilbert's syndrome (18 males, 10 females). There were 29 healthy subjects with no mutation in UGT1A1, and 27 healthy subjects and 26 Gilbert's syndrome patients with mutations in the coding and/or promoter (TATA box) regions of UGT1A1. RESULTS: The mean increment of serum bilirubin (DeltaSB) was 7.6 micromol/L [corrected] (males) and 4.1 micromol/L (females) in subjects with no UGT1A1 mutation. Subjects with mutation in UGT1A1 showed higher levels of DeltaSB than individuals without mutation. Among healthy subjects, gender difference in DeltaSB values was observed only in individuals with the wild type of UGT1A1, but not in those with mutations in this gene. CONCLUSION: The results of the present study suggest that UGT1A1 mutation has a role in the development of high-grade fasting hyperbilirubinemia after caloric restriction.

Adult↗
PubMed2001

Useful polyclonal antibodies against synthetic peptides corresponding to immunoglobulin light chain constant region for immunohistochemical detection of immunoglobulin light chain amyloidosis.

For the immunohistochemical detection of immunoglobulin (Ig) light chain amyloidosis on formalin-fixed, paraffin-embedded tissue sections, we prepared polyclonal antibodies against synthetic peptides corresponding to positions 118-134 of Ig lambda light chain and positions 116-133 of Ig kappa light chain. Nineteen cases of systemic Ig lambda light chain amyloidosis (Alambda amyloidosis), 10 cases of systemic Ig kappa light chain amyloidosis (Akappa amyloidosis), one case of immunohistochemically unclassified systemic amyloidosis and five cases of localized Alambda amyloidosis were tested with these antibodies. Anti-lambda (118-134) antiserum and the affinity-purified antibody both reacted with 18 of the 19 cases of systemic Alambda amyloidosis and all cases of localized Alambda amyloidosis, although the immunoexpression was somewhat variable in intensity in different areas within the same specimen in both systemic and localized amyloidosis. The signal intensities in plasma cells and serum reacted for anti-lambda (118-134) antiserum were weaker than signals obtained with commercially available anti-Ig lambda light chain antibodies. Anti-kappa (116-133) antiserum and the affinity-purified antibody reacted with nine of the 10 cases of systemic Akappa amyloidosis. We conclude that these antibodies against synthetic peptides corresponding to the Ig light chain constant region are useful for the classification of amyloidosis on formalin-fixed, paraffin-embedded tissue sections.

Amyloidosis↗
PubMed2001

Maturation of macrophages from peripheral blood monocytes in Kawasaki disease: immunocytochemical and immunoelectron microscopic study.

Kawasaki disease (KD) is regarded as a cytokine-associated disorder. Despite intensive investigation into the etiology of KD, this remains unclear, although monocytes and macrophages are thought to play an important role. We examined peripheral blood monocytes using a monoclonal antibody, PM-2K, which recognizes mature macrophages but not monocytes. This study was conducted in 12 patients with KD, three patients with sepsis and 12 control subjects. Approximately 8% of whole peripheral blood monocytes from patients with acute KD were observed to be PM-2K positive. Approximately 15-20% of peripheral blood CD14+ monocytes from these patients were positive for PM-2K antibody (as determined by immunoelectron microscopy). PM-2K-positive monocytes had significantly fewer numbers of intracytoplasmic peroxidase-positive granules than monocytes from control subjects. In contrast, PM-2K-negative monocytes from patients with acute KD had a significantly greater number of peroxidase-positive granules in the cytoplasm than in those from controls. Monocytes from patients with sepsis displayed PM-2K immunocytochemical staining, similar to that in monocytes from patients with KD. These results suggest that during the acute stage of KD, monocytes partly differentiate into macrophages in the peripheral circulation.

Cellular Senescence↗
PubMed2001

Vascular amyloid of unknown origin and senile transthyretin amyloid in the lung and gastrointestinal tract of old age: histological and immunohistochemical studies.

