Search PubMed⌕ Search

Biomedical subjects

H Shindo

Publications and source records attributed to H Shindo.

At least 37 records · Page 2Linked to original sources
PubMed2000

High-speed counter-current chromatography of apple procyanidins.

Apple procyanidins were separated by high-speed counter-current chromatography using a type-J multilayer coil planet centrifuge. Several two-phase solvent systems with a wide range of hydrophobicities from a non-polar hexane system to polar n-butanol systems were evaluated their performance in terms of the partition coefficient and the retention of the phase. The best separation of procyanidins B and C was achieved with a two-phase solvent system composed of n-butanol-methyl tert.-butyl ether-acetonitrile-0.1% trifluoroacetic acid (2:4:3:8) using the lower phase as a mobile at a flow-rate of 1.0 ml/min.

Biflavonoids↗
PubMed2000

Destabilization of nucleosomes by an unusual DNA conformation adopted by poly(dA) small middle dotpoly(dT) tracts in vivo.

Poly(dA) small middle dotpoly(dT) tracts are common and often found upstream of genes in eukaryotes. It has been suggested that poly(dA) small middle dotpoly(dT) promotes transcription in vivo by affecting nucleosome formation. On the other hand, in vitro studies show that poly(dA) small middle dotpoly(dT) can be easily incorporated into nucleosomes. Therefore, the roles of these tracts in nucleosome organization in vivo remain to be established. We have developed an assay system that can evaluate nucleosome formation in yeast cells, and demonstrated that relatively longer tracts such as A(15)TATA(16) and A(34) disrupt an array of positioned nucleosomes, whereas a shorter A(5)TATA(4) tract is incorporated in positioned nucleosomes of yeast minichromosomes. Thus, nucleosomes are destabilized by poly(dA) small middle dotpoly(dT) in vivo in a length-dependent manner. Furthermore, in vivo UV footprinting revealed that the longer tracts adopt an unusual DNA structure in yeast cells that corresponds to the B' conformation described in vitro. Our results support a mechanism in which a unique poly(dA) small middle dot poly(dT) conformation presets chromatin structure to which transcription factors are accessible.

Base Sequence↗
PubMed2000

Long-term results of fractionated strontium-90 radiation therapy for pterygia.

PURPOSE: The long-term safety and effectiveness of fractionated strontium-90 radiation therapy (RT) for pterygium were reviewed retrospectively. METHODS AND MATERIALS: Between 1984 and 1996, 399 patients with 490 pterygia were treated with a strontium-90 eye applicator following surgical removal of the pterygium. The median follow-up period was 61 months (range 2-178). Of the 490 pterygia, 452 were fresh, 17 were recurrences after surgical removal alone, and 21 were recurrences after surgical removal plus postoperative RT. Fractionated RT of 31-42 Gy/4-5 fractions/22-29 days was given for 95.1% of the pterygia. RESULTS: In total, 58 (11.8%) local recurrences of pterygia were noted. The median time of local recurrences was 10 months, ranging from 2 to 93 months, and 16 recurrences (28%) were noted later than 24 months after treatment. The interval between surgery and the start of RT (1-3 days vs. >3 days) and recurrent pterygia were significant variables for local control in the multivariate analysis, while total RT dose (7-29 Gy vs. 31-50 Gy) was a marginally significant variable. Late toxicities that may be associated with strontium-90 RT were scleromalacia (scleral thinning) in 4 eyes, adhesion of eyelids in 3 eyes, and scleral ulcer in 2 eyes. CONCLUSION: Fractionated strontium-90 RT of approximately 40 Gy/4-5 fractions was safe and effective for preventing recurrence of pterygia, when RT was started within 3 days of surgery.

Adult↗
PubMed2000

Etiologic factors in femoral head osteonecrosis in growing rats.

