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Biomedical subjects

Y Tanji

Publications and source records attributed to Y Tanji.

At least 55 records · Page 3Linked to original sources
PubMed1994

Identification of the domain required for trans-cleavage activity of hepatitis C viral serine proteinase.

A serine proteinase, Cpro-2, encoded in the hepatitis C virus (HCV) genome, is considered to be located in the N-terminal part of HCV p70, one of the putative nonstructural (NS) proteins of HCV. Cpro-2 is suggested to be responsible for producing several kinds of NS proteins by processing of the HCV precursor polyprotein. We identified the active domain of Cpro-2 and clarified the mechanism of HCV polyprotein processing; various HCV mutants deleted around this serine proteinase structure were cosynthesized with unprocessed HCV polypeptides containing Cpro-2-dependent cleavage sites in COS-1 cells. We showed that Cpro-2 cleaved the HCV precursor polyprotein intermolecularly (trans) and that Cpro-2 domain which is necessary and sufficient for that cleavage mapped to within 167 aa, from Gly1049 to Ser1215 of the HCV precursor polyprotein.

Animals↗
PubMed1994

Processing of hepatitis C viral polyprotein in Escherichia coli.

Two proteinase activities, encoded by hepatitis C virus (HCV), Cpro-1 and Cpro-2. Cpro-1 and Cpro-2 appear to process the precursor polyprotein from which they originate. Mutant HCV polypeptides containing the region for these proteinases were produced in Escherichia coli as fusion proteins. The N- and C-terminal ends of the HCV polypeptides were fused with the E. coli maltose-binding protein (MBP) and E. coli dihydrofolate reductase (DHFR), respectively. The proteinase activities cleaved the fusion polypeptides by the same processing pathway used in eukaryotic protein production systems. The N-terminal amino acid (aa) sequences of the processed fusion proteins were determined. A comparison of those N-terminal sequences with the aa sequence of the HCV precursor polyprotein showed that the N-terminal and C-terminal cleavage sites of p70(NS3), one of the HCV nonstructural (NS) proteins, were the same as those identified in other processing studies: cleavages were estimated to be between aa 1026 and 1027 and between aa 1657 and 1658 of the HCV precursor protein, which are known to be cleaved by Cpro-1 and Cpro-2, respectively. Cpro-1 and Cpro-2 both functioned in E. coli and possessed authentic characteristic features.

ATP-Binding Cassette Transporters↗
PubMed1994

Use of human leukocyte antigen-mismatched allogeneic lymphokine-activated killer cells and interleukin-2 in the adoptive immunotherapy of patients with malignancies.

Clinical effects and side effects were studied in the adoptive immunotherapy of patients bearing malignant diseases using human leukocyte antigen (HLA)-mismatched allogeneic lymphokine-activated killer (LAK) cells. Allogeneic LAK cells were induced from peripheral blood lymphocytes (PBL) of normal donors by means of initial stimulation with pokeweed mitogen (PWM). Six of 15 patients applied in the adoptive immunotherapy showed clinical effects such as partial or complete regression of pulmonary metastasis, pleural effusion and primary tumor. All pulmonary metastatic lesions were eliminated in one case by this adoptive immunotherapy combined with chemotherapy. Generally toxic effects were chillness, fever and general fatigue which were reversible, and no allergic side effects occurred even though allogeneic LAK cells were injected frequently except one patient who showed preshock like symptom accompanied with leukocytopenia and continuous hypotension immediately after infusion but was finally rescued. In the patients who received more than 10(11) of allogeneic LAK cells, anti-HLA class I antibodies appeared without any evidence of autoantibody. However, immunological side effects were never experienced after injection of allogeneic LAK cells even when the anti-HLA class I antibodies appeared in the patients. Taken together, allogeneic LAK cells could be considered as alternative therapy for patients with malignancies who could not supply sufficient materials of autologous LAK cells.

Adult↗
PubMed1994

Hepatitis C virus polyprotein processing: kinetics and mutagenic analysis of serine proteinase-dependent cleavage.

