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T Tsuji

Publications and source records attributed to T Tsuji.

At least 181 records · Page 10Linked to original sources
PubMed2000

Profiling of Caenorhabditis elegans proteins using two-dimensional gel electrophoresis and matrix assisted laser desorption/ionization-time of flight-mass spectrometry.

The nematode Caenorhabditis elegans (C. elegans) is the first animal whose whole 97 Mb genome sequence, encoding ca. 19000 open reading frames (ORF's), has been essentially determined. We tried to establish a 2-DE map of the nematode proteome by means of two-dimensional polyacrylamide gel electrophoresis (2-D PAGE). A soluble protein fraction of mixed stages of the worm, wild-type strain N2, was applied to 2-D PAGE. After Coomassie Brilliant Blue (CBB) staining, 1200 spots were detected and 140 major spots were excised from the gel and subjected to in-gel digestion with Achromobacter protease I (lysyl endopeptidase). Resulting peptides were analyzed by matrix assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS) followed by peptide mass fingerprinting for protein identification. With this approach we have obtained a two-dimensional electrophoresis (2-DE) protein map in which 69 spots were localized as landmarks for comparison of expression profiles to elucidate the basis of various biological events.

Animals↗
PubMed2000

IL-12-encoding plasmid has a beneficial effect on spontaneous autoimmune disease in MRL/MP-lpr/lpr mice.

Systemic lupus erythematosus (SLE) is characterized by immune abnormalities explained by the overproduction of Th(2)cytokines such as autoantibody production and polyclonal B cell activation. We examined the effect of administering a DNA plasmid encoding IL-12 on the lupus-like disease of MRL/MP-lpr/lpr (MRL/lpr) mice. Treatments were delivered intramuscularly every 4 weeks, starting at 4 weeks of age. This intervention significantly inhibited the accumulation of CD4(-)CD8(-)T cells, and reduced lymphadenopathy and splenomegaly. A significant decrease in serum IgG anti-DNA autoantibody titers was observed, and plasmid IL-12 therapy was also associated with a reduction in the proteinuria and glomerulonephritis characteristic of this disease. Serum IFN-gamma level was increased by inoculating IL-12 encoding plasmid, suggesting that the cytokine balance was skewed towards Th(1). The clinical implications of this suppression of autoimmune disease are also discussed.

Animals↗
PubMed2000

Expression of perlecan proteoglycan in the infarct zone of mouse myocardial infarction.

Perlecan, a basal lamina proteoglycan, has been shown to interact with other extracellular matrix (ECM) components, especially type IV collagen, and is thus involved in ECM formation. Perlecan has also been postulated to promote growth factor-receptor interactions, including the binding of basic fibroblast growth factor (bFGF) to its receptor, and to enhance mitogenesis and angiogenesis. To test our hypothesis that perlecan is increased in the myocardial infarct zone, we examined perlecan expression after experimentally induced myocardial infarction in BALb/c mice by the methods of in situ hybridization, Northern blotting, and immunohistochemistry. In situ hybridization revealed mRNA signals for perlecan in the infarct marginal zone on day 2 and in the infarct interior zone around infarct granulation tissue on day 7. On day 14 the signals were observed at the center point of the infarct. The signals were detected in spindle-shaped mesenchymal cells (fibroblasts and myofibroblasts). Some surviving myocytes in the infarct marginal zone also showed positive signals. The sequential changes in the perlecan mRNA signal distribution paralleled those for type IV collagen mRNA. Northern blotting demonstrated increased expression of perlecan consistent with the observations of in situ hybridization. Immunopositive staining for perlecan was observed in the infarct zone around granulation tissue on day 7 and in the entire infarct zone on days 14-28. Immunostaining for bFGF was localized surrounding the infarct granulation tissue on day 7 and overlapped with perlecan immunostaining. The present results demonstrated the expression of perlecan by spindle-shaped mesenchymal cells (fibroblasts and myofibroblasts) and some surviving myocytes in the myocardial infarct, indicating the contribution of perlecan to the pathological course of myocardial infarction.

Animals↗
PubMed2000

Increased adrenomedullin in cerebrospinal fluid after traumatic brain injury in children: a preliminary report.