Osteonecrosis of the epiphyseal nucleus of the femoral head, which resembles that in Perthes' disease in children, was studied in spontaneously hypertensive rats (SHRs) and ordinary Wistar Kyoto rats (WKYs). The SHRs were kept in ordinary cages and the WKYs in custom-made high cages from 5 weeks to 15 weeks after birth. The WKYs had to stand on their hind limbs to feed because the feed box and the drinking aperture were placed at a high level. At 15 weeks, the femurs were resected and examined radiographically and histologically. There was a relatively high incidence of avascular necrosis in the epiphyseal nuclei of the femoral heads; 45.8% in SHRs and 33.3% in WKYs. In the SHRs, there were ossification disturbances of the proximal femoral epiphysis and deformities of the proximal femurs, such as a flattened femoral head and short neck. These findings suggest that constitutional cartilage disorder in the SHRs and excessive mechanical stress on the femoral heads in the WKYs participated in the etiology of the osteonecrosis.

Animals↗
PubMed2000

Apoptosis of articular chondrocytes in rheumatoid arthritis and osteoarthritis: correlation of apoptosis with degree of cartilage destruction and expression of apoptosis-related proteins of p53 and c-myc.

To investigate the relationship of chondrocyte apoptosis and cartilage destruction, we performed in situ nick end labeling (ISNEL), electron microscopy, and immunohistochemistry against apoptosis-related proteins, p53 and c-myc, in the articular cartilages of patients with rheumatoid arthritis (RA; n = 12) and osteoarthritis (OA; n = 12), and in control articular cartilages from patients with femoral neck fracture (n = 8). The distribution of stained chondrocytes was evaluated semiquantitatively in relation to the degree of cartilage destruction. ISNEL-positive chondrocytes with apoptotic morphological features were identified in a relatively early phase of cartilage destruction, and correlated positively and significantly in a number with the degree of cartilage degeneration. Comparison of RA and OA revealed a significantly greater number of ISNEL-positive chondrocytes in RA cartilage. In contrast, the specimens of normal subjects contained few cells with apoptotic changes. Similarly to the distribution of ISNEL staining, the expression of p53 and c-myc proteins was observed in chondrocytes within the degraded lesions, and showed a positive correlation with the number of ISNEL-stained cells. These results suggest that the degree of chondrocyte apoptosis is closely related to cartilage destruction and that chondrocytes in RA more readily undergo apoptosis than those in OA. The expression of p53 and c-myc proteins in ISNEL-positive areas may reflect the involvement of these proteins in the apoptotic process in articular chondrocytes in inflammatory arthritis.

Adult↗
PubMed2000

Deformation and vascular occlusion of the growing rat femoral head induced by mechanical stress.

To investigate the effects of mechanical stress on the extent of deformation and decrease of the blood supply to the femoral head in the growing rat, we conducted a histological and microangiographic study, using a custom-made hip-joint loading device. Application of a 1-kg to 3-kg load to the hip joint caused compression not only of the articular cartilage but also of the growth plate of the femoral head. These changes were particularly prominent in the lateral portion of the femoral head. The compression also caused complete occlusion of the lateral epiphyseal arteries, representing the blood supply to the femoral head, at the point of penetration into the epiphyseal cartilage. Our results indicate that the sum of forces during mechanical load application to the hip joint in growing rats acts to deform the cartilage of the lateral side of the femoral head. Our results also indicate the presence of a mechanically weak point in the femoral head which may lead to occlusion of the lateral epiphyseal arteries.

Animals↗
PubMed2000

How amino acids control the binding of Cu(II) ions to DNA. Part III. A novel interaction of a histidine complex with DNA.

