PubMed Health⌕ Search

Biomedical subjects

N Emi

Publications and source records attributed to N Emi.

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

No significant association between HA-1 incompatibility and incidence of acute graft-versus-host disease after HLA-identical sibling bone marrow transplantation in Japanese patients.

We retrospectively examined HA-1 typing with polymerase chain reaction using sequence-specific primers in 120 samples from 60 HLA-A2-positive Japanese bone marrow transplantation recipients who received short-term methotrexate and cyclosporin A for graft-versus-host disease (GVHD) prophylaxis and their HLA-identical sibling donors. HA-1-incompatible pairs were observed in 22% of the samples. The probability of developing acute GVHD (grade II to IV) in HA-1-incompatible and -compatible patients was 0% and 19%, respectively (P = .10). In a comparison between HA-1-incompatible and -compatible patients with standard-risk leukemia, in whom age, patient/donor sex, and use of a total body irradiation-containing regimen were equivalent, the probability of developing acute GVHD (grade II to IV) was 0% and 10%, respectively (P = .38). No evidence of recurrent leukemia was observed in the HA-1-incompatible patients with standard-risk leukemia, compared with 37% in HA-1-compatible patients (P = .11). In conclusion, HA-1 incompatibility may not be a risk factor for grade II to IV acute GVHD in Japanese patients who receive methotrexate and cyclosporin A and undergo bone marrow transplantation from HLA-identical sibling donors.

Adolescent↗
PubMed2000

Nutritional therapy of chronic hepatitis by whey protein (non-heated).

In an open study the clinical efficacy of milk serum (whey) protein (Immunocal; cysteine content: 7.6-fold higher than that of casein) isolated from fresh milk and purified without heating was evaluated in 25 patients with chronic hepatitis B or C. Immunocal (12 g as protein) food (mousse) was given twice a day, in the morning and evening, for 12 weeks (test period). Casein (12 g as protein) food (mousse) was similarly given for two weeks prior to the start of the supplement with Immunocal food (induction period) and for four weeks after the end of the supplement with Immunocal food (follow-up period). Serum alanine aminotransferase (ALT) activity was reduced, and plasma glutathione (GSH) levels increased in six and five of eight patients with chronic hepatitis B, respectively, 12 weeks after the start of the supplement with Immunocal food. Serum lipid peroxide levels significantly decreased, and interleukin (IL)-2 levels and natural killer (NK) activity significantly increased. However, there were no significant Immunocal-related changes in 17 patients with chronic hepatitis C. These findings suggest that the long-term supplement with Immunocal alone may be effective for improving liver dysfunctions in patients with chronic hepatitis B.

Adult↗
PubMed1999

Conjugation of plasmid DNAs with lactose via diazocoupling enhances resistance to restriction enzymes and acquires binding affinity to galactose-specific lectin.

Oligosaccharide-plasmid DNA conjugates were synthesized simply and effectively via the diazocoupling method. Plasmids (pUC19, pTRI-beta-actin, and pEGFP-C1) were treated with an N-beta-lactoside-substituted diazonium salt to yield diazocoupling products with degree of substitutions of 2.5-3.1 mol% of overall nucleobases. The lactose-pUC19 conjugate was found to resist restriction enzymes more strongly than the nonconjugated plasmid DNA and to acquire a strong binding affinity to galactose-specific lectin RCA(120). The diazocoupling modification of pTRI-beta-actin plasmid DNA little influenced in vitro transcription with T7 RNA polymerase. When lactose-pEGFP-C1 conjugate was transfected to baby hamster kidney (BHK) cells by means of cationic lipids, transduced gene was expressed in BHK cells similarly with the nonconjugated pEGFP-C1. The modification of plasmid DNA with carbohydrate enhanced the resistance to restriction enzymes and developed a strong binding affinity to galactose-specific lectin.

Actins↗
PubMed1999

Successful culture and sustainability in vivo of gene-modified human oral mucosal epithelium.