Adrenomedullin is a recently discovered 52-amino-acid peptide that is a potent vasodilator. Infusion of adrenomedullin increases regional cerebral blood flow and reduces infarct volume after vascular occlusion in rats. Adrenomedullin may represent an endogenous neuroprotectant since it is increased after focal brain ischemia. Cerebral hypoperfusion is present after traumatic brain injury (TBI) in children. We hypothesized that adrenomedullin levels would be increased in children with severe TBI. Total adrenomedullin concentrations were measured using a radioimmunometric assay. Thirty-six samples of ventricular cerebrospinal fluid (CSF) from 10 pediatric patients were collected during the first 10 days after severe TBI (GCS < 8). Control CSF was obtained from 5 children undergoing lumbar puncture, who had normal CSF parameters and no evidence of central nervous system infection. Patients underwent standard neuro-intensive care, including cerebrospinal fluid drainage. Data were analyzed using a univariate regression model. Adrenomedullin concentration was markedly elevated in CSF of children following TBI versus control (mean level 10.65 vs 1.51 fmol/ml, p = 0.006). All 36 case samples had an adrenomedullin concentration above the median value for the controls (1.52 fmol/ml). We conclude adrenomedullin is elevated in the CSF of children following severe TBI. We speculate that it participates in the endogenous response to cerebral hypoperfusion after TBI.

Adrenomedullin↗
PubMed2000

Establishment of a human hepatoma cell line, HLE/2E1, suitable for detection of p450 2E1-related cytotoxicity.

By transfection of an expression vector of human cytochrome P450 2E1 (CYP2E1) into a human hepatoma cell line (HLE), a new cell line (HLE/2E1) that stably expresses activity of CYP2E1 has been established. The HLE/2E1 cell line expressed a higher level of CYP2E1 messenger ribonucleic acid than did the mother HLE cell line. CYP2E1 enzyme activity determined by a p-nitrophenol oxidation assay was also higher in HLE/2E1 cells than in HLE cells. In addition, the enzyme activity of the HLE/2E1 cells was increased by ethanol treatment. Exposure to acetaminophen (APAP) or buthionine sulfoximine (BSO) caused a greater decrease in viability of the HLE/2E1 cells than that of the HLE cells, as determined by the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide assay. The cytotoxicity of APAP or BSO to HLE/2EI cells was inhibited by the addition of ethanol or vitamin E. However, the cytotoxicity of both APAP and BSO was enhanced by 24-h preincubation of HLE/2E1 cells with ethanol. These results show that this cell line provides a useful model for studying catalytic properties of CYP2E1 and cytotoxic mechanisms of chemicals metabolized by CYP2E1.

Acetaminophen↗
PubMed2000

Active intestinal tuberculosis with esophageal candidiasis due to idiopathic CD4(+) T-lymphocytopenia in an elderly woman.

We describe a case of intestinal tuberculosis and esophageal candidiasis in an 85-year-old Japanese woman with idiopathic CD4+ T-lymphocytopenia (ICL). The patient exhibited clinical symptoms of odynophagia, bloody diarrhea, and high fever. Physical examination on admission showed a poor nutritional status. Endoscopic examination of the upper digestive tract revealed the esophageal mucosa to be covered with yellowish-white plaque-like lesions. Colonoscopic examination revealed multiple annular ulcerations with bleeding. She was diagnosed with intestinal tuberculosis by polymerase chain reaction (PCR) and fecal culture. Her CD4+ T-lymphocyte count was 178/mm3 and no evidence of human immunodeficiency virus (HIV) infection was found. She was successfully treated with fluconazole and antituberculosis drugs. This case emphasizes the importance of opportunistic infections in elderly patients with predisposing conditions such as ICL.

Aged↗
PubMed2000

Alterations of extracellular matrix induced by tobacco smoke extract.

