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Rishu Takimoto

Publications and source records attributed to Rishu Takimoto.

At least 19 recordsLinked to original sources
PubMed2006

Ex vivo large-scale generation of human platelets from cord blood CD34+ cells.

In the present investigation, we generated platelets (PLTs) from cord blood (CB) CD34(+) cells using a three-phase culture system. We first cultured 500 CB CD34(+) cells on telomerase gene-transduced human stromal cells (hTERT stroma) in serum-free medium supplemented with stem cell factor (SCF), Flt-3/Flk-2 ligand (FL), and thrombopoietin (TPO) for 14 days. We then transferred the cells to hTERT stroma and cultured for another 14 days with fresh medium containing interleukin-11 (IL-11) in addition to the original cytokine cocktail. Subsequently, we cultured the cells in a liquid culture medium containing SCF, FL, TPO, and IL-11 for another 5 days to recover PLT fractions from the supernatant, which were then gel-filtered to purify the PLTs. The calculated yield of PLTs from 1.0 unit of CB (5 x 10(6) CD34(+) cells) was 1.26 x 10(11) - 1.68 x 10(11) PLTs. These numbers of PLTs are equivalent to 2.5-3.4 units of random donor-derived PLTs or 2/5-6/10 of single-apheresis PLTs. The CB-derived PLTs exhibited features quite similar to those from peripheral blood in morphology, as revealed by electron micrographs, and in function, as revealed by fibrinogen/ADP aggregation, with the appearance of P-selectin and activated glycoprotein IIb-IIIa antigens. Thus, this culture system may be applicable for large-scale generation of PLTs for future clinical use.

Antigens, CD34↗
PubMed2006

Human herpesvirus-6 hepatitis associated with cyclosporine-A encephalitis after bone marrow transplantation for chronic myeloid leukemia.

A 31-year-old man referred to our hospital for treatment of his chronic myeloid leukemia (CML) in the first chronic phase by bone marrow transplantation. We pretreated him with cyclophosphamide and total body irradiation and bone marrow transplantation (BMT) was carried out. On day 31, the engraftment was confirmed and on day 52, acute graft versus host disease (GVHD) was observed. On day 189, he lost consciousness due to cyclosporine A-induced leukoencephalopathy and 375 mg cyclosporine A was changed to 100 mg prednisolone. On day 199, liver dysfunction (AST 410 IU/L, ALT 557 IU/L, gammaGTP 385 IU/L, ALP 363 IU/L, D-Bil 0.3 mg/dl) developed and a liver biopsy was performed. PCR analysis of DNA from the liver biopsy specimen was positive for HHV-6 and immunostaining using anti-HHV-6 and anti-HHV-6b antibodies showed positive staining in the cytosol of hepatocytes. No other viruses were found to induce hepatitis. From these results, he was diagnosed as having HHV-6 hepatitis and it was successfully treated with gancyclovir (GCV) administration.

Adult↗
PubMed2006

A phase I/II study of nedaplatin and 5-fluorouracil with concurrent radiotherapy in patients with esophageal cancer.

PURPOSE: To determine the recommended dose (RD) of cis-diammine-glycolatoplatinum (nedaplatin) when given concurrently with 5-FU and high dose radiation therapy in the treatment of esophageal cancer. The purpose of the phase II trial is to determine efficacy and further define the side effect profile. METHODS: Twenty-six patients with clinical stage I to IVA squamous cell carcinoma of the esophagus were enrolled in a non-surgical treatment comprised of a fixed dose of fluorouracil (400 mg/m2 administered as continuous intravenous infusion on days 1-5 and days 8-12) plus escalating doses of nedaplatin (40 mg/m2 in level 1, 50 mg/m2 in level 2, or 60 mg/m2 in level 3 on days 1 and 8), repeated twice every 3 weeks with concurrent radiotherapy (60 Gy). RESULTS: Between July 1998 and February 2004, a total of 26 patients entered this trial, all of whom were considered evaluable for toxicity assessment. In phase I of the study, 12 patients were treated in sequential cohorts of three to six patients per dose level. The maximum tolerated dose was reached at level 3 with two grade 4 neutropenia and one grade 4 thrombocytopenia. Thus, the recommended dosing schedule is level 2. Of the 20 patients treated at the RD level 2, including 6 patients of the RD phase I portion, 8 out of 20 patients (40%) had grade 3-4 neutropenia, 5 patients (25.0%) had grade 3-4 thrombocytopenia, 4 patients (20.0%) had grade 3 anemia and 4 patients (20.0%) had grade 3-4 esophagitis. Other toxicities were relatively mild and usually of grade 2 or less. Objective responses were noted in the 26 patients (overall response rate, 88.5%) including 11 (42.3%) complete remissions. The 1- and 3-year survival rates were 65.1 and 37.2%, respectively, with a median survival time of 21.2 months. CONCLUSIONS: The combination of nedaplatin and 5-FU with radiation is a feasible regimen that shows promising antitumor activity with an acceptable safety profile in patients with esophageal cancer.