Hepatitis C virus (HCV) serine proteinase (Cpro-2) is responsible for the processing of HCV nonstructural (NS) protein processing. To clarify the mechanism of Cpro-2-dependent processing, pulse-chase and mutation analyses were performed by using a transient protein production system in cultured cells. Pulse-chase study revealed the sequential production of HCV-NS proteins. Production of p70(NS3) and p66(NS5B) were rapid. An 89-kDa processing intermediate protein (p89) was observed during the early part of the chase. p89 seemed to be cleaved first into a 31-kDa protein (p31) and a p58/56(NS5A). p31 was further processed into p4(NS4A) and p27(NS4B). Mutation analysis of cleavage sites of NS4A/4B, NS4B/5A, and NS5A/5B revealed that cleavage at each site is essentially independent from cleavage occurring at the other site.

Animals↗
PubMed1994

Analysis of N-terminal processing of hepatitis C virus nonstructural protein 2.

We determined the partial amino (N)-terminal amino acid sequence of hepatitis C virus p21 (nonstructural protein 2 [NS2]). Cleavage at the p21 (NS2) N terminus depended on the presence of microsomal membranes. The amino-terminal position of p21 (NS2) was assigned to amino acid 810 of the hepatitis C virus strain IIJ precursor polyprotein. Mutation of the alanine residue at position P1 of the putative cleavage site inhibited membrane-dependent processing. This alteration in processing together with the fact that hydrophobic amino acid residues are clustered upstream of the putative cleavage site suggested the involvement of a signal peptidase(s) in the cleavage. Furthermore, mutation analysis of this possible cleavage site revealed the presence of another microsome membrane-dependent cleavage site upstream of the N terminus of p21 (NS2).

Amino Acid Sequence↗
PubMed1994

[Therapeutic strategy for anaplastic carcinoma and malignant lymphoma of the thyroid].

Therapeutic strategies for anaplastic carcinoma and malignant lymphoma of the thyroid are reviewed. Because of the rarity of both diseases among thyroid malignancies, the generally accepted treatment policy has not been established, although the importance of radiation therapy and chemotherapy has been well recognized for the treatment of both diseases. Hyperfractionated radiation therapy combined with suitable systemic chemotherapy will counter anaplastic thyroid carcinoma and prolong survival. Most patients with thyroid malignant lymphoma have better prognoses than anaplastic carcinoma patients. But delayed diagnosis and inadequate treatment still results in a poor prognosis, in spite of the more treatable characteristics of thyroid malignant lymphoma than anaplastic thyroid carcinoma. At present, early detection of the tumor and selection of a suitable therapy is necessary for the improvement of prognosis, and further clinical studies are required to establish effective regimens for multidisciplinary treatment.

Carcinoma↗
PubMed1993

Proteolytic processing and membrane association of putative nonstructural proteins of hepatitis C virus.

By using a plasmid-based transient protein expression system in cultured cells and an in vitro transcription/translation system, we analyzed the proteolytic processing of the putative nonstructural protein region of the precursor polyprotein from a Japanese type of hepatitis C virus. In addition to the previously reported viral proteins, p21 and p70, we identified products of 4 kDa (p4), 27 kDa (p27), 56 kDa (p56), 58 kDa (p58), and 66 kDa (p66). These products were produced in a viral serine proteinase (proteinase 2)-dependent manner from the region downstream of p70 in the precursor polyprotein and were arranged as NH2-p70-p4-p27-p58(p56)-p66-COOH as determined with region-specific antibodies. We showed that p56 was an N-terminally truncated form of p58, which suggested that a small polypeptide of 2 kDa (p2) was produced from the N-terminal part of p58. Cleavage between p4 and p27 was inefficient in vitro and we saw the 31-kDa precursor polypeptide (p31) accumulate. Furthermore, efficient cleavage at this site in vivo required the presence of p58/p56. Immunoprecipitation analysis in vitro also suggested the mutual interaction of those nonstructural protein products. An especially close association of p4 with p70 may contribute to association of p70 with microsomal membranes.