L-Histidine Cu(II) complex bound to DNA showed broad EPR signals characteristic of the aggregated Cu(II) species, which could be observed even when the molar ratio of L-histidine to Cu(II) ion was smaller than unity. The signal for the DNA fibers changed with the orientation of the fibers in the static magnetic field. Based on these results, the signal was assigned to a mono-histidine Cu(II) complex stereospecifically aggregated in a groove or along a phosphodiester chain of the double helical DNA. In contrast to the L-histidine complex, the D-histidine complex bound to DNA did not show such broad signals and the observed spectra for the complex on B-form DNA fibers at -150 degrees C were simulated assuming that the g1 axis of the mono-D-histidine complex tilts by about 55 degrees from the DNA-fiber axis. Addition of some deoxy-nucleotides, but not deoxy-nucleosides, to a solution of a mono-histidine complex resulted in the formation of a dinuclear ternary complex with different structures for L- or D-histidine, suggesting the possibility that the stereospecific aggregation of the L-histidine complex on a double helical DNA was mediated by the phosphodiester backbones.

Copper↗
PubMed2000

Altered in vitro apoptosis of cultured mast cells prepared from an inbred strain of mice, NC/Kuj.

BACKGROUND: An inbred strain of mice, NC, develops dermatitis associated with highly elevated serum IgE and dermal mast cell hyperplasia. OBJECTIVES AND METHODS: To clarify the mechanisms for the dermal mast cell hyperplasia in NC, we prepared bone marrow-derived mast cells (BMMCs) from three strains of mice, NC/Kuj, C57BL/6 and BALB/c, and compare histamine contents, histamine release abilities, adhesive properties and apoptosis of the BMMCs. RESULTS: Compared with BMMCs obtained from C57BL/6 and BALB/c, NC/Kuj BMMC possessed higher histamine content and higher adhesive ability to plastic plates, although histamine release from BMMCs was found to be similar in the three strains. The most intriguing finding is the lack of apoptosis in the BMMCs from NC/Kuj upon growth factor deprivation as determined by DNA ladder formation and loss of membrane phospholipid asymmetry. CONCLUSION: The altered in vitro properties of mast cells in NC/Kuj partially account for an increase of dermal mast cells, which might be involved in the development of skin lesions in NC.

Animals↗
PubMed2000

Activation of specific MEK-ERK cascade is necessary for TGFbeta signaling and crosstalk with PKA and PKC pathways in cultured rat articular chondrocytes.

OBJECTIVE: TGFbeta is a potent stimulator of cell growth in cultured rat articular chondrocytes (CRAC). The stimulatory effect is mediated through the immediate induction of c-fos gene by activating ERK of MAPK. The present study was undertaken to investigate the upstream regulators involved in TGFbeta-induced ERK activation in CRAC and to compare the results with the events in HepG2 cells. RESULTS: In vitro kinase and trans-reporting assays showed that TGFbeta preferentially activated ERK and JNK pathways in CRAC and HepG2, respectively. ERK activation in CRAC was selectively inhibited by PD98059, a MEK inhibitor. Overexpression of wild or active forms of MEKK1, the upstream activator of ERK and JNK, decreased the TGFbeta-induced 3TP-luciferase activity in CRAC. In contrast, in HepG2 dominant negative form of MEKK1 or SEK1 ligand-dependent reporter activity was diminished. Transfection of TAK1, another MAPKKK, also positively and negatively regulated 3TP transcriptional activity of HepG2 and CRAC, respectively. Activation of PKA by 8-bromo-cyclic AMP or forskolin, and inhibition of PKC by calphostin C, resulted in a significant decrease in 3TP activity as well as in vitro ERK kinase activity in CRAC. CONCLUSIONS: The results indicate that TGFbeta transduces a predominant signal pathway through MEK-ERK-Elk1, independent of MEKK1 or TAK1 pathway in CRAC. However, in HepG2, activation of MEKK1 and TAK1 is essential for TGFbeta-induced signal transmission. The results also demonstrated that in CRAC, MEK-ERK pathway activated by TGFbeta is negatively regulated by PKA cascade but transactivated by PKC.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed2000

Dexamethasone inhibition of TGF beta-induced cell growth and type II collagen mRNA expression through ERK-integrated AP-1 activity in cultured rat articular chondrocytes.