Aged↗
PubMed2006

Ex vivo expansion of G-CSF-mobilized peripheral blood CD133+ progenitor cells on coculture with human stromal cells.

OBJECTIVE: The pentaspan molecule CD133 has been shown to be a marker of more primitive hematopoietic progenitors in mobilized peripheral blood (PB). Our objective was to assess the efficacy of PB CD133(+) cells in our coculture system using human telomerized stromal (HTS) cells. METHODS: Five thousand PB CD133(+) cells or conventional cord blood (CB) CD34(+) cells were expanded with or without HTS cells in the presence or absence of stem cell factor, thrombopoietin, and Flk-2/Flt-3 ligand. RESULTS: The coculture was significantly superior in expanding PB clonogenic cells as compared with the stroma-free culture (CFU-C, 2 +/- 0 vs 111 +/- 15-fold of initial cell number, p < 0.01), and the fold increase of PB clonogenic cells was comparable to that for CB cells after two weeks of coculture (BFU-E, 54 +/- 3 vs 56 +/- 4-fold; CFU-GM, 156 +/- 26 vs 83 +/- 9-fold; CFU-Mix, 30 +/- 11 vs 80 +/- 36-fold). However, proliferation of CFU-Mk from PB on coculture with HTS cells was modest as compared with stroma-free culture. Concomitantly, multiple hematopoietic cells transmigrated below the stromal layer and formed cobblestone areas (CAs). The production of hematopoietic progenitor cells from CAs after coculture with PB was significantly lower than that seen in cells cocultured with CB for four weeks (CFU-Mix, 0 +/- 0 vs 9 +/- 5-fold on day 28, p < 0.01), although a similar number of CAs derived from PB and CB were observed. CONCLUSION: PB CD133(+) cells proliferated efficiently above the stromal layer, while the characteristics of PB CD133(+) cells underneath the human stromal layer were likely to be maintained, even after long-term hematopoietic-stromal interaction.

AC133 Antigen↗
PubMed2006

[De novo CD5-positive diffuse large B-cell lymphoma associated with autoimmune hemolytic anemia presenting as erythroid hypoplasia].

A 54-year-old woman was admitted due to high-grade fever, cervical lymphadenopathy and general malaise in May 2003. On examination, severe anemia was noted, direct Coombs and cold hemagglutinin tests were positive and the haptoglobin level was low in the peripheral blood. However, a bone marrow examination revealed marked erythroid hypoplasia. A diagnosis was made of co-existing combined type autoimmune hemolytic anemia (AIHA) and erythroid hypoplasia. A pathologic diagnosis of de novo CD5-positive diffuse large B-cell lymphoma (de novo CD5+ DLBCL) was made based on a cervical lymph node biopsy. The patient was treated with CHOP accompanied by rituximab (R-CHOP), resulting in complete remission after 3 courses of chemotherapy. The AIHA and erythroid hypoplasia subsided after 2 courses of R-CHOP. The sera obtained during erythroid hypoplasia significantly inhibited the growth of erythroid progenitor cells (erythroid colony-forming units, CFU-E) from her bone marrow collected after recovery. We report here a patient with de novo CD5+ DLBCL associated with both AIHA and erythroid hypoplasia, suggesting that the lymphoma triggered an abnormal immunity which generated some humoral inhibitors against erythropoiesis.

Anemia, Hemolytic, Autoimmune↗
PubMed2006

[Enteropathy-type T-cell lymphoma with CD8 and CD56 expression].

A 54-year-old male presented at a previous hospital with abdominal pain, where the patient was diagnosed as having abdominal and retroperitoneal lymphadenopathies by CT scan, then, he was referred to our hospital for further examination. Upper gastrointestinal endoscopy showed a tumor formation in the second portion of the duodenum, and there were medium sized atypical lymphocytes in biopsy specimens of the tumor. Since the atypical lymphocytes in the biopsy specimens were positive for CD3, CD8, CD56 and CD103 by immunostain, and had a rearrangement of the T-cell receptor 7-chain, the patient was diagnosed as having enteropathy-type T-cell lymphoma (ETL). Although ETL usually occurs as a complication of celiac disease, malabsorption and anti-gliadin antibodies, which are normally present in celiac disease, were not observed in this patient.