Human oral mucosal cells are an attractive site for tissue engineering because they are the most accessible cells in the body and easy to manipulate in vitro. They thus have possibilities for targeting by somatic gene therapy. We examined the efficiency of retrovirus-mediated gene transfer and the construction of mucosal epithelium in vivo. Human oral mucosal cells were transduced with a retroviral vector carrying the lacZ gene at high efficiency and constructed epithelium after G418 selection with 3T3 cells in vitro. The cultured oral mucosal epithelium membrane was then grafted onto immunodeficient mice. Beta-Gal expression was detected histochemically in vivo 5 weeks after grafting. Furthermore, we transduced factor IX cDNA into the mucosal epithelium membrane, and it was then transplanted into nude mice. Between 0.6 and 1.8 ng of human factor IX per milliliter was found in mouse plasma, and the production was continued for 23 days in vivo. These results confirmed that the oral mucosal epithelium is an ideal target tissue for gene therapy or tissue engineering.

Animals↗
PubMed1999

Immune response of post-transplant peripheral lymphocytes against the patient pre-B cell line, NAGL-1.

We have established a pre-B acute lymphoblastic leukemia (ALL) cell line, NAGL-1, from the bone marrow of a patient diagnosed with pre-B ALL. The patient has been disease-free for the 4 years since allogeneic bone marrow transplantation from her HLA-genotypically identical sister. NAGL-1 showed a pre-B cell phenotype (CD19+, CD10+, c mu+, s mu-) mostly identical to freshly isolated leukemic cells from the patient. This cell line strongly expressed HLA class I and HLA-DR molecules, as well as the costimulatory molecules CD54, CD40, and CD86. Cytotoxic T-lymphocyte (CTL) lines were generated by stimulating the donor-derived peripheral blood mononuclear cells with either irradiated leukemic cells or NAGL-1. Both CTL lines showed specific lysis against NAGL-1 in 51Cr release assays. Lytic activity was partially inhibited by anti-CD8 and anti-HLA class I monoclonal antibodies. Treatment of NAGL-1 with TNF-alpha increased its susceptibility to the CTL line. One CD8+ T cell clone derived from the CTL line killed both the patient phytohemagglutinin (PHA) blasts and NAGL-1 but not the donor PHA blasts, suggesting that the clone recognized the patient-specific minor antigen presented on both PHA blasts and NAGL-1. Utilization of leukemic cell lines could be a useful model for the development of CTL lines and clones for immunological study and potential immunotherapy.

Adolescent↗
PubMed1999

CD4- and CD56-positive T-cell line, MTA, established from natural killer-like T-cell leukemia/lymphoma.

We have established a T-cell line, MTA, from the peripheral blasts of a patient with natural killer (NK)-like T-cell leukemia/lymphoma. The MTA cell displays a T-cell and NK-cell phenotype (CD2+, CD3+, CD4+, CD56+) identical to freshly isolated leukemic blasts from the patient. This cell line shows clonal T-cell receptor rearrangement and a distinguishable character by light microscopy with May-Grünwald Giemsa staining. G-banding analysis showed that the MTA cells had a karyotype of 94(4N), XXXX, add (1) (p36), del (5) (q14q23), add (17) (p11), add (19) (q13). However, unlike NK malignancy, we could not show a direct pathogenic role for Epstein-Barr virus (EBV) with EBV-encoded small RNA and polymerase chain reaction analysis. The MTA cell line is a novel cell line with which to study NK-like T-cell ontogenecity.

CD4 Antigens↗
PubMed1998

Feasibility of double-expression retroviral vector using complement regulatory factor gene.

The donor source of vascular endothelial cells for hybrid blood vessels seeded with genetically engineered endothelial cells is generally considered to be autologous. The purpose of this study was to determine whether porcine endothelial cells transduced with double-expression retroviral vector using complement-resistant gene could be substituted for autologous endothelial cells. Decay-accelerating factor (DAF) and tissue plasminogen activator (tPA) cDNA were inserted into retroviral vector with homologous restriction factor 20 cDNA as a complement regulatory factor gene. Porcine aortic endothelial cells were transduced with these double-expression retroviral vectors, followed by the complement-dependent selection. Porcine endothelial cells transduced withdouble-expression retroviral vectors showed a high gene expression of both DAF and tPA. Complement-dependent cytotoxicity and adherence of U937 were significantly inhibited by the transduction of double-expression vectors with complement regulatory factor gene. Double-expression retroviral vector using complement regulatory factor gene was efficacious in substituting porcine endothelial cells for the autologous endothelial cells.