Animals↗
PubMed1993

Two proteinase activities in HCV polypeptide expressed in insect cells using baculovirus vector.

Processing of HCV viral precursor protein requires at least two viral proteinases, Cpro-1 and Cpro-2, in addition to cellular proteinases. The HCV polypeptide that covers the region for the two viral proteinase domains was expressed in insect cells using a baculovirus expression system. The two proteinase activities were demonstrated in the infected cells. The Cpro-1-dependent cleavage site was estimated from the amino acid sequence of the N-terminus of the processed product. Analyses of the susceptibilities of various mutants altered at position P1 and P1' of the putative cleavage site suggested that amino acid residues at these positions is not essential for recognition and cleavage by Cpro-1-dependent activity.

Amino Acid Sequence↗
PubMed1991

Effect of OmpA signal peptide mutations on OmpA secretion, synthesis, and assembly.

In previous investigations, we have examined the effect of OmpA signal peptide mutations on the secretion of the two heterologous proteins TEM beta-lactamase and nuclease A. During these studies, we observed that a given signal peptide mutation could affect differentially the processing of precursor OmpA-nuclease or precursor OmpA-lactamase. This observation led us to further investigate the influence of the mature region of a precursor protein on protein export. Preexisting OmpA signal peptide mutations of known secretion phenotype when directing heterologous protein export (nuclease A or beta-lactamase) were fused to the homologous mature OmpA protein. Four signal peptide mutations that have previously been shown to prevent export of nuclease A and beta-lactamase were found to support OmpA protein export, albeit at reduced rates. This remarkable retention of export activity by severely defective precursor OmpA signal peptide mutants may be due to the ability of mature OmpA to interact with the cytoplasmic membrane. In addition, these same signal peptide mutations can affect the level of OmpA synthesis as well as its proper assembly in the outer membrane of Escherichia coli. Two signal peptide mutations dramatically stimulate the rate of precursor OmpA synthesis three- to fivefold above the level observed when a wild-type signal peptide is directing export. The complete removal of the OmpA signal peptide does not result in increased OmpA synthesis. This finding suggests that the signal peptide mutations function positively to stimulate OmpA synthesis, rather than bypass a down-regulatory mechanism effected by a wild-type signal peptide. Overproduction of wild-type precursor OmpA or precursors containing signal peptide mutations which lead to relatively minor kinetic processing defects results in accumulation of an improperly assembled OmpA species (imp-OmpA). In contrast, signal peptide mutations which cause relatively severe processing defects accumulate no or only small quantities of imp-OmpA. All mutations result in equivalent levels of properly assembled OmpA. Thus, a strong correlation between imp-OmpA accumulation and cell toxicity was observed. A mutation in the mature region of OmpA which prevents the proper outer membrane assembly of OmpA was suppressed when export was directed by a severely defective signal peptide. These findings suggest that signal peptide mutations indirectly influence OmpA assembly in the outer membrane by altering both the level and rate of OmpA secretion across the cytoplasmic membrane.

Amino Acid Sequence↗
PubMed1990

Successful direct PTCA on LAD after first episode of acute myocardial infarction: does it improve cardiac function?

Patients who received direct percutaneous transluminal coronary angioplasty (PTCA) after acute mycardial infarction and maintained potency but with unimproved cardiac function were studied. In 15 patients, the first episode of acute myocardial infarction was caused by a left anterior descending branch lesion; 11 had an ejection fraction of 50% or more in the left ventriculogram in the follow-up period (improved group), and 4 patients had ejection fraction of less than 50% (unimproved group). There was so significant difference between the groups in the mean time between the onset of infarction and revascularization (improved group, 259.3 +/- 76.9 min; unimproved group, 168.0 +/- 101.6 min) or in the sigma Q. which was the sum of the Q wave depth of V2, V3, and V4 at the time of admission (improved group, 12.1 +/- 15.6 mm; unimproved group 29.8 +/- 13.4 mm). The maximum creatine kinase concentration was significantly higher in the unimproved group (improved group 2670 +/- 893 IU/L; unimproved group, 7243 +/- 1928 IU/L, p less than 0.05), and the time taken from the onset to reach its peak was significantly shorter in the unimproved group (improved group, 13.0 +/- 5.1 hr; unimproved group, 6.8 +/- 1.3 hr, p less than 0.05.) These results suggest the probability of sudden deterioration of myocardium, and factors other than microcirculatory thromboembolism should be considered as the cause of unimproved cardiac function after successful direct PTCA.