The histological and immunohistochemical characteristics and the incidence of amyloid deposits in the tissues of the lung and gastrointestinal tract were investigated in 64 autopsied individuals who were 80 years and older (age range: 80-92 years; mean: 83.3 years). Immunohistochemical examination was performed with antibodies against amyloid A, transthyretin, immunoglobulin lambda and kappa light chain amyloid fibril proteins, beta2-microglobulin, beta protein, apolipoprotein AI, apolipoprotein AII, atrial natriuretic peptide, apolipoprotein E, and amyloid P component. Transthyretin amyloid fibril protein (ATTR) deposits were observed in five cases (7.8%). Gastrointestinal amyloid deposits of unknown origin were observed in the veins of the gastrointestinal tract in 26 cases (40.6%). This amyloid was regarded as portal amyloid with respect to distribution pattern. Pulmonary vascular amyloid deposits of unknown origin were observed in 12 cases (18.8%). These amyloid deposits were found mainly in medium-sized veins in the lungs and did not react with any antibodies against amyloid fibril proteins except apolipoprotein E and amyloid P component. Eleven of the 26 cases (42.3%) showing portal amyloid also showed pulmonary vascular amyloid of unknown origin. The pulmonary vascular amyloid deposits were similar to the portal amyloid deposits with respect to their morphological features and their relation to elastic fibers in the vessels. Further morphological investigation and biochemical analysis of the pulmonary vascular amyloid and portal amyloid will resolve questions of their origins and relation.

Aged↗
PubMed2001

Comparative immunochemical study of lectin-binding sites and cytoskeletal filaments in static and reactive mesothelium and adenocarcinoma.

In cytological preparations, reactive mesothelial cells (RMC) in serous effusions are sometimes difficult to distinguish from adenocarcinoma cells (AC). RMC and AC can be distinguished by lectin-binding patterns, but the pattern of binding of lectins to normal mesothelium is not well defined. We investigated the expression of cytoskeletal filaments, cytokeratin (CK) and vimentin (VM), and the cell surface binding pattern of 10 lectins (HPA, SBA, ABA, DSA, PNA, RCA-I, UEA-I, LTA, WGA and ConA) in the serosa of 48 adenocarcinoma specimens. We also investigated the usefulness of six lectins (HPA, SBA, RCA-I, UEA-I, LTA and WGA) in identification of RMC and AC in 16 serous effusions. DSA reactivity was significantly higher (P < 0.05) in static mesothelial cells (SMC) than in RMC. Reactivity for LTA and ConA was significantly lower (P < 0.05) in SMC than in RMC. Anti-CK and anti-VM immunoreactivity was always positive in RMC and almost negative in SMC. In serous effusions, HPA, SBA and UEA-I binding was evident in 100, 88 and 81% of AC, respectively. Little to no binding of HPA, SBA or UEA-I was detected in RMC. Our results suggest that the morphological differences between SMC and RMC are likely to be due to differences in cytoskeletal composition, with accompanying changes in cell-surface lectin-binding patterns. HPA, SBA and UEA-I are likely to be useful markers for identification of RMC and AC in cytology.

Adenocarcinoma↗
PubMed2001

Localized malignant mesothelioma of the pleura.

We describe a case of malignant pleural mesothelioma appearing as a solitary pleural tumor in a 56-year-old Japanese man with no history of exposure to asbestos. A chest radiograph revealed an isolated extrapulmonary mass in the left hemithorax. The patient underwent tumor resection, but the tumor later recurred on the contralateral pleura. The patient developed cerebral metastases and died 16 months after the initial surgery. The resected tumor was sessile with broad-based pleural attachment. Microscopically, the tumor was composed of interlacing fascicles of plump spindle cells intermixed with few polygonal cells. Most of the tumor cells showed positive immunoreactivity for cytokeratins (AE1 and AE3) and vimentin. Many of the tumor cells were positive for epithelial membrane antigen, and a few were positive for desmin. In contrast, the tumor cells were consistently negative for carcinoembryonic antigen, epithelial antigen BerEP4, calretinin, S-100 protein, neuron-specific enolase, muscle actin antigen HHF35, alpha-smooth muscle actin antigen and CD34. Ultrastructurally, the tumor cells had diffusely distributed cytoplasmic intermediate filaments, desmosome-like junctions, and a few microvilli. Some tumor cells contained cytoplasmic tonofilaments. Immunohistochemical and ultrastructural findings supported the mesothelial nature of the tumor, and led us to diagnose this tumor as a sarcomatoid localized malignant mesothelioma.