Epidemiologic studies have indicated the association between tobacco smoking and skin aging, but the exact mechanism of tobacco smoke-induced premature skin aging is currently unknown. In this study, we investigated the alterations of collagen, matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) in human fibroblasts treated with tobacco smoke extract. Human fibroblasts were exposed to different concentrations of water-soluble extract from tobacco smoke. Human fibroblasts irradiated with ultraviolet A1 (UVA1) were used as positive controls because the mechanism of UVA1-mediated MMP expression has been well characterized. The expression of MMP and TIMP was analyzed semiquantitatively following reverse transcriptase-polymerase chain reaction. Production of type I and type III collagens was detected by Western blotting and biosynthesis of new collagen was assessed by 3H-proline incorporation. Upon treatment with tobacco smoke extract or UVA1 irradiation, the expression of MMP-1 and MMP-3 mRNA was significantly increased in a dose-dependent manner. Maximum induction was observed with 25 microl/ml tobacco smoke extract. In contrast, the expression of TIMP-1 and TIMP-3 mRNA remained unchanged. Western blotting of the supernatant revealed that type I and type III collagens were decreased as compared with untreated controls. Collagen biosynthesis was significantly reduced by 40.1% following treatment with 25 microl/ml tobacco smoke extract. Sodium azide, L-ascorbic acid and Trolox (a water-soluble vitamin E) prevented both the UVA1- and the tobacco-induced alteration of MMP-1. These observations suggest that the imbalance of connective tissue matrix components might contribute to the molecular basis for premature skin aging in smokers. They also suggest that reactive oxygen species including singlet oxygen mediate this process.

Adult↗
PubMed2000

The relationship between circulating sialyl Tn antigen and polypoid or nonpolypoid growth characteristics in colorectal cancer.

Recent studies delineated two different patterns of tumor growth in colorectal carcinoma characterized as polypoid and nonpolypoid (PG-type and NPG-type, respectively). We quantified serum sialyl Lewis (Le)a (CA19-9), sialyl Lex (SLX), sialyl Tn (STN), and carcinoembryonic antigen (CEA) in 269 colorectal cancer patients to establish whether their levels correlated with any biological or clinical differences between PG-type and NPG-type cancer. Patients were divided into high and low antigen groups (higher or lower than a selected diagnostic-based cut-off value) and compared. Statistical testing was by univariate and multivariate (logistic regression) analyses. Forty-seven (17.5%) patients with PG-type and 222 (82.5%) with NPG-type cancer were studied. In contrast to NPG-type, the characteristics of the PG-type cancers included a low rate of lymph node metastasis and a high serum STN level. In contrast to a low STN level, a high STN level was independently related to the presence of distant metastasis in patients with PG-type cancer, and also to the presence of distant metastasis and large-sized tumor in patients with NPG-type cancer. These data suggest that differences in STN levels in the serum of patients with PG-type or NPG-type colorectal carcinomas may be at least partly responsible for different tumor progression behavior.

Antigens, Tumor-Associated, Carbohydrate↗
PubMed2000

Carcinoma of the splenic flexure: multivariate analysis of predictive factors for clinicopathological characteristics and outcome after surgery.

The clinicopathological characteristics and outcome of splenic flexure cancer after surgery have yet to be fully elucidated. The aim of the current study was, therefore, to establish predictive factors related to splenic flexure cancer and outcome after surgery. We compared the clinicopathological characteristics and outcome of 34 patients with splenic flexure cancers (which represents 3.7% of the total number of colon cancers in our series) with those of 418 patients with right colon and 475 patients with left colon cancers by univariate and multivariate analyses, using logistic regression analysis and Cox's proportional hazards model. Splenic flexure cancers had a high risk of obstruction (26.5% of patients), and had a more advanced stage and lower cure rate than left colon cancers. Logistic regression analysis revealed that two independent factors, colonic obstruction and the presence of distant metastases, were related to the splenic flexure tumor site. Splenic flexure cancer patients had a poorer outcome than those with left colon cancer (P = 0.0361). However, there was no difference in survival between patients with splenic flexure, those with right colon cancer and those with left colon cancer who underwent curative surgery. Cox's regression analysis revealed that neither the site of splenic flexure nor colonic obstruction was an independent prognostic factor. In conclusion, splenic flexure cancer is characterized by a high risk of obstruction and the presence of distant metastases. However, after curative resection, splenic flexure cancer has a similar outcome to colon cancer at other sites. In addition, neither the splenic flexure site nor colonic obstruction had an independent influence on patient survival after surgery.

