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Biomedical subjects

K Shimizu

Publications and source records attributed to K Shimizu.

At least 289 records · Page 16Linked to original sources
PubMed2000

Blockade of TGF-beta by in vivo gene transfer of a soluble TGF-beta type II receptor in the muscle inhibits corneal opacification, edema and angiogenesis.

Accumulating evidence suggests the involvement of TGF-beta in the process of corneal opacity, which is one of the serious causes of visual loss. However, whether TGF-beta is indeed critical for the pathogenesis remains unknown. We constructed an adenovirus expressing an entire ectodomain of the human type II TGF-beta receptor fused to Fc portion of human IgG (AdTbeta-ExR): this soluble receptor is secreted from AdTbeta-ExR-infected cells, binds to TGF-beta and inhibits TGF-beta signaling. When AdTbeta-ExR was injected into the femoral muscle of Balb/c mice, a high level of the soluble receptor protein (2.0-3.5 x 10(3) pM) was detectable in the serum and in the ocular fluid for at least 10 days. In the mice subjected to corneal injury with silver nitrate and to intramuscular injection with either saline or a control adenovirus expressing beta-galactosidase (AdLacZ), corneal opacification composed of extracellular matrix (ECM) accumulation, of infiltration of neutrophils and monocytes/macrophages, and of angiogenesis were all induced. In contrast, they were markedly reduced in the mice injected with AdTbeta-ExR. Immunohistochemical analysis revealed that TGF-beta, fibronectin, macrophage chemoattractant protein-1, and vascular endothelial growth factor were densely stained in the edge of wounded cornea, but they were scarcely present in the injured-cornea of AdTbeta-ExR-treated mice. Our results demonstrate that TGF-beta indeed plays a critical role in the process of cornea opacification, and that adenovirus-mediated expression of a soluble TGF-beta receptor can be therapeutically useful.

Adenoviridae↗
PubMed2000

The effect of sofalcone on indomethacin-induced gastric ulcers in a Helicobacter pylori-infected gnotobiotic murine model.

BACKGROUND: Sofalcone has been reported to exert anti-ulcer and gastroprotective actions, but its exact mechanism of action remains unknown. In our laboratory, we found that indomethacin-induced gastric ulcers become worse when associated with Helicobacter pylori infection. METHODS: We employed the H. pylori-infected gnotobiotic murine model to examine the effect of sofalcone on indomethacin-induced gastric ulcers in the presence of H. pylori infection. In vitro experiments were also done to evaluate the effects of sofalcone on H. pylori growth, adherence of H. pylori to the MKN45 cells (a human gastric epithelial cell line), and these cells' IL-8 production in the presence of H. pylori. RESULTS: We found that sofalcone produced a significant improvement in ulcer size as well as a substantial reduction in the number of H. pylori colonies in H. pylori-infected gnotobiotic mice. In vitro sofalcone has a significant bacteriocidal effect against H. pylori and can also significantly prevent adherence of this bacterium to MKN45 cells, thus remarkably reducing IL-8 production of these cells in response to stimulation by H. pylori. CONCLUSION: Our results suggest that sofalcone can improve ulcer healing by the mechanisms mentioned above.

Animals↗
PubMed2000

Extrachromosomal transposition of the transposable element Minos occurs in embryos of the silkworm Bombyx mori.

To assess the ability of the transposable element Minos to act as a vector for genetic manipulation of the silkworm Bombyx mori, an extrachromosomal transposition assay based on three plasmids was performed. The three plasmids - helper, donor and target - were co-injected into preblastoderm embryos. Low molecular weight DNA was extracted from the embryos at the stage of blastokinesis and used to transform Escherichia coli. High frequency of transposition was observed in the presence of a helper plasmid possessing an intronless Minos transposase gene, whereas transposition did not occur in the presence of a helper plasmid with the intron-bearing transposase gene. Sequence analysis of the insertion sites showed that Minos always inserts into a TA dinucleotide. Although the insertions are distributed throughout the target gene, there was a preference for certain insertion sites. However, no consensus could be identified in the sequence flanking the target site. The results strongly suggest that the transposable element Minos has the potential to be used as a vector in the silkworm and probably in other lepidopteran insects.