3T3 Cells↗
PubMed1998

Recombinant adeno-associated virus-mediated gene transfer into human leukemia cell lines.

Adeno-associated virus (AAV)-based vector is a promising gene transfer vehicle by virtue of the characteristics of wild-type AAV:tropism to a wide range of human tissues and locus-specific integration at chromosome 19q13.3. To elucidate the nature of the recombinant AAV (rAAV), transduction of neomycin phosphotransferase enzyme gene (NeoR gene) into seven human leukemia cell lines was performed. Transduction efficiencies were assessed by colony formation assay and limiting dilution assay. The results suggested that both assays are comparable. Transduction efficiencies of the NeoR gene into K-562, MEG-O1, Raji, MOLT-3, HL-60, U937 and NKM-1 at a multiplicity of infection (MOI) of 0.1 were 0.27, 0.25, 0.015, 0.009, < 0.0025 and < 0.0025%, respectively. After purification and concentration of rAAV, 27% efficiency was observed in K562 at an MOI of 7 and a linear relationship between MOI and efficiency was confirmed, suggesting that this system may be useful for gene transduction into leukemia cells. Integration of the NeoR gene into the host genome was detected by Southern blotting analysis, which showed various sizes of digested fragments. A fluorescent in situ hybridization (FISH) study was carried out on 11 clones, in all of which the NeoR gene was integrated out of chromosome 19q13.3. In five of the clones, whole chromosome painting probes revealed that the integration sites were chromosomes 1q, 2q, 2q, 11p, 12p and 13q.

Dependovirus↗
PubMed1998

Establishment of complement-resistant retroviral vector by homologous restriction factor 20 gene.

Homologous restriction factor 20 (HRF20, CD59) is one of the complement regulatory factors. In this study, the complement-resistant retroviral vector, which possesses the HRF20 gene as a selection gene, was constructed and examined. The virus-producing cell, transduced with complement-resistant retroviral vector, was established after complement-dependent selection. NIH3T3 and PK15 cells transduced with this virus-producing cell were successfully selected by complement-dependent selection, which showed significant expression of the HRF20 antigen. In addition, these cells, transduced with tissue plasminogen activator (tPA) cDNA using complement-resistant retroviral vector, expressed tPA antigen after complement-dependent selection. These findings suggest that complement-resistant retroviral vector can be used for the double transduction of HRF20, as well as other genes.

3T3 Cells↗
PubMed1998

Semliki Forest virus-based DNA expression vector: transient protein production followed by cell death.

We have constructed a novel DNA expression vector based on Semliki Forest virus (SFV). SFV produces non-structural proteins (nsPs) which replicate genomic RNA and amplify the mRNA encoding the structural proteins of SFV. A recombinant cDNA genome of SFV, in which the SFV structural genes were replaced by a polylinker cassette to allow for insertion of heterologous DNA, was placed under the control of a cytomegalovirus immediate-early enhancer/promoter with a polyadenylation signal. Transfection of mammalian cells with this SFV-based plasmid vector, pSFV3-CMV-lacZ-pA, resulted in transient high-level expression of a beta-galactosidase reporter gene. The expression level of beta-galactosidase from pSFV3-CMVlacZ-pA was more than 20-fold higher than that obtained from the plasmid with deleted nsPs genes, pSFV3A5976-lacZ, demonstrating that the nsPs genes were essential for the high level of expression. Substantial beta-galactosidase activity was detected in the medium of pSFV3-CMV-lacZ-pA-transfected cells, suggesting that the overproduction of beta-galactosidase caused cell death and release of the protein into the medium. We have demonstrated a high-level expression of the exogenous beta-galactosidase gene from pSFV3-CMV-lacZ-pA constructed using an SFV replication system.