Intraarticular injection of dexamethasone (DEX) accelerates cartilage degradation due to the suppression of chondrocyte proliferation and extracellular matrix formation. The present study first demonstrated the interaction between DEX and TGF beta, a potent growth factor for cultured rat articular chondrocytes (CRAC), and then investigated the molecular mechanism by which DEX counteracts TGF beta-induced chondrocyte proliferation and differentiation through the regulation of AP-1 activity. DEX reduced serum-deprived and TGF beta-stimulated cell growth and [(3)H]-thymidine incorporation of CRAC. DEX also inhibited the expression of (alpha)1 type II collagen with concomitant suppression of the promoter activity. Transfection studies using a reporter vector with AP-1 responsive elements showed that DEX reduced TGF beta-activated but not basal luciferase activities. Activation of 3TP-luc, another AP-1 responsive element containing reporter was also blocked by DEX. GAL4-Elk1 studies revealed that DEX suppressed TGF beta-induced ERK activation which led to c-fos gene expression followed by increase in AP-1 complex formation, whereas the Smad pathway was not involved in DEX-dependent negative regulation of AP-1 in a reporter assay that requires FAST1-Smad2 for the activation. DEX also eliminated TGF beta-induced c-fos mRNA expression and ERK activation in Northern analysis and in vitro kinase assay, respectively. Further, DNA synthesis and transactivation of type II collagen by TGF beta were inhibited by PD98059, an inhibitor of MEK. Our results indicate that DEX suppressed TGF beta-induced chondrocyte proliferation and type II collagen expression, probably through selective inhibition of ERK integrated AP-1 activation.

Animals↗
PubMed2000

Immunohistochemical localisation of protein tyrosine kinase receptors Tie-1 and Tie-2 in synovial tissue of rheumatoid arthritis: correlation with angiogenesis and synovial proliferation.

OBJECTIVE: To investigate the involvement of Tie-1 and Tie-2, receptor tyrosine kinases required for angiogenesis, in synovial proliferation and angiogenesis of rheumatoid arthritis (RA). METHODS: Synovial tissues from 10 patients with RA and three control subjects were analysed by double immunohistochemistry and reverse transcriptase polymerase chain reaction (RT-PCR). RESULTS: Expression of Tie-1 and Tie-2 was seen in all synovia, but predominantly in papillary projected portions. In synovial lining cells, Tie-2 was expressed mainly in the basal layer and frequently colocalised with vimentin and proliferating cell nuclear antigen (PCNA), whereas Tie-1 was also expressed in the superficial layer. In stromal cells, Tie-2 immunoreactivity was restricted to vimentin positive fibroblast-but not macrophage derived cells, whereas Tie-1 expression was not dependent on the phenotype. Tie receptors were also highly expressed in the endothelium and surrounding pericytes of capillaries scattered over the papillary proliferated synovium without notable difference in the expression of the two receptors. Furthermore, Tie positive vessels often overexpressed PCNA. In normal synovia, expression of Tie receptors was restricted to the capillary endothelium. RT-PCR confirmed the expression of Tie-1 and Tie-2 in RA synovial tissues and also in the cultured synoviocytes. CONCLUSION: The results suggest the possible involvement of overexpressed Tie-1 and Tie-2 in synovial lining and stromal cells in the pathophysiology of RA synovitis, probably through distinct mechanisms. Furthermore, expression of Tie receptors in actively growing vasculature may reflect the direct involvement of these receptors in angiogenesis and subsequent vascularisation.

Adult↗
PubMed2000

Deletion of the yhhP gene results in filamentous cell morphology in Escherichia coli.