Colonic Neoplasms↗
PubMed2000

Differences in release mechanisms and distributions for sialyl Le(a) and sialyl Le(x) antigens in colorectal cancer.

BACKGROUND: To investigate colorectal cancer-related carbohydrate antigen release and distribution, we evaluated serum levels of sialyl Le(a) (CA19-9) and sialyl Le(x) antigen (SLX) in blood samples obtained from both a peripheral vein and a tumor's draining vein. METHODS: Blood samples were obtained during surgery from 126 patients. Based on these samples, patients were placed into a high-antigen group, with a concentration above a selected cutoff value, or into a low-antigen group, with a tumor marker concentration below that same value. The blood samples obtained from peripheral veins were designated by the "p" prefix, and samples from drainage veins were designated by the "d." RESULTS: Serum d-SLX levels were significantly higher than p-SLX levels (P < .0001), although there was no difference between those of d-CA19-9 and p-CA19-9. Only 1 (3.6%) of 28 patients in the high d-CA19-9 group had a low p-CA19-9. In contrast, 6 (33.3%) of 18 patients in the high d-SLX group had low p-SLX levels (P = .0103). Correlations between pathological variables and either p-CA19-9 levels or d-CA19-9 levels were similar. However, both distant metastasis and venous invasion did prove to be independent variables related to d-SLX levels, as shown by logistic regression analysis. CONCLUSIONS: SLX may drain predominantly via the draining veins of colorectal tumors into portal circulation, whereas CA19-9 may drain via another route.

Aged↗
PubMed2000

Expression of MAGE, GAGE and BAGE genes in human liver diseases: utility as molecular markers for hepatocellular carcinoma.

BACKGROUND/AIMS: The MAGE, GAGE and BAGE genes encode tumor antigens recognized by autologous cytotoxic T lymphocytes. The aim of this study was to evaluate the possibility of using these genes as molecular markers and as the targets of specific immunotherapy for human hepatocellular carcinoma (HCC). METHODS: The expressions of MAGE-1, MAGE-3, GAGE1-6, GAGE1-2 and BAGE mRNA in 33 surgically resected HCC samples and 26 of their corresponding non-cancerous samples (11 liver cirrhosis and 15 chronic hepatitis) were studied by a reverse-transcription polymerase chain reaction, and were compared with clinicopathological parameters. The expression of MAGE-1 was also examined in 16 biopsied HCC samples. RESULTS: MAGE-1, MAGE-3, GAGE1-6, GAGE1-2 and BAGE mRNA were expressed in 67%, 39%, 36%, 30%, and 21% of the HCC, respectively. At least one transcript was detected in 88% of the HCC, while no expression was observed in the non-cancerous livers. There was no significant correlation between the expression of any of the tumor antigens examined and the differentiation stage or size of the HCC. Especially, MAGE-1 was highly expressed in small HCC with a diameter of less than 2 cm and in well-differentiated HCC (81% and 70%, respectively), and was also expressed even in alpha-fetoprotein-negative and PIVKA-II-negative HCC (58% and 76%, respectively). The MAGE-1 expression was detected in 69% of biopsied HCC samples and the expression was high in both small and well-differentiated HCC. CONCLUSIONS: These tumor-specific antigens can be useful as molecular markers and as the possible target molecules for the specific immunotherapy of human HCC.

Adult↗
PubMed2000

Induction of apoptosis in lymphoid tissues of mice after intramuscular injection of enterotoxigenic Escherichia coli enterotoxin.