Animals↗
PubMed1998

Protection of xeno-hepatocytes from complement-mediated cytolysis by transduction with homologous restriction factor 20 gene using retroviral vector.

Xeno-hepatocyte transplantation has the possibility to substitute for clinical liver transplantation in certain restricted hepatic diseases such as inherited metabolic disorders. To overcome human complement-dependent cytotoxicity on xeno-hepatocytes, the effectiveness of ex vivo transfer with the homologous restriction factor 20 (HRF20, CD59) gene was examined on primary-cultured xeno-hepatocytes using a retroviral vector. Primary-cultured rat hepatocytes transduced with HRF20 cDNA expressed HRF20 antigen by flow cytometric analysis and showed the integration of HRF20 cDNA to the genomic DNA by the polymerase chain reaction. The viability of rat hepatocytes incubated with 50% human serum was decreased due to complement-dependent cytotoxicity, whereas that of the transfectant was significantly protected (77.2+/-9.4 vs. 97.8+/-5.2%, p < 0.01). It was concluded that primary-cultured xeno-hepatocytes transduced with HRF20 cDNA using a retroviral vector could escape complement-dependent cytolysis by human serum.

Animals↗
PubMed1998

Bystander tumoricidal effect and gap junctional communication in lung cancer cell lines.

Tumor cells expressing the herpes simplex virus-thymidine kinase (HSV-tk) gene become sensitive to ganciclovir (GCV), and the phenomenon by which tumor cells surrounding the HSV-tk expressing cells also become sensitive to GCV is known as the "bystander effect." The purpose of this study was to investigate the bystander effect in human lung-cancer cell lines, and the role of gap-junctional intercellular communication as the mechanism responsible for it. Gap-junctional intercellular communication was measured both with a dye-transfer assay involving single-cell microinjection of Lucifer Yellow and with a PKH26/calcein-AM double-dye-transfer assay. Significant bystander tumoricidal effect was observed in lung-cancer cell lines when cultured cells contained only 10% HSV-tk expressing cells. This was also observed to occur with cell lines of different origin or from different species. Although gap-junctional intercellular communication characterized by rapid transfer of Lucifer Yellow was not observed, we did detect gap-junctional communication marked by the slow transfer of calcein-AM in lung-cancer cell lines. However, neither an inhibitor (1-octanol) nor an enhancer (all trans-retinoic acid [ATRA]) of gap-junctional communication affected the extent of the bystander effect. These findings suggest that low levels of gap-junctional communication may be efficient for producing the bystander effect in lung-cancer cells, or that other mechanisms may underlie this effect. Although gap-junctional communication may play an important role in generating the bystander effect in tumor cells expressing the HSV-tk gene, further knowledge of the mechanism of this effect may help improve the treatment of lung cancer with an HSV-tk system.

1-Octanol↗
PubMed1998

Analysis of prognostic factors in newly diagnosed acute promyelocytic leukemia treated with all-trans retinoic acid and chemotherapy. Japan Adult Leukemia Study Group.

PURPOSE: We conducted a multicenter study of differentiation therapy with all-trans retinoic acid (ATRA) followed by intensive chemotherapy in patients with newly diagnosed acute promyelocytic leukemia (APL) and analyzed the prognostic factors for predicting complete remission (CR), event-free survival (EFS), and disease-free survival (DFS). PATIENTS AND METHODS: All patients received ATRA until CR. If patients had an initial leukocyte count greater than 3.0 x 10(9)/L, they received daunorubicin (DNR) and behenoyl cytarabine (BHAC). During therapy, if patients showed blast and promyelocyte counts greater than 1.0 x 10(9)/L, they received additional DNR and BHAC. After achieving CR, patients received three courses of consolidation and six courses of maintenance/intensification chemotherapy. RESULTS: Of 198 registered, 196 were assessable (age range, 15 to 86 years; median, 46) and 173 (88%) achieved CR. Multivariate analysis showed that no or minor purpura at diagnosis (P = .0046) and age less than 30 years (P = .0076) were favorable factors for achievement of CR. Predicted 4-year overall survival and EFS rates were 74% and 54%, respectively, and the 4-year predicted DFS rate for 173 CR patients was 62%. Multivariate analysis showed that age less than 30 years (P = .0003) and initial leukocyte count less than 10 x 10(9)/L (P = .0296) were prognostic factors for longer EFS, and initial leukocyte count less than 10.0 x 10(9)/L was a sole significant prognostic factor for longer DFS (P = .0001). CONCLUSION: Our results show that age, hemorrhagic diathesis, and initial leukocyte count are prognostic factors for APL treated with ATRA followed by intensive chemotherapy.