CD3 Complex↗
PubMed2006

[Peripheral T-cell lymphoma presenting with eosinophilia due to interleukin-5 produced by lymphoma cells].

A 72-year-old male was admitted to our hospital to determine the cause of neck lymphadenopathies. He was diagnosed pathologically by a lymph node biopsy as having peripheral T-cell lymphoma (PTCL). He also had developed eosinophilia and his serum concentration of Interleukin-5 (IL-5) was extremely high. Immunostaining of a resected lymph node revealed that there were lymphoma cells which were stained with anti-IL-5 antibody. After 2 courses of CHOP therapy, the lymphadenopathies and eosinophilia disappeared and the concentration of serum IL-5 was normalized. It was suggested that IL-5 produced by lymphoma cells had induced his eosinophilia.

Aged↗
PubMed2005

Augmentation of antitumor effects of p53 gene therapy by combination with HDAC inhibitor.

We have previously shown that the HDAC inhibitors (HDACI) activate the p53 molecule through acetylation of 320 and 373 lysine residues, upregulate PIG3 and NOXA and induce apoptosis in cancer cells expressing wild and pseudo-wild type p53 genes (Terui T, et al. Cancer Res 2003; 63:8948-54). It has also been reported that expression of the Coxsackie adenovirus receptor and subsequent transfection efficiency of the adenovirus in cancer cells were enhanced by HDACI treatment. In this study, we extended these observations to explore the combination effect of adenoviral vector carrying wild type p53 (Ad-p53) gene therapy with a HDACI, sodium butyrate (SB), on xenografted human gastric cancer cells (KATO-III) and hepatocellular carcinoma cells (HuH7) in nude mice. We first confirmed an increased expression of Coxsackie adenovirus receptors with an associated increment of transgene (X-gal) expression by SB treatment in KATO-III cells. We then injected Ad-p53 into subcutaneous tumors of KATO-III and HuH7 combined with intraperitoneal administration of SB and found a significantly higher growth suppressive effect than single treatments of each. Even a complete regression of tumors was observed in three of five mice treated with this combination while with single treatment no tumor regression was observed. Tumors treated with the combination showed higher numbers of TUNEL positive cells than those treated with a single modality. Moreover, necrotic changes were more evident in tumors treated with the combination than separately, a compatible finding to the observation that vascularity revealed by CD34 staining was poorer in tumors treated with the combination than those treated with p53 gene or SB alone. This was further supported by the finding that BAI-1 (brain specific angiogenesis inhibitor-1), an inhibitor of vascularization, was induced by SB treatment in KATO-III and HuH7 cells transfected with Ad-p53. Thus SB was shown to be an efficient potentiator of p53 gene therapy for cancer.

Adenoviridae↗
PubMed2005

Human mesenchymal stem cells xenografted directly to rat liver are differentiated into human hepatocytes without fusion.

Hepatic transdifferentiation of bone marrow cells has been previously demonstrated by intravenous administration of donor cells, which may recirculate to the liver after undergoing proliferation and differentiation in the recipient's bone marrow. In the present study, to elucidate which cellular components of human bone marrow more potently differentiate into hepatocytes, we fractionated human bone marrow cells into mesenchymal stem cells (MSCs), CD34+ cells, and non-MSCs/CD34- cells and examined them by directly xenografting to allylalcohol (AA)-treated rat liver. Hepatocyte-like cells, as revealed by positive immunostaining for human-specific alpha-fetoprotein (AFP), albumin (Alb), cytokeratin 19 (CK19), cytokeratin 18 (CK18), and asialoglycoprotein receptor (AGPR), and by reverse transcription-polymerase chain reaction (RT-PCR) for expression of AFP and Alb mRNA, were observed only in recipient livers with MSC fractions. Cell fusion was not likely involved since both human and rat chromosomes were independently identified by fluorescence in situ hybridization (FISH). The differentiation appeared to follow the process of hepatic ontogeny, reprogramming of gene expression in the genome of MSCs, as evidenced by expression of the AFP gene at an early stage and the albumin gene at a later stage. In conclusion, we have demonstrated that MSCs are the most potent component in hepatic differentiation, as revealed by directly xenografting into rat livers.

Albumins↗
PubMed2005

Telomerized human bone marrow-derived cell clones maintain the phenotype of hematopoietic-supporting osteoblastic and myofibroblastic stromal cells after long-term culture.