Aged↗
PubMed1989

[Mass culture of LAK cells by a hollow-fiber bioreactor system].

We attempted to culture LAK cells by the use of the hollow-fiber bioreactor system, "Acusyst-P". The number of LAK cells increased by 30-40 times. The majority of LAK cells cultured by this hollow fiber system originated in T cell. LAK cells cultured by hollow-fiber system were different from LAK cells statically cultured in their cytotoxicity and change of phenotype. We obtained LAK cells more efficiently and safely which have higher viability and cytotoxicity by the use of hollow-fiber bioreactor system than by static culture.

Cytological Techniques↗
PubMed1987

Augmentation of cytotoxic activity of recombinant human tumor necrosis factor (rHu-TNF) by recombinant human interferon-gamma (rHu-IFN-gamma).

The present study was undertaken to examine the effect of recombinant human interferon-gamma (rHu-IFN-gamma) on the in vitro antitumor activity of recombinant human tumor necrosis factor (rHu-TNF) against rHu-TNF-sensitive and resistant tumor cells. rHu-IFN-gamma augmented both cytostatic and cytocidal activity of rHu-TNF. When 14 tumor cells were tested, augmentation by rHu-IFN-gamma was observed in most of rHu-TNF-sensitive tumor cells and in few cases in rHu-TNF-resistant tumor cells. Among rHu-TNF-sensitive tumor cells, there was no relationship between the degree of augmentation and the susceptibility to rHu-TNF. Augmentation was marked when tumor cells were treated with rHu-IFN-gamma either before the exposure to rHu-TNF or during the early period of the exposure to rHu-TNF. On the other hand, augmentation was not observed when tumor cells were treated with rHu-IFN-gamma during a late period of the exposure to rHu-TNF. From these results, a combined treatment with both agents can be expected to provide an approach to get better results in clinical trials.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed1987

[Adoptive immunotherapy of malignant diseases with LAK cells].

Peripheral blood lymphocytes obtained from patients by leukapheresis were cultured in RPMI 1640 containing human plasma and interleukin 2. The morphology, phenotypes and cytotoxicity of induced LAK cells were studied. Lymphoblastoid cells mainly proliferated were OKIa1+ cells and were thought to be LAK cells. Maximal cytotoxicity was obtained after two weeks of incubation. IL-2 enhanced the cytotoxicity of LAK cells. Autologous LAK cells induced by two weeks of incubation were injected into patients. One case of pulmonary metastases of breast cancer showed reduction and two lesions showed partial regression. Also, no new lesions appeared in the lungs of a patient with alveolar soft-part sarcoma.

Adult↗
PubMed1987

[Adoptive immunotherapy of malignant diseases using normal allogeneic LAK cells].

Allogeneic LAK cells induced from PBL of normal donors were used for adoptive immunotherapy of malignant diseases. The possibility of an antibody against allogeneic LAK cells was suggested. However, no immune reaction was observed and LAK activity was maintained in vitro in the presence of patient's plasma sampled during the course of therapy.

Graft vs Host Reaction↗
PubMed1987

[Blocking factors in human blood which inhibit the cytotoxicity of lymphocytes].

The cytotoxicity of human peripheral blood lymphocytes against malignant tumor cell lines was significantly inhibited by human plasma or sera. In addition to so-called blocking factors such as antigens, antibodies, antigen-antibody complexes, IAP and ferritin, some substances which could be removed by heat or centrifugation were considered to play a significant role.

Antibody-Dependent Cell Cytotoxicity↗