Brain Neoplasms↗
PubMed2001

Mutations affecting nerve attachment of Caenorhabditis elegans.

Using a pan-neuronal GFP marker, a morphological screen was performed to detect Caenorhabditis elegans larval lethal mutants with severely disorganized major nerve cords. We recovered and characterized 21 mutants that displayed displacement or detachment of the ventral nerve cord from the body wall (Ven: ventral cord abnormal). Six mutations defined three novel genetic loci: ven-1, ven-2, and ven-3. Fifteen mutations proved to be alleles of previously identified muscle attachment/positioning genes, mup-4, mua-1, mua-5, and mua-6. All the mutants also displayed muscle attachment/positioning defects characteristic of mua/mup mutants. The pan-neuronal GFP marker also revealed that mutants of other mua/mup loci, such as mup-1, mup-2, and mua-2, exhibited the Ven defect. The hypodermis, the excretory canal, and the gonad were morphologically abnormal in some of the mutants. The pleiotropic nature of the defects indicates that ven and mua/mup genes are required generally for the maintenance of attachment of tissues to the body wall in C. elegans.

Animals↗
PubMed2001

Polycation graft copolymers accelerating DNA strand exchange: involvement of ionic interaction.

In the previous study (Chem. Eur. J., 7, 176 (2001)) we demonstrated that the comb-type polycationic copolymer (PLL-g-Dex) which is composed of poly (L-lysine) backbone and dextran graft chains expedited the DNA strand exchange reaction. In this study, fluorescence resonance energy transfer (FRET) was employed to explore the copolymer-mediated DNA strand exchange with higher time-resolution. To initiate strand exchange reaction the duplex prepared from 3'-fluorescein isothiocyanate (FITC)--and 5'-carboxytetramethylrhodamine(TAMRA)-labeled complementary DNAs was added to its non-labeled complementary single strand. DNA strand exchange was monitored by observing the recovery of the FITC quenching. More than 20,000 times increase in strand exchange rate at 37 degrees C by the copolymer was estimated. To investigate the accelerating mechanisms of the copolymer, the same reactions but at various ionic strenghts were studied. With increasing ionic strengths the strand exchange rate in the absence of the copolymer increased, suggesting that ionic repulsion among DNAs is unfavorable for the strand exchange to occur. Hence, alleviation of the electrostatic repulsion through interpolyelectrolyte complex formation is probably a role of the copolymer for accelerating the strand exchange reaction.

DNA↗
PubMed2001

Protein/oligonucleotide conjugates as a cell specific PNA carrier.

We have focused on proteineus ligand conjugate with oligonucleotides (ODNs) as a cell-specific delivery vector for peptide nucleic acids (PNAs). Asialofetuin (AF), a hepatocyte-specific proteineus ligand, was conjugated with ODNs that served as binding sites for PNAs. Succinimidyl-transe-4(N-maleimidylmethyl)-cyclohexane-1-carboxylate (SMCC) modified AF was coupled with 5'-thiolated oligodeoxynucleotide (HS-ODN). The resulting conjugate held PNAs with sequence-specific manner. The PNA/DNA conjugate complex has resistance against nucleases in serum. The efficient release of PNA from the complex was observed when the complex was made in contact with a target nucleotide. PNA uptake to hepatocytes was greatly enhanced when hepatocytes was incubated with PNA/conjugate complex. Free AF thoroughly inhibited PNA uptake with the conjugate, evidencing asialoglycoprotein receptor (ASGP-R) mediated endocytosis to be a major-route for the cellular uptake.

Animals↗
PubMed2001

Degenerated neurilemoma (ancient schwannoma).