Although it has been reported that intravenous injection of Escherichia coli enterotoxin induces atrophies of the thymus and spleen by necrosis, the toxin injected intramuscularly to mice induced atrophies of both tissues, which were associated with apoptosis of lymphocytes. Apoptosis predominantly occurred in the thymus and increased in a time-dependent manner up to 26 h and faint ladder band patterns of DNA were observed at 36 h. Although the high dose of toxin also induced in vitro apoptosis in cultured thymocytes, the toxin was not detected in the serum at levels sufficient to cause in vitro apoptosis after intramuscular administration. By flow cytometric analysis, CD4+ CD8+ double-positive T cell and CD45+ positive B cell numbers were found to be mainly decreased in thymus and spleen, respectively, of mice. These results suggest that the atrophies of thymus and spleen by intramuscular administration of the toxin to mice are due to apoptosis of CD4+ CD8+ double-positive T and CD45+ positive B cells, respectively, but the toxin does not reach these cells via the circulation. A different mechanism from that in vitro in cultured cells might be involved in the induction of apoptosis in vivo.

Adjuvants, Immunologic↗
PubMed2000

doc-1--mediated apoptosis in malignant hamster oral keratinocytes.

PURPOSE: Cell cycle mediators involved in inducing apoptosis are frequently deregulated during carcinogenesis. Deleted in oral cancer-1 (doc-1) is an S-phase regulator that is inactivated during oral carcinogenesis. Transfection of doc-1 into malignant oral keratinocytes leads to increased cell loss. It is hypothesized that ectopic expression of doc-1 in hamster oral cancer cells induces apoptosis. MATERIALS AND METHODS: Malignant hamster oral keratinocytes (wt-HCPC-1), which lack measurable doc-1 mRNA and protein, were previously transfected with either a CMV-doc-1 expression vector construct (doc-HCPC-1) or the parental control vector pcDNA3 (cv-HCPC-1). A trypan blue exclusion assay was performed to examine cell death in the parental or wild-type HCPC-1 keratinocytes, HCPC-1 transfected with the parental pcDNA3 vector, and the doc-1 transfected HCPC-1 cells. To examine whether ectopic expression of doc-1 mediates gross cellular changes consistent with apoptosis, toluidine blue-safranin differential staining and the quantitative fluorescent microscopy assays were performed. To identify early apoptotic cytochemical changes observed in the cell membrane and nucleus, annexin V/propidium iodide (PI) fluorescence-activated cell sorter (FACS) analysis and the terminal deoxytransferase-mediated dUTP nick-end labeling (TUNEL) assay were performed. RESULTS: Doc-HCPC-1 showed elevated numbers of dead cells over wt-HCPC-1 and cv-HCPC-1 in the trypan blue exclusion assay. Toluidine blue-safranin staining and quantitative fluorescent microscopy showed significant morphologic changes in the doc-1 transfectants consistent with apoptosis (P < .05). TUNEL assays (P < .05) and annexin V/PI FACS analysis (P < .05) also showed early cytochemical changes in the doc-HCPC-1 transfectants, confirming that ectopic expression of doc-1 induces apoptosis. CONCLUSIONS: These data suggest that doc-1 induces apoptosis in malignant hamster oral keratinocytes. It is hypothesized that doc-1 is a mediator of apoptosis that is inactivated during hamster oral carcinogenesis.

Animals↗
PubMed2000

Extensive generation of human cord blood CD34(+) stem cells from Lin(-)CD34(-) cells in a long-term in vitro system.

Human CD34(-) hematopoietic stem cells (HSCs) have been identified as potential precursors of CD34(+) HSCs by using xenogeneic transplantation systems. However, the properties of CD34(+) cells generated from CD34(-) cells have not been precisely analyzed due to the lack of an in vitro system in which CD34(+) cells are continuously produced from CD34(-) cells. We conducted this study to determine whether CD34(+) cells generated in vitro from CD34(-) cells have long-term multilineage reconstitution abilities. Lin(-)CD34(-) population isolated from human cord blood was cultured in the presence of murine bone marrow stroma cell line, HESS-5, and human cytokines, thrombopoietin, Flk2/Flt3 ligand, stem cell factor, granulocyte colony-stimulating factor, interleukin 3 (IL-3), and IL-6. They were analyzed weekly for their surface markers expressions, colony-forming cells, long-term culture initiating cells (LTC-IC), and SCID repopulating cells (SRC) abilities up to 30 days of culture. In this culture system, more than 10(7) CD34(+) cells can be continuously generated from 10(4) CD34(-) cells over 30 days. These CD34(+) cells produce colony-forming units, LTC-IC, and SRC with multi-lineage differentiation, all of which are characteristic features of hematopoietic stem/progenitor cells. These findings suggest that CD34(-) HSCs have extensive potential for the generation of CD34(+) HSCs in vitro. This system provides a novel and potentially useful procedure to generate CD34(+) cells for clinical transplantation and gene therapy.