Animals↗
PubMed2000

Relationship of decrease in fecundity with advancing age to structural changes in mouse endometrium.

The aim of this study was to determine whether a relationship exists between decrease in fecundity and structural changes in the antimesometrial endometrium of the mouse. Fecundity was calculated as the number of animals showing a placental sign/number of copulated animals x 100 (%). Structural changes in the endometrium were examined by electron microscopy. A negative correlation between age and fecundity was found. Fecundity was 50% at 7 mo of age. At this age, amorphous material appeared in the region between the basement membrane deep to the luminal epithelium and the subepithelial cells. This material was sometimes attached to the basement membrane. It increased in amount with advancing age, as fecundity decreased. The structure of the uterine luminal epithelial cells did not alter with age. The results indicated that decrease in fecundity with advancing age is correlated with the appearance of amorphous material beneath the basal lamina of the endometrial epithelium. It is suggested that this could impair communication between the luminal epithelium and the endometrial stroma, which plays an important role in implantation.

Aging↗
PubMed2000

Preferential down-regulation of phospholipase C-beta in Ewing's sarcoma cells transfected with antisense EWS-Fli-1.

EWS-Fli-1, a fusion gene found in Ewing's sarcoma and primitive neuro-ectodermal tumour (PNET), encodes a transcriptional activator and promotes cellular transformation. We have made stable Ewing's sarcoma cells expressing antisense EWS-Fli-1 transcripts by transfecting the antisense EWS-Fli-1 expression plasmid. These cells showed partial loss of endogenous EWS-Fli-1 proteins and suppression of the cell growth. To elucidate the molecular mechanisms underlying the growth inhibition, we examined the changes of signal transducing proteins by immunoblot analysis in Ewing's sarcoma cells stably expressing antisense EWS-Fli-1 transcripts. Western blotting of the cell proteins revealed that expressions of phospholipase Cbeta2 and beta3 (PLCbeta2, PLCbeta3), and also protein kinase C alpha and beta (PKCalpha, beta) were significantly reduced by transfecting with antisense EWS-Fli-1. The inositol phosphates production by bradykinin (BK), but not platelet-derived growth factor (PDGF), was suppressed in these cells. These results suggest that the PLCbeta2 and PLCbeta3 may play a role in tumour proliferation in Ewing's sarcoma cells.

DNA, Antisense↗
PubMed2000

Inhibition of tyrosinase by flavonoids, stilbenes and related 4-substituted resorcinols: structure-activity investigations.

Several flavonoids, stilbenes and related 4-substituted resorcinols, obtained from Artocarpus incisus and other plants or synthesized, were tested for their inhibitory activity against tyrosinase. The structure-activity relationships suggested that specific natural or synthesized compounds having the 4-substituted resorcinol skeleton have potent tyrosinase inhibitory ability. Kinetic studies have indicated that specific compounds having the 4-substituted resorcinol skeleton exhibit competitive inhibition of the oxidation of DL-beta-(3,4-dihydroxyphenyl)alanine (DL-DOPA) by mushroom tyrosinase. These findings could lead to the design and discovery of new tyrosinase inhibitors.

Enzyme Inhibitors↗
PubMed2000

The 5 alpha-reductase inhibitory components from heartwood of Artocarpus incisus: structure-activity investigations.

The methanol extract of heartwood of Artocarpus incisus showed potent 5 alpha-reductase inhibitory activity. We investigated the 5 alpha-reductase inhibitory effects of nine compounds isolated from A. incisus. Chlorophorin (IC50 = 37 microM) and artocarpin (IC50 = 85 microM) showed more potent inhibitory effects than did alpha-linolenic acid, which is known as a naturally occurring potent inhibitor. Structure-activity investigations suggested that the presence of an isoprene substituent (prenyl and geranyl) would enhance 5 alpha-reductase inhibitory effects.

5-alpha Reductase Inhibitors↗
PubMed2000

A classification of circadian blood pressure profiles using a cumulative chi-square technique.

