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Y Hinuma

Publications and source records attributed to Y Hinuma.

At least 109 records · Page 6Linked to original sources
PubMed1984

Microplate method for retrovirus-induced transformation of normal human T-cells.

An adult T-cell leukemia virus (ATLV) producer cell line (TL-Su) derived from a healthy ATLV-carrier, was used for virus-induced transformation of normal human T-cells. The transformation was carried out on microplates by co-cultivation of a limiting number of normal peripheral blood leukocytes (PBL) as transformed target cells and 2 X 10(4)/well of TL-Su cells irradiated lethally. The frequency of establishment of transformed cell lines correlated with the dose of PBL. The transforming target cell number was estimated to be approximately one per 500 PBL by the microplate method using TL-Su effector cells. When 1 X 10(3) PBL per well were plated, more than 90% of wells showed immortalizing transformation of PBL. Twenty transformed cell lines obtained from microplates with 500 PBL per well were shown to be of T-cell lineage. Ten of them were defined as belonging to the helper/inducer T-cell subset with Leu 3a antigen, and 4 were demonstrated to be of the cytotoxic/suppressor T-cell subset with Leu 2a antigen. The transformed cells always expressed Tac and OKT9 antigens (considered to be IL2 and transferrin receptors respectively). ATLV antigens were also expressed in all these cell lines tested, but their molecular species detected by radioimmunoprecipitation varied regardless of PBL donors and T-cell subsets. We propose this microplate transformation method as an advantageous system for obtaining abundant T-cell lines and for screening their immunological functions.

Antigens, Surface↗
PubMed1984

28,000-dalton polypeptide (p28) of adult T-cell leukemia-associated antigen encoded by 24 S mRNA of human T-cell leukemia virus has an associated protein kinase activity.

Human T-cell leukemia virus producer cell line MT-2 was labeled with [32P]phosphoric acid, and its cell extracts were immunoprecipitated with mouse monoclonal antibodies (GIN-7, and KK-1) and rabbit sera (anti-p24, and anti-gp68). Analysis of the immunocomplexes on sodium dodecyl sulfate-polyacrylamide gell electrophoresis revealed that p53, p28, and p19 of adult T-cell leukemia-associated antigens were phosphorylated in vivo. Immunocomplexes of MT-2 cell extract with monoclonal antibody KK-1 were incubated with [gamma-32P]ATP in vitro and it was revealed that the phosphokinase activity was associated with p28. The phosphokinase activity of p28 was specific to the serine residue but was not to the tyrosine residue.

Animals↗
PubMed1984

Cell surface phenotypes and expression of viral antigens of various human cell lines carrying human T-cell leukemia virus.

Human T-cell leukemia/lymphoma virus (HTLV)-carrying cells from various origins were characterized by cell surface markers and expression of HTLV antigens. Eight cell lines named TCL were obtained by transformation of peripheral blood leukocytes (PBL) of healthy donors or HTLV carriers in cocultures with HTLV-producer MT-2 cells. Nine cell lines named ILT were interleukin 2 (IL2)-dependent cell lines cloned from PBL of ATL patients and healthy HTLV-carriers. Tc-Kan9 cell line was also an IL2-dependent cell line clonally established from PBL culture stimulated with autologous TCL cells. Five cell lines named TL were established in vitro directly from PBL of an adult T-cell leukemia (ATL) patient and from ILT cells of an ATL patient and three HTLV-carriers, respectively, to grow autonomously without IL2. All the TCLs, ILTs, TLs and Tc-Kan9 possessed Leu-I antigen, a pan-T-cell marker. Leu3a antigen, a helper/inducer T-cell marker, was expressed on five of eight TCLs and all of the ILTs and TLs. Leu-2a, a cytotoxic/suppressor T-cell marker, was detected only on Tc-Kan9 but not others. Fresh ATL leukemic cells of patients had a helper/inducer T-cell marker. Ia, OKT9 and Tac antigens, markers for activated and differentiated T cells, were strongly expressed on all of the cell lines tested and fresh ATL leukemic cells were weakly positive for these antigens. Expression of HTLV antigens detected by mouse monoclonal antibodies and an ATL-patient serum varied among these cell lines. One TL, two ILTs and most of the fresh ATL leukemic cells did not express HTLV antigens on the cell surface. The other cell lines were all positive for the surface viral antigens. However, molecular species of antigens defined by radioimmunoprecipitation with an ATL-patient serum were not always identical among the cell lines. Molecular weights of polypeptides detectable in most of the cell lines were 62K, 46K, 40K, 24K, 21K and 19K which could never be detected in several control T-cell lines. 68K and 28K polypeptides were frequently detected in MT-2 and TGLs. GIN14, a mouse monoclonal antibody against HTLV core protein (p19) detected not only p19 in various cell lines but also p28, p29, p31 or p40 in certain cell lines tested. B-cell lines named LCL were established and cloned from PBL of two HTLV-carriers by EB-virus-induced transformation and they also expressed HTLV antigens, Ia, OKT9 and Tac antigens.(ABSTRACT TRUNCATED AT 400 WORDS)