The Escherichia coli yhhP gene was predicted to encode a small hypothetical protein of 81 amino acids, the cellular function of which is not known. To gain insight into the function of this uncharacterized YhhP protein, genetic and biochemical studies were done. We first tried to express and purify the YhhP protein to prepare an anti-YhhP antiserum. Western blotting showed that the hypothetical yhhP gene is indeed transcribed and translated as a minor cytoplasmic protein. YhhP-deficient (delta yhhP) cells formed colonies poorly on a rich medium (e.g., Luria-Bertani medium) containing a relatively low concentration of NaCl, while they can grow normally either in LB containing 3% NaCl or in a synthetic medium (e.g., M9-glucose). During exponential growth in rich medium, an early step of cell division was inhibited in delta yhhP cells, forming filaments. For the YhhP-deficient filamentous cells, the FtsZ-ring formation was analyzed with immunofluorescence microscopy. The FtsZ-ring formation did not occur normally in the delta yhhP filaments, although the filamentous cells contained the FtsZ protein at a certain level comparable to that in the wild-type cells. The ftsZ gene was found to function as a multicopy suppressor of the delta yhhP mutant. Another multicopy suppressor gene was identified as the dksA gene. Provided that either the ftsZ or dksA gene was introduced into the mutant cells with its multicopy state, the resulting transformants were capable of growing in rich medium, formed wild-type short rods. These results are discussed with regard to the presumed function of this ubiquitous protein.

Amino Acid Sequence↗
PubMed2000

Differential effects of IGF-binding proteins, IGFBP-3 and IGFBP-5, on IGF-I action and binding to cell membranes of immortalized human chondrocytes.

Insulin-like growth factor-I (IGF-I) is an important anabolic factor for cartilage tissue and its action is, in part, regulated by IGF-binding proteins (IGFBPs). The object of this study was to investigate the effects of IGFBPs on IGF-I action and on binding of IGF-I to cells using a reproducible immortalized human chondrocyte culture model. Treatment of the C-28/I2 cells with IGF-I or des(1-3)IGF-I in serum-free medium stimulated cell proliferation in a dose-dependent manner. However, the effect of des(1-3)IGF-I was more potent, thereby suggesting that endogenously produced IGFBPs inhibited IGF action. The stimulatory effect of IGF-I was inhibited significantly by addition of IGFBP-3 but enhanced slightly by IGFBP-5. However, neither IGFBP-3 nor IGFBP-5 had an effect on basal cell growth. Binding of (125)I-labeled IGF-I to the cells was displaced by both IGFBP-3 and IGFBP-5, although higher concentrations of unlabeled IGFBP-5 were required to displace IGF-I to the same extent as IGFBP-3. Treatment of the cells with IGF-I increased the levels of IGFBP-5 protein measured by Western ligand blotting, and stimulated a corresponding increase in IGFBP-5 mRNA while increasing type II collagen mRNA. Our findings indicate that the balance between IGFBP-3 and IGFBP-5 influences IGF receptor binding and its action on chondrocyte proliferation, and may thereby modulate cartilage metabolism.

Binding, Competitive↗
PubMed2000

Magnetization transfer measurements of the hippocampus in patients with Alzheimer's disease, vascular dementia, and other types of dementia.

BACKGROUND AND PURPOSE: Although atrophy of structures in the medial temporal lobe has been considered an indication of Alzheimer's disease (AD), atrophic changes on MR images have also been associated with other dementing diseases and are not specific to AD. This study was undertaken to determine whether characteristic alterations in the hippocampus of patients with AD are detectable with magnetization transfer (MT) imaging. METHODS: Coronal MT imaging was performed in 35 patients with probable AD, in 14 patients with vascular dementia, in 13 patients with other types of dementia, and in 23 control subjects to measure MT ratios of the hippocampus. Medial temporal lobe atrophy was graded subjectively on a five-point scale. RESULTS: Scores of medial temporal lobe atrophy in all dementia groups were significantly higher than those in control subjects, but no differences were found among the dementia groups. MT ratios in the hippocampus were significantly lower in patients with AD than in those with non-AD dementia and in the control subjects; however, no differences were found between the non-AD dementia patients and the control subjects. MT ratio measurements were better than visual analysis of atrophy for differentiating AD patients from those with non-AD dementia (an overall discrimination rate of 77% versus 65%). MT ratios significantly correlated with scores on the Mini-Mental State Examination and with medial temporal lobe atrophy in AD patients but not in patients with non-AD dementia. CONCLUSION: MT measurements may be more specific than visual analysis in detecting structural damage of the hippocampus in AD patients and might be useful in discriminating AD from vascular dementia and other types of dementia.