Non-invasive ambulatory blood pressure monitoring has been used to assess circadian rhythms in blood pressure in a qualitative fashion. However, there are no established methods for assessing circadian changes in blood pressure in a quantitative fashion. In this study, we developed a quantitative method to evaluate the circadian rhythm based on profile analysis and a cumulative chi-square technique. This method was used to analyse the circadian blood pressure variations in 100 normotensive volunteers and 127 hypertensive patients. Three blood pressure profiles were identified for the normotensive group, while four were identified for the hypertensive group. Furthermore, there was a relationship between the discriminate blood pressure profiles and the severity of hypertension. We conclude that profile analysis of 24-h ambulatory blood pressure monitoring may be used to stratify patients with regard to the risk of complications of hypertension.

Adult↗
PubMed2000

A subpopulation of fibroblast growth factor-2-binding heparan sulfate is lost in human papillary thyroid carcinomas.

We hypothesized that a change in composition of proteoglycans can regulate the bioactivity of fibroblast growth factor (FGF)-2 in the thyroid. In order to test this hypothesis, we established a simple and sensitive method for detecting FGF-2-binding heparan sulfates and characterized them in papillary thyroid carcinomas and normal thyroids. The thyroid extracts were applied to a Q-Sepharose anion exchange column. After the column was washed with 10 mM of phosphate buffer, 1 microgram of human recombinant FGF-2 was added onto the column. The column was eluted with a gradient of NaCl (0.3-1.5 M). Each fraction was blotted onto nitrocellulose membrane. Immunoreactivity of heparan sulfate and FGF-2 was revealed by the incubation of membranes with the specific antibodies, and quantitatively estimated by measuring the density of the color product. In normal thyroids, immunoreactivity of heparan sulfate was detected as two peaks at 0.7 and 0.9 M of NaCl. Heparan sulfate-containing fractions also showed FGF-2 immunoreactivity, indicating the complex formation of FGF-2 and heparan sulfate. In papillary thyroid carcinomas, immunoreactivity of heparan sulfate showed various elution profiles on Q-Sepharose chromatography, including single peak at 0.7 M of NaCl and the one similar to that of the normal thyroids. However, FGF-2 immunoreactivity was detected only in the fractions eluting at 0.7 M of NaCl. This loss of a subpopulation of FGF-2-binding heparan sulfate in human papillary thyroid carcinomas may lead to the increase of free FGF-2 bioavailable in extracellular matrix.

Anion Exchange Resins↗
PubMed2000

Identification of remains by sequencing of mitochondrial DNA control region.

The maternity of two newborns who were murdered and abandoned >5 and 10 years were analyzed by amplification and direct sequencing of mitochondrial DNA (mtDNA) control regions. Sequences of two hypervariable segments from each femur bone sample and the blood of the putative mother showed four mutations in hypervariable region I and two mutations in addition to two nucleotide insertions in hypervariable region II compared with the reference sequence, and all sequences were identical. The genotype of these individuals is found to be relatively rare in the Japanese population, and it was strongly suggested that both sets of newborn remains really were children of the putative mother. Sexes of the remains were determined to be female and male by amplifying a segment of the X-Y homologous gene, amelogenin. These results demonstrate that sequencing of mtDNA is a useful tool for genetic identification of aged and decomposed materials.

Amelogenin↗
PubMed2000

Tc-99m PMT hepatobiliary scintigraphy in the differential diagnosis of extrahepatic metastases and hepatocellular carcinoma.