OBJECTIVE: Gene transfer of the telomerase catalytic subunit (TERT) into primary human stromal cells prolonged their lifespan. However, primary human stromal cells are actually composed of adipocytes, myofibroblasts, osteoblasts, etc. Our objective was to investigate the phenotype and hematopoietic-support of the human telomerized stromal cell (HTS) in clonal level. MATERIALS AND METHODS: We established HTS clones (HTS-1 to HTS-9) from a parental population of HTSs by limiting dilution. Hematopoietic-supporting activity of the HTS clones was examined by coculturing with CD34(+) cells. RESULTS: HTS-1 to HTS-3 contained alkaline phosphatase (ALP)(+) cells, and HTS-4 to HTS-9 were composed of both ALP(+) and alpha-smooth muscle actin-positive cells. Although all HTS clones exhibited normal growth kinetics, one of the HTS clones exhibited a chromosomal abnormality. Moreover, the parental population of the HTS cells acquired an increased growth rate and anchorage independence after 4 years of culturing. Expression of hematopoietic growth factors, such as stem cell factor, angiopoietin-1, and hedgehog mRNA was detected in all HTS clones. The degree of hematopoietic progenitor support differed between the HTS clones, and the expansion level of CD34(+) cells was the highest in HTS-8. CONCLUSION: Human telomerized stromal cell clones exhibited the phenotype of hematopoietic supporting osteoblastic and myofibroblastic cells after long-term culture. Clinical application of HTS cells should be limited because of their potential for neoplastic transformation after hTERT gene transfer. HTS cells may be useful for analyzing the molecular mechanism of hematopoietic support of human stromal cells.

Bone Marrow Cells↗
PubMed2005

A patient with TP53 germline mutation developed Bowen's disease and myelodysplastic syndrome with myelofibrosis after chemotherapy against ovarian cancer.

Here we report a case of myelodysplastic syndrome (MDS) with myelofibrosis associated with Bowen's disease. A female patient had undergone an operation and chemotherapy for ovarian cancer when she was 65 years old, and she developed MDS at the age of 70 years old. PCR-single strand conformation polymorphism (SSCP) analysis of peripheral blood mononuclear cells, a Bowen's disease lesion, and normal skin showed an abnormal peak in TP53 exon5. Direct sequencing revealed that they all had missense mutation in codon 175 (G to A) of arginine switched to histidine, suggesting a germline mutation of TP53. It was speculated that p53 function was lost by TP53 germline mutation with the loss of a wild type allele induced by the chemotherapy against ovarian cancer, leading to the development of MDS. No therapeutic effects of low dose melphalan or cyclosporine A on MDS were observed, however one month of 30 mg/day prednisolone administration induced a hematological response.

Aged↗
PubMed2005

[Essential thrombocythemia associated with incomplete type intestinal Behçet disease during hydroxyurea treatment].

A 77-year-old man was diagnosed as having essential thrombocythemia in 1992. Treatment with hydroxyurea was started in 1997, which stabilized the platelet count. The patient then suffered from pharyngalgia and rhinitis with a high fever, immediately after which he developed tarry stools and anemia and was admitted to our hospital. The physical examination revealed splenomegaly, oral aphthous ulcers, genital ulcers and skin lesions on the lower limbs. His hematological and biochemical tests revealed anemia and increased level of C-reactive protein. He also had an HLA-B51 phenotype. The findings of gastro-intestinal and colon fiberoscopy showed a duodenal ulcer and multiple ulcers on ascending colon. He was thus diagnosed as having intestinal tract-type Behçet disease. After withdrawal of the hydroxyurea administration, the intestinal ulcers, oral aphthous ulcers and genital ulcers improved.

Aged↗
PubMed2004

Plasma glutathione S-Transferase P1-1 as a prognostic factor in patients with advanced non-Hodgkin's lymphoma (stages III and IV).