Aged↗
PubMed1999

Triplex formation of chemically modified homopyrimidine oligonucleotides: thermodynamic and kinetic studies.

We have investigated effects of chemical modifications of a third strand on the thermodynamic and kinetic properties of the triplex formation between a 23-bp duplex and each of four kinds of 15-mer chemically modified third strands using isothermal titration calorimetry and interaction analysis system. The chemical modifications of the third strand included one base modification, with replacement of thymine by uracil; two sugar moiety modifications, RNA and 2'-O-methyl-RNA; and one phosphate backbone modification, with replacement of phosphodiester by phosphorothioate backbone. The thermodynamic and kinetic parameters obtained were similar in magnitude at room temperature for the triplex formation with the base-modified and the sugar-modified third strands. By contrast, binding constant for the triplex formation with the third strand containing phosphorothioate backbone was much smaller by a factor of 10 than that for the other triplex formations. Kinetic analyses have also demonstrated that the third strand containing phosphorothioate backbone was much slower in the association step and much faster in the dissociation step than the other third strands, which resulted in the much smaller binding constant. The reason for the instability of the triplex with the third strand containing phosphorothioate backbone will be discussed. We conclude that, at least in the triplex formation with the chemically modified third strands studied in the present work, the modification of phosphate backbone of the third strand produces more significant effect on the triplex formation than the modifications of base and sugar moiety.

Base Sequence↗
PubMed1999

Diffusion-weighted and magnetization transfer imaging of the corpus callosum in Alzheimer's disease.

We investigated structural changes of the corpus callosum in patients with Alzheimer's disease (AD) using sagittal diffusion-weighted (DW) and magnetization transfer (MT) imaging. Patients with AD (n=23) had a significantly decreased area only in the posterior portion of the corpus callosum. Apparent diffusion coefficient (ADC) values perpendicular to the commisural fiber orientation were significantly higher in the anterior portion of the corpus callosum without definite atrophy, as well as in the posterior portion with significant atrophy, in patients with AD than in controls (n=16) and thus diffusion in these regions showed a significantly lower degree of anisotropy in patients than in controls. MT ratios were also significantly lower in patients with AD in the anterior and posterior portions of the corpus callosum than in controls. These findings probably reflect structural changes in the corpus callosum including axonal loss and/or demyelination. DW and MT imagings may be useful in detecting degeneration of the corpus callosum in AD.

Aged↗
PubMed1999

Identification of the DNA binding surface of H-NS protein from Escherichia coli by heteronuclear NMR spectroscopy.

The DNA binding domain of H-NS protein was studied with various N-terminal deletion mutant proteins and identified by gel retardation assay and heteronuclear 2D- and 3D-NMR spectroscopies. It was shown from gel retardation assay that DNA binding affinity of the mutant proteins relative to that of native H-NS falls in the range from 1/6 to 1/25 for H-NS(60-137), H-NS(70-137) and H-NS(80-137), whereas it was much weaker for H-NS(91-137). Thus, the DNA binding domain was defined to be the region from residue A80 to the C-terminus. Sequential nuclear Overhauser effect (NOE) connectivities and those of medium ranges revealed that the region of residues Q60-R93 in mutant protein H-NS(60-137) forms a long stretch of disordered, flexible chain, and also showed that the structure of the C-terminal region (residues A95-Q137) in mutant H-NS(60-137) was nearly identical to that of H-NS(91-137). 1H and 15N chemical shift perturbations induced by complex formation of H-NS(60-137) with an oligonucleotide duplex 14-mer demonstrated that two loop regions, i.e. residues A80-K96 and T110-A117, play an essential role in DNA binding.

Amino Acid Sequence↗