PURPOSE: The authors evaluated the utility of hepatobiliary scintigraphy for tissue characterization of extrahepatic metastases from hepatocellular carcinoma (HCC) using Tc-99m N-pyrydoxyl-5-methyltriptophane (Tc-99m PMT). METHODS: We examined 13 patients with HCC (29 extrahepatic metastases and 3 benign bone lesions) and 5 patients with other cancers (15 extrahepatic metastases). Thirty minutes to 6 hours after intravenous administration of Tc-99m PMT, planar (all 47 lesions) and SPECT (42 lesions) images were obtained. Accumulation of Tc-99m PMT in the lesion was evaluated visually by comparing bone scintigraphy, computed tomography, magnetic resonance imaging, or all of these. RESULTS: Findings were positive in 12 of 13 patients with HCC and extrahepatic metastases (16 of 29 on planar imaging and 21 of 26 on SPECT). Findings in all three benign bone lesions and 15 metastatic lesions from non-HCC primary lesions were negative (0 of 18 on planar imaging, 0 of 16 on SPECT). There were no false-positive findings in these lesions. Lesion-by-lesion sensitivity, specificity, accuracy, and positive and negative predictive values were 55%, 100%, 72%, 100%, and 58% by planar imaging and 81%, 100%, 88%, 100%, and 76% by SPECT, respectively. CONCLUSIONS: Because of the high specificity and reasonable sensitivity, Tc-99m PMT appears to be useful for the differential diagnosis of extrahepatic metastases from HCC. SPECT improves the detectability of small or faint accumulation in metastases from HCC.

Aged↗
PubMed2000

Effect of troglitazone on exocrine pancreas in rats with streptozotocin-induced diabetes mellitus.

Abnormality of pancreatic exocrine secretion has been observed in patients with diabetes mellitus. Troglitazone is a novel insulin-sensitizing agent that improves hyperglycemia and hyperinsulinemia in insulin-resistant diabetes mellitus. We investigated the effect of troglitazone on exocrine pancreas in streptozotocin (STZ)-induced diabetic rats. Diabetes mellitus was induced by intraperitoneal injection of STZ (25 mg/kg), and then 0.2% troglitazone containing rat chow was given for 2 weeks. Control diabetic animals received normal rat chow for 2 weeks. Glucose tolerance tests were performed before and after the administration of troglitazone. Pancreas weight, enzyme, protein, and insulin contents in the pancreas were measured. For the exocrine secretory study, pure pancreatic juice was collected hourly. Plasma glucose concentrations stimulated by the oral administration of 2.5 g/kg glucose in the troglitazone-treated group were significantly lower than those in the control group, but not plasma insulin concentrations. Pancreas weight in diabetic rats was less than that in normal rats. Administration of troglitazone resulted in a significant increase in pancreas weight and amylase and trypsin output. However, protein and insulin contents were not affected by the treatment with troglitazone. Both basal and cholecystokinin (CCK-8; 26 pmol/kg/h) stimulated exocrine secretion in juice volume, amylase, and trypsin output were markedly decreased in diabetic rats, compared with those in normal rats. Impaired basal and CCK-stimulated pancreatic exocrine secretion in diabetic rats recovered to the normal levels when troglitazone was given. In conclusion, troglitazone might be effective to restore exocrine pancreatic insufficiency in STZ-diabetic rats.

Animals↗
PubMed2000

BodyMap: a collection of 3' ESTs for analysis of human gene expression information.

BodyMap is a collection of site-directed 3' expressed sequence tags (ESTs) (gene signatures, GSs) that contains the transcript compositions of various human tissues and was the first systematic effort to acquire gene expression data. For the construction of BodyMap, cDNA libraries were made, preserving abundance information and histologic resolutions of tissue mRNAs. By sequencing 164,000 randomly selected clones, 88,587 GSs that represent chromosomally coded transcripts have been collected from 51 human organs and tissues. They were clustered into 18,722 independent 3' termini from transcripts, and more than 3000 of these were not found among ESTs assembled in UniGene (Build 75). Assessment of the prevalence of polyadenylation signals and comparison with GenBank cDNAs indicated that there was no significant contamination by internally primed cDNAs or genomic fragments but that there was a relatively high incidence (12%) of alternative polyadenylation sites. We evaluated the sensitivity and resolution of expression information in BodyMap by in silico Northern hybridization and selection of tissue-specific gene probes. BodyMap is a unique resource for estimation of the absolute abundance of transcripts and selection of gene probes for efficient hybridization-based gene expression profiling.

3' Untranslated Regions↗
PubMed2000

Absence of germline CHK2 mutations in familial gastric cancer.