PURPOSE: This study aims to investigate whether the plasma level of glutathione S-transferase P1-1 (GSTP1-1), which is a phase II detoxifying enzyme known to be a resistance factor for anticancer drugs, could be a prognostic factor of de novo non-Hodgkin lymphoma (NHL) in clinical stages (CSs) III and IV. EXPERIMENTAL DESIGN: Study population consisted of 80 NHL patients with no prior treatment: 12 patients were at CS I, 14 at CS II, 25 at CS III, and 29 at CS IV. All 54 patients at CS III or CS IV were treated with cyclophosphamide, doxorubicin, vincristine, and prednisolone (CHOP). Plasma GSTP1-1 concentration was measured by ELISA. We stained lymph node tissues for GSTP1-1 using anti-GSTP1-1 monoclonal antibody 5F and quantitatively assessed the intensity of immunostaining by using the KS-400 image analyzing system. RESULTS: There was a significant stepwise increment of plasma GSTP1-1 concentration from CS I to CS IV (P < 0.05). Of the 54 patients with CS III or IV treated with CHOP, 28 (52%) had elevated plasma GSTP1-1 levels. Plasma GSTP1-1 concentration tended to correlate with the intensity of GSTP1-1 expression in lymphoma tissues as assessed by immunostaining (P = 0.07). The CR rates in patients at CS III and CS IV treated by CHOP, 55.2% (14 of 26) and 16.0% (5 of 28) for the low and high plasma GSTP1-1 groups, respectively, were significantly different (P < 0.01). For these two groups, the median survival times were 64 and 25 months, respectively (P < 0.01), and the median times to progression were 58 and 12 months, respectively (P < 0.01). There was no significant correlation between plasma GSTP1-1 concentrations and other NHL prognostic indicators in these patients as determined by univariate and multivariate analyses. CONCLUSION: These results showed that plasma GSTP1-1 is a useful prognostic factor for CS III and IV advanced NHL. Thus, it may be a promising strategy to treat NHL concomitantly with anticancer drugs and GSTP1-1-specific inhibitors.

Adult↗
PubMed2004

Fas-mediated apoptosome formation is dependent on reactive oxygen species derived from mitochondrial permeability transition in Jurkat cells.

Generation of reactive oxygen species (ROS) and activation of caspase cascade are both indispensable in Fas-mediated apoptotic signaling. Although ROS was presumed to affect the activity of the caspase cascade on the basis of findings that antioxidants inhibited the activation of caspases and that the stimulation of ROS by itself activated caspases, the mechanism by which these cellular events are integrated in Fas signaling is presently unclear. In this study, using human T cell leukemia Jurkat cells as well as an in vitro reconstitution system, we demonstrate that ROS are required for the formation of apoptosome. We first showed that ROS derived from mitochondrial permeability transition positively regulated the apoptotic events downstream of mitochondrial permeability transition. Then, we revealed that apoptosome formation in Fas-stimulated Jurkat cells was clearly inhibited by N-acetyl-L-cysteine and manganese superoxide dismutase by using both the immunoprecipitation and size-exclusion chromatography methods. To confirm these in vivo findings, we next used an in vitro reconstitution system in which in vitro-translated apoptotic protease-activating factor 1 (Apaf-1), procaspase-9, and cytochrome c purified from human placenta were activated by dATP to form apoptosome; the formation of apoptosome was markedly inhibited by reducing reagents such as DTT or reduced glutathione (GSH), whereas hydrogen peroxide prevented this inhibition. We also found that apoptosome formation was substantially impaired by GSH-pretreated Apaf-1, but not GSH-pretreated procaspase-9 or GSH-pretreated cytochrome c. Collectively, these results suggest that ROS plays an essential role in apoptosome formation by oxidizing Apaf-1 and the subsequent activation of caspase-9 and -3.

Apoptosis↗
PubMed2004

Indian hedgehog gene transfer augments hematopoietic support of human stromal cells including NOD/SCID-beta2m-/- repopulating cells.

Hematopoietic stem cells (HSCs) are a subset of bone marrow cells that are capable of self-renewal and of giving rise to all types of blood cells. However, the mechanisms involved in controlling the number and abilities of HSCs remain largely unknown. The Indian hedgehog (Ihh) signal has an essential role in inducing hematopoietic tissue during embryogenesis. We investigated the roles of the Ihh in coculture with CD34+ cells and human stromal cells. Ihh mRNA was expressed in primary and telomerized human (hTERT) stromal cells, and its receptor molecules were detected in CD34+ cells. Ihh gene transfer into hTERT stromal cells enhanced their hematopoietic supporting potential, which was elevated compared with control stromal cells, as indicated by the colony-forming units in culture (CFU-Cs) (26-fold +/- 2-fold versus 59-fold +/- 3-fold of the initial cell number; mixed colony-forming units [CFU-Mix's], 63-fold +/- 37-fold versus 349-fold +/- 116-fold). Engraftments of nonobese diabetic/severe combined immunodeficiency-beta2m-/- (NOD/SCID-beta2-/-) repopulating cells (RCs) expanded on Ihh stromal cells were significantly higher compared with control coculture results, and engraftment was neutralized by addition of an antihedgehog antibody. Limiting dilution analysis indicated that NOD/SCID-beta2m-/- RCs proliferated efficiently on Ihh stromal cells, compared with control stromal cells. These results indicate that Ihh gene transfer could enhance the primitive hematopoietic support ability of human stromal cells.

Animals↗