Adult↗
PubMed1984

Characterization of African green monkey B-cell lines releasing an adult T-cell leukemia-virus-related agent.

Eight lymphoblastoid cell lines were established from the peripheral blood of individual African green monkeys (AGM). The AGM-2206 line grew out spontaneously. The others - AGM-6, 7, 8, 10, 12, 13, and 16 - were obtained after infection of peripheral AGM lymphocytes with cell-free culture supernatant of AGM-2206. All lines contained, and were probably transformed by, AGM-EBV. Moreover, they expressed immunoglobulins but lacked the Leu l T-cell marker. Thus they were B cells. Since a high percentage of AGMs are naturally infected with a virus similar to adult T-cell leukemia virus (ATLV), we examined these cell lines for ATLV. With immunofluorescence tests we detected ATLV-related antigens (ATLA) in three of the eight cell lines. EBV membrane antigen was present in three out of four. The highest percentage (40%) of ATLA-positive cells was found in the AGM-13 line. After metabolic labelling of these cells, ATLV-specific polypeptides p24, p19, p15, and p10 were detected. Hybridization experiments showed that both AGM-2206 and AGM-13 cell lines contained ATLV-proviral DNA. Electron micrographs of AGM-13 revealed a few type-C particles morphologically similar to the MT-2 virus. By cocultivation this AGM virus was able to infect and immortalize human peripheral blood lymphocytes. One such human cell line, NA-13, expressed polypeptides closely related to ATLV core antigens but a 68,000 mol.wt. glycopolypeptide was serologically distinct from MT-2 ATLV gp68.

Animals↗
PubMed1984

Prevalence of possible adult T-cell leukemia virus-carriers among volunteer blood donors in Japan: a nation-wide study.

A nation-wide survey of carriers of adult T-cell leukemia virus (ATLV) detected as anti-ATLA (ATLV-associated antigens) in volunteer blood donors was made in Japan. A total of 200 sera from donors between 40 and 64 years of age, collected in each of 64 blood centers located in eight districts, were tested for anti-ATLA by an indirect immunofluorescence test. The highest rate (8%) of seropositive donors was found in Kyushu and lower rates of 0.3 to 1.2% in other districts. However, the positive rate varied considerably in different localities even within districts with a high or low seropositive incidence. Further studies of all the donors (16 to 64 years old) in eight centers in Kyushu demonstrated two facts: (1) that the frequency of seropositive donors increased with age, and (2) that four centers in this district showed distinctly high incidences of seropositive donors, while the other four centers showed low incidences. The seropositive rate among all donors was estimated to be about 3% in Kyushu and 0.08 to 0.3% in all other districts.

Adolescent↗
PubMed1984

Prevalence of antibody to adult T-cell leukemia virus-associated antigens (ATLA) in Japanese monkeys and other non-human primates.