Recently, the CHK2 gene was identified as being a candidate gene responsible for Li-Fraumeni syndrome (LFS). Gastric cancer is often clustered in families with LFS, so it is possible that germline CHK2 mutation is also present in familial gastric cancer (FGC). We therefore defined the genomic structure of the CHK2 gene, designed intronic primers, and searched for germline CHK2 mutations in 25 FGC cases by polymerase chain reaction-single strand conformational polymorphism analysis of the entire coding region. In all of the 25 cases, at least two siblings had histories of gastric cancer. There were no FGC cases that showed germline CHK2 mutations. Thus, it was indicated that germline CHK2 mutations do not contribute to the familial clustering of gastric cancer.

Aged↗
PubMed2000

Binding of Delta1, Jagged1, and Jagged2 to Notch2 rapidly induces cleavage, nuclear translocation, and hyperphosphorylation of Notch2.

Delta1, Jagged1, and Jagged2, commonly designated Delta/Serrate/LAG-2 (DSL) proteins, are known to be ligands for Notch1. However, it has been less understood whether they are ligands for Notch receptors other than Notch1. Meanwhile, ligand-induced cleavage and nuclear translocation of the Notch protein are considered to be fundamental for Notch signaling, yet direct observation of the behavior of the Notch molecule after ligand binding, including cleavage and nuclear translocation, has been lacking. In this report, we investigated these issues for Notch2. All of the three DSL proteins bound to endogenous Notch2 on the surface of BaF3 cells, although characteristics of Jagged2 for binding to Notch2 apparently differed from that of Delta1 and Jagged1. After binding, the three DSL proteins induced cleavage of the membrane-spanning subunit of Notch2 (Notch2(TM)), which occurred within 15 min. In a simultaneous time course, the cleaved fragment of Notch2(TM) was translocated into the nucleus. Interestingly, the cleaved Notch2 fragment was hyperphosphorylated also in a time-dependent manner. Finally, binding of DSL proteins to Notch2 also activated the transcription of reporter genes driven by the RBP-Jkappa-responsive promoter. Together, these data indicate that all of these DSL proteins function as ligands for Notch2. Moreover, the findings of rapid cleavage, nuclear translocation, and phosphorylation of Notch2 after ligand binding facilitate the understanding of the Notch signaling.

Animals↗
PubMed2000

Spontaneous and cytokine regulated c-fos gene expression in rheumatoid synovial cells: resistance to cytokine stimulation when the c-fos gene is overexpressed.

OBJECTIVE: To study the effect of cytokines on the transactivation of the c-fos gene in relation to the contribution of overexpression of c-fos/AP-1 in rheumatoid joint destruction. METHODS: The promoter region (-447 to +109) of the human c-fos gene was integrated upstream of the chloramphenicol acetyltransferase (CAT) reporter gene, and the effect of cytokines on the expression of the c-fos gene was studied in the rheumatoid synovial cells of early (3-4) or late (14-18) passages, in the presence or absence of cytokines, by the transient transfection assay. RESULTS: Expression of c-fos gene was enhanced by tumour necrosis factor alpha (TNF alpha) and interleukin 6 (IL6) in the synovial cells of early passage, whereas it was not enhanced in the synovial cells of late passage. The c-fos gene expression was also enhanced by 13-O-tetradecanoyl phorbol-13-acetate (TPA) in early passage but was somewhat suppressed in the late passage. It was found that the c-fos gene and c-Fos protein were both increased in the synovial cells of late passage. Similarly, c-fos gene expression was also not increased by TPA or cytokine stimulation in the stable c-fos transformants (fos-pH8) or H-ras transformed NIH3T3 cells (NIH H-ras cells) that constitutively expressed c-fos genes. CONCLUSIONS: Although TNF alpha and IL6 augmented c-fos gene expression of rheumatoid synovial cells, transactivation of c-fos gene became resistant against cytokine stimulation under prolonged expression of c-fos gene, which may impart a tumour-like characteristic to rheumatoid synovial cells.

Arthritis, Rheumatoid↗