We report a case of degenerated neurilemoma in a 65-year-old Japanese woman. A tumor in the right thigh histopathologically showed a mixture of Antoni type A and B patterns, and contained prominent blood vessels with hyalinized walls, but no calcification. The tumor cell nuclei of middle areas were large, hyperchromatic and often multilobed but lacked mitotic figures. The mucinous areas had few tumor cells. Antoni type A and B areas were S-100 protein positive, while mucinous areas and hyalinized vessels were negative.

Aged↗
PubMed2001

Sentinel lymph nodes study in 30 cases of melanoma.

In 30 cases of melanoma, we attempted to detect sentinel lymph nodes using 1-2% patent blue dye and were able to detect them in 27 cases (90%): 19 cases out of 21 cases in the groin area (90%), 5 out of 5 cases in the axilla area (100%), and 3 out of 4 cases in the neck area (75%). The numbers of sentinel lymph nodes were one in 16 cases, two in 7 cases, three in 2 cases, and four in 2 cases. The cases with three and four nodes were all in the groin area. In 22 cases, tumor metastasis was negative in sentinel lymph nodes. Sentinel lymph nodes were detected in 36 out of 174 samples, and tumor metastasis was negative except in these sentinel lymph nodes (false negative 0%). In the groin area, sentinel lymph nodes were located around the femoral and great saphenous vein junction. In the axilla area, sentinel lymph nodes were located in the central, lateral and subscapular lymph nodes. In the head and neck area, sentinel lymph nodes were found in the submandibular and occipital lymph nodes. The positions of sentinel lymph nodes differed a little with tumor location. By accumulating cases, it should become possible to predict the positions of sentinel lymph nodes before operations. Sentinel lymph node biopsy is easy and requires only a small incision.

Adult↗
PubMed2001

Analysis of the vascularity of an atelocollagen sponge substitute dermis in the human.

Atelocollagen sponge had been developed as a new beneficial dermal substitute. However, histologic studies of the vascularity of an atelocollagen sponge substitute dermis in humans are very rare. The purpose of this study was to determine the distribution of blood vessels in an artificial dermis made of atelocollagen affixed to human tissue. The artificial dermis was used in 21 cases and at 23 sites, and biopsies were performed. The samples were stained with anti-human blood type A, B, and H monoclonal antibodies. Angiogenesis in the upper layer of the artificial dermis was recognizable within 14 days after affixing the artificial dermis. We concluded that a skin graft taken 14 days after affixing the artificial dermis would be successful, even though the granulation tissue formation might not be completed and the color would most likely remain white. The lower layer of the angiogenesis spread in a horizontal direction like an expanded network of blood vessels, while the middle and upper layers of angiogenesis spread perpendicularly to form upward growth when two cases were examined in various spots. The data suggested that the sponge construction of artificial dermis not only became a scaffold for the invasion of collagen, but it also facilitated construction of near normal dermis with regard to the construction of blood vessels.

Adult↗
PubMed2001

Thallium-201 single photon emission computed tomography (SPECT) in patients with duchenne's progressive muscular dystrophy: a histopathologic correlation study.

The pathomorphologic mechanism responsible for abnormal perfusion imaging during thallium-201 myocardial single photon emission computed tomography (201Tl-SPECT) in patients with Duchenne's progressive muscular dystrophy (DMD) was investigated. Hearts from 7 patients with DMD were evaluated histopathologically at autopsy and the results correlated with findings on initial and delayed resting 201Tl-SPECT images. The location of segments with perfusion defects correlated with the histopathologically abnormal segments in the hearts. Both the extent and degree of myocardial fibrosis were severe, especially in the posterolateral segment of the left ventricle. Severe transmural fibrosis and severe fatty infiltration were common in segments with perfusion defects. In areas of redistribution, the degree of fibrosis appeared to be greater than in areas of normal perfusion; and intermuscular edema was prominent. Thus, the degree and extent of perfusion defects detected by 201Tl-SPECT were compatible with the histopathology. The presence of the redistribution phenomenon may indicate ongoing fibrosis. Initial and delayed resting 201Tl-SPECT images can predict the site and progress of myocardial degeneration in patients with DMD.

Adolescent↗