Adolescent↗
PubMed1997

Fusion of the platelet-derived growth factor receptor beta to a novel gene CEV14 in acute myelogenous leukemia after clonal evolution.

Chromosomal translocations involving band 5q31-35 occur in several hematologic disorders. A clone with a t(5; 14)(q33; q32) translocation appeared at the relapse phase in a patient with acute myelogenous leukemia who exhibited a sole chromosomal translocation, t(7; 11), at initial diagnosis. After the appearance of this clone, the leukemia progressed with marked eosinophilia, and combination chemotherapy was ineffective. Southern blot analysis showed a rearrangement of the platelet-derived growth factor receptor beta (PDGFRbeta) gene at 5q33 which was not observed at initial diagnosis. This translocation resulted in a chimeric transcript fusing the PDGFRbeta gene on 5q33 with a novel gene, CEV14, located at 14q32. Expression of the 5' region of the PDGFRbeta cDNA, upstream of the breakpoint, was not detected. However, the 3' region of PDGFRbeta, which was transcribed as part of the CEV14-PDGFRbeta fusion gene, was detected. A partial cDNA for a novel gene, CEV14, includes a leucine zipper motif and putative thyroid hormone receptor interacting domain and is expressed in a wide range of tissues. The expression of a CEV14-PDGFRbeta fusion gene in association with aggressive leukemia progression suggests that this protein has oncogenic potential.

Amino Acid Sequence↗
PubMed1997

Controlled gene expression with a reverse tetracycline-regulated retroviral vector (RTRV) system.

A retroviral vector was constructed with an autoregulatory cassette to allow expression of the gene of interest in response to oral administration of doxycycline (Dox) in vivo. The cassette contains all the components of the reverse tetracycline-regulated (rtTA) system, a drug selectable marker with the internal ribosome entry site and the gene of interest (GFP). FACS analyses showed an induction of GFP-fluorescence of two orders of magnitude in retrovirus-infected 208F cells dependent on the amount of Dox in the medium. Furthermore, oral administration of Dox resulted in GFP expression in transplanted 208F cells in the peritoneal cavity of nude mice. Thus this reverse tetracycline-regulated retroviral vector (RTRV) system simplifies the delivery of controllable genes to cultured and implanted cells. It is hoped that this approach may pave the way to controlled gene expression during a particular window of time in gene therapy applications.

Animals↗
PubMed1997

VSV-G-pseudotyped retroviral packaging through adenovirus-mediated inducible gene expression.

It has been technically difficult to generate recombinant adenoviruses encoding genes for cytotoxic products such as vesicular stomatitis virus G-protein (VSV-G), which is too toxic for the host cells to allow adenoviral propagation. In our companion paper (Yoshida, Y., and Hamada, H., Biochem. Biophys. Res. Commun., 230, 426-430, 1997), a tetracycline-inducible adenovirus system is reported. The inducible expression system enabled us to generate recombinant adenoviruses encoding genes for the cytotoxic viral VSV-G product. In this study, we generated recombinant adenoviruses encoding VSV-G and MoMLV gag-pol genes, both under the tetracycline-controllable promoter, and attempted retroviral packaging. Simultaneous infection of these adenoviruses together with tetracycline-transactivator (NtTA) expression resulted in efficient VSVG-pseudotyped retroviral packaging. Adenovirus-mediated recombinant retrovirus generation will be useful in studies with various pseudotyped mutants, as well as in assays for retrovirus-related genes and their products.

Adenoviridae↗