The prevalence of adult T-cell-leukemia virus (ATLV) infection was examined in Japanese monkeys living naturally in various parts of Japan and in other species of non-human primates imported into and kept in Japan. Sera of 2,650 Japanese monkeys from 41 troops throughout Japan were tested. High incidences of anti-ATLV-associated antigen (ATLA)-positive monkeys were found in most troops, not only in the endemic area of human ATL(Southwestern Japan), but also in non-endemic areas. The incidence of sero-positive individuals increased gradually with age, reaching a maximum when the animals became adult, indicating age dependency, like that found by epidemiological studies on humans. Anti-ATLA antibodies were also detected in 90 of 815 sera of imported non-human primates of 33 species other than Japanese monkeys. All the anti-ATLA sero-positive monkeys were Catarrhines (Old World monkeys), mainly macaques of Asian origin. Some sero-positive monkeys were also found among animals of African origin, but no antibody was detected in Prosimians and Platyrrhines (New World monkeys). The clear-cut difference between the geographical distribution of sero-positive simians and that of humans indicates the improbability of direct transmission of ATLV from simians to humans.

Age Factors↗
PubMed1984

Titers of Epstein-Barr virus-related antibodies in nasopharyngeal carcinoma in Japan.

It is thought that nonkeratinizing or undifferentiated squamous cell carcinoma in the nasopharynx (NPC) is intimately correlated with Epstein-Barr Virus (EBV). Twenty-one patients with NPC were followed in Kyoto University Hospital and 4 in Osaka Red Cross Hospital during the past 2 years from 1980 to 1981. These patients were classified histopathologically according to the WHO classification in 1978 and staged with the TNM classification in Union Internationale Contre le Cancer (UICC) in 1978. The incidence rate of NPC among the head and neck tumors was 5.6% in the authors' university from 1980 to 1981. The sex ratio of male to female was nearly equal. The mean age of NPC patients was 56.7 years. Sera from these 25 patients with nasopharyngeal carcinoma were collected at intervals of 3 to 8 months over a 2-year period, and were examined for their spectra and titers of antibodies of EBV-related antigens. They were titrated for IgG, IgA and IgM antibodies to EB viral capsid antigen (VCA), for IgG and IgA antibodies to early antigen-DR component (EA) and for antibodies to EBV-associated nuclear antigen (EBNA). All of these patients were primarily treated with radiation, while a few who did not respond to this therapy were subsequently treated with surgery or chemotherapy. EBV antibodies of VCA-IgG, -IgA, EA(DR)-IgG, and -IgA and EBNA were elevated in 73% and 90% of the nonkeratinizing and undifferentiated NPC patients, respectively. The VCA-IgM was elevated in almost none of the cases. In contrast to this, these values were all in a normal range in the NPC patients with keratinizing squamous cell carcinoma and malignant lymphoma. Also 9% and 10% of nonkeratinizing and undifferentiated carcinomas showed the normal ranges of EBV antibodies, possibly indicating a nonassociation with EBV. When NPC disappeared with radiation therapy, EBV antibodies became normal for 6-18 months. However, those whose NPC did not respond to the combined therapy with radiation, surgery and chemotherapy maintained high titers of EBV antibodies. The prognosis was the poorest in the patients with undifferentiated carcinoma, 40% of whom died within 4 years after diagnosis.

Adult↗
PubMed1984

Sera from adult T-cell leukemia patients react with envelope and core polypeptides of adult T-cell leukemia virus.

Sera from five Japanese patients with adult T-cell leukemia (ATL) showed in the immunofluorescence test for ATL-associated antigen (ATLA) titers ranging from 320 to 1280. Control sera from three healthy adults were negative. These eight sera were used to immunoprecipitate radiolabeled polypeptides from three cell lines infected with adult T-cell leukemia virus (ATLV) and two noninfected human cell lines. Cells were labeled either metabolically with [35S]cysteine, [35S]methionine, or [3H]glucosamine, or chemically with 125-iodine. Immunoprecipitates from cells, virus, and concanavalin A-enriched supernatants were analyzed by polyacrylamide gel electrophoresis. Cells producing ATLV contain gp68, the putative precursor to ATLV envelope polypeptides, gp46, and possibly p15, in addition to several nonglycosylated polypeptides between 40 to 70 kDa. Gp46 is shed into the culture medium and appears to be loosely attached to the viral and cellular surface. After purification on density gradients viral particles contain immunoreactive p24, p19, p15, and small amounts of gp46. Kinetics of synthesis, distribution, size, biochemical characteristics, and immunoreactivity of these polypeptides strongly suggest that most of them are structural components of ATLV or their precursors. Apparently, the intracellular ATLA complex predominantly represents precursors of viral structural polypeptides and gp46 is the viral envelope glycopolypeptide.

Antibodies, Viral↗
PubMed1984

Experimental infection of cynomolgus monkeys with a human retrovirus, adult T-cell leukemia virus.

The experimental infection of six cynomolgus monkeys with adult T-cell leukemia virus (ATLV) was attempted. Three animals were inoculated with living MT-2 cells and three with cell-free ATLV. All animals developed an antibody response to virus-specific glycopolypeptides and viral core polypeptides. ATLV-specific antigens appeared in peripheral lymphocytes from all six animals. Virus expression persisted in all animals. Up to 40 weeks after inoculation no animal developed any symptom of leukemia.

Animals↗
PubMed1984

Expression of HTLV-specific polypeptides in various human T-cell lines.

The adult T-cell leukemia (ATL)-associated antigen complex (ATLA) was first discovered with indirect immunofluorescence by Hinuma et al. (1981). Biochemical analysis with MT-2 cells revealed that ATLA consisted mainly of human T-cell leukemia virus (HTLV) structural polypeptides and their precursors (Yamamoto and Hinuma 1982a; Schneider et al. 1984). In this study, we have investigated the molecular nature of the ATLA antigen complex in various HTLV-positive human cell lines established by different methods including independently established HTLV-infected HUT 102 cells. We found that HTLVs infecting these cell lines have similar core polypeptides, p24 and p19, as well as an envelope glycopolypeptide, gp46, in all these cells. The intracellular gp61 and p53 appear to be precursors of the viral envelope and core polypeptides, respectively. Interestingly, MT-2 and MT-2 related T-cell lines contain two different species of envelope proteins, gp68 and gp61, whereas cell lines not related to MT-2 express only gp61.

Antigens, Viral↗
PubMed1984

Binding of adult T-cell leukemia virus to various hematopoietic cells.

A newly found human retrovirus, adult T-cell leukemia virus (ATLV) was shown by means of membrane immunofluorescence to bind to various hematopoietic cells including T-, B- and non-T, non-B-cell lines. Partially purified viral gp46 from culture fluids of ATL virus producer lines also bound efficiently to an ATLV-negative T-cell line, CCRF-CEM cells. When the viruses were pre-incubated with anti-ATLV-positive human sera, ATLV binding to the cells was clearly inhibited but not by pre-incubation with anti-ATLV-negative sera. These data suggest that: (1) ATLV binds not only to T-cells but also to multiple types of cells of hematopoietic origin; (2) anti-ATLV antibody-positive human sera have the blocking antibody for the binding of ATLV to lymphoid cells.

Antigens, Neoplasm↗
PubMed1984

In vitro infection of human B lymphocytes with adult T-cell leukemia virus.

Experimental transmission of adult T-cell leukemia (ATL) virus (ATLV) into human B lymphocytes was attempted. Cocultivation of B-cell rich fraction of peripheral blood from a healthy adult with X-ray irradiated ATLV producer MT-2 cells resulted in the establishment of OKA(B) cell line co-infected with both Epstein-Barr virus (EBV) and ATLV. OKA(B) cells and its subclones contained: (1) B cell markers exclusively; (2) both EBV-specific antigen, EBNA and ATLV-specific antigen, ATLA detected by immunofluorescence test; (3) ATLV-specific polypeptides, p24 and p19; (4) ATLV-specific mRNA in ATLA-positive clones; (5) ATLV and EBV particles detectable by electron microscopy. These data clearly show that human B lymphocytes are susceptible to ATLV infection.

Antigens, Viral↗
PubMed1984

Retrovirus-induced expression of interleukin 2 receptors on cells of human B-cell lineage.

We provide direct evidence for interleukin 2 receptor (IL2R) induction by human T-cell leukemia/lymphoma virus (HTLV) in human B-cell lines. A lymphoblastoid B-cell line (LCL-Ter) was established by Epstein-Barr virus-induced transformation of peripheral blood lymphocytes derived from a healthy HTLV carrier and cloned in vitro. HTLV gp21 and/or p19 antigens were detected in eight LCL-Ter clones, all of which also expressed the IL2R antigens Tac and Hiei (defined by monoclonal antibodies). However, five other LCL-Ter clones, which were negative for the HTLV antigens, did not express the IL2R antigens. Furthermore, when the IL2R-negative B-cell line LCL-Kan, derived form a normal donor, was cocultured with HTLV-producer cells, three HTLV-carrying clones were obtained and found to constitutively express IL2R. IL2R induced by HTLV on these B-cell lines bound recombinant interleukin 2 and were similar in apparent molecular mass (approximately equal to 60 kDa) to the IL2R of peripheral blood lymphocytes stimulated with phytohemagglutinin.

Antigens, Surface↗
PubMed1984

Characterization of human B-cell lines harbouring both adult T-cell leukaemia (ATL) virus and Epstein-Barr virus derived from ATL patients.

Human B-cell lines, designated ATLB cell lines, were spontaneously established from peripheral blood of adult T-cell leukaemia (ATL) patients. The cell lines consistently expressed ATL-associated antigen (ATLA) and Epstein-Barr virus-associated nuclear antigen (EBNA). A cloned ATLB line, designated ATLB 2, showed that both ATLA and EBNA antigens were present in the same B-cell clone. In this study, we have further characterized ATLV and EBV in the cloned ATLB 2 cell line by biochemical techniques. The ATLA antigens in these clones, initially shown by immunofluorescence, were studied by immunoprecipitation with human sera from ATL patients and the Western blotting technique using a mouse monoclonal antibody (GIN-14). We identified ATLV core polypeptides 24K and 19K in the ATLB cell extracts. Furthermore, total cellular DNA and poly(A) RNA from the ATLB cell lines were analysed for the presence of viral genomes with molecularly cloned DNA probes containing the ATLV and EBV sequence, respectively. The results showed that all ATLB 2 clones contained highly conserved ATLV proviral DNA irrespective of whether or not they expressed ATLA. They also contained several copies of EB virus DNA and DNA-DNA reassociation kinetics studies clearly showed that most of the EBV DNA sequences were present in ATLB cells. ATLV-related mRNA was detected in only ATLA-positive clones (ATLB 2-3 and 2-21) but not in a negative clone (ATLB 2-45).

B-Lymphocytes↗
PubMed1984

Precursor polypeptides of adult T-cell leukaemia virus: detection with antisera against isolated polypeptides gp68, p24 and p19.

Sera of individuals infected with adult T-cell leukaemia virus (ATLV) react predominantly with the polypeptides gp68, p24 and p19. These polypeptides were isolated from ATLV-infected MT-2 cells and virus. The radioiodinated polypeptides were used to quantify respective antibodies in individual ATLV carrier sera. Heteroantisera prepared in rabbits against isolated polypeptides facilitated studies on the biosynthesis of the core and envelope polypeptides of ATLV. Pulse-chase experiments revealed a polypeptide of mol. wt. 48 000 (48K) as the precursor to the core polypeptides p24 and p19. A 28K polypeptide related to p19 appeared to be an early side-product of the gag gene or a translate of a defective viral message. Antiserum to the putative env gene product gp68 recognized gp68, gp66 and small amounts of gp62. In tunicamycin-treated cells gp68, gp66 and gp62 were no longer synthesized, but a 54K polypeptide reacted with antiserum to gp68. Polypeptide p54 is structurally related to gp68 and therefore apparently represents the unglycosylated form of gp68. Moreover, the apparent mol. wt. of p54 and p48 agree with those predicted for respective env and gag precursors from the nucleotide sequence of an ATLV provirus.

Antibodies, Viral↗
PubMed1984

Human adult T-cell leukaemia virus is distinct from a similar isolate of Japanese monkeys.

We have compared the structural polypeptides of an adult T-cell leukaemia (ATL) virus (ATLV) isolate from a Japanese patient with ATL with those of a similar virus derived from a Japanese macaque monkey. Both are distinct but related entities. Their core polypeptides p19 could not be distinguished, but p24, another core polypeptide, and their envelope glycopolypeptides differ. The human virus directs the synthesis of a single intracellular glycopolypeptide, gp68, while the macaque virus specifies two such glycopolypeptides, gp57 and gp50. Furthermore, the glycopolypeptides of both viruses are serologically distinct. Thus, these viruses