Animals↗
PubMed2000

Time-dependent changes of decorin in the infarct zone after experimentally induced myocardial infarction in rats: comparison with biglycan.

Decorin, a small dermatan sulphate proteoglycan, has been postulated to interact with other components of the extracellular matrix. We examined time-dependent changes of decorin in the infarct zone after experimentally induced myocardial infarction in rats by Northern blotting, in situ hybridization, and immunohistochemistry. The expression of decorin mRNA was compared to that of biglycan mRNA. Northern blotting demonstrated that the decorin mRNA expression was not increased in the infarct zone on day 2, while increased biglycan mRNA was observed at that time (average 3.1-fold increase). Decorin mRNA expression was increased on day 7, and reached a peak (average 2.2-fold increase) around day 14. Biglycan mRNA expression also reached a peak level around day 14 (average 13.3-fold increase). In situ hybridization revealed that mRNA signals for decorin did not appear in the infarct zone on day 2, while biglycan mRNA signals were observed. Decorin mRNA signals were observed in spindle-shaped mesenchymal cells in the infarct peripheral zone on day 7. The decorin mRNA signals appeared later than those of biglycan. Immunopositive staining for decorin was observed in the infarct zone on day 7. The present results demonstrated a time-dependent increase in decorin mRNA expression in mesenchymal cells in the infarct zone in rats. Decorin mRNA appeared later and was increased to a lower extent in the infarct zone than biglycan mRNA.

Animals↗
PubMed2000

Serial magnetic resonance imaging in a non-traumatic rabbit osteonecrosis model: an experimental longitudinal study.

We investigated the time-dependent natural course of experimental osteonecrosis (ON), including initial changes in ON and the reparative process, using in vivo serial repetitive magnetic resonance imaging (MRI) in a non-traumatic rabbit serum sickness ON model. Some necrotic lesions were detected at 1 week (3 of 16 femora with necrotic lesions) and some in the metaphysis were detected by 12 weeks (2 of 6 femora with lesions) on T(1)-weighted, T(2)-weighted, and fat suppression T(1)-weighted images. On contrast-enhanced MRI, extravasation of the erythrocytes was detected at 72 h (7 of 26 femora with lesions) as a small, focal enhanced area. Necrotic lesions were detected in all abnormal femora by 6 weeks (16 of 16 femora with lesions) as focal, homogeneously or inhomogeneously enhanced areas. Reparative tissue replaced with new vascular and trabecular formation in necrotic areas was detected as an extended marginal enhanced area at 12 weeks. These results suggest that the enhancement patterns on contrast-enhanced MRI may provide helpful information about the developmental and reparative process of clinical ON.

Animals↗
PubMed2000

A preliminary report of the treatment of blue nevus with dermal injection of riboflavin and exposure to near-ultraviolet/visible radiation (ribophototherapy).

Dye lasers are useful for treating pigmented skin lesions, but their equipment is expensive and bulky. A simple and cheap phototherapy would be acceptable to dermatologists for treating pigmented skin lesions such as nevus of Ota. We investigated as a pilot study whether dermal injection of riboflavin and exposure to near-ultraviolet/visible radiation (ribophototherapy) decreases the dermal pigment of blue nevi which are recalcitrant to laser therapy. The therapeutic efficacy was assessed by comparison of the amount of dermal pigment in hematoxylin-eosin specimens taken before and after treatment. Pigmentation of the nevus became faint to the depth of 1 mm with little noticeable epidermal change after 21 treatments. At the deeper dermis somewhere between 3 and 4 mm from the epidermis, ballooning degeneration of the dermal cells was observed in hematoxylin-eosin specimens. Ribophototherapy is hopeful for treating pigmented skin lesions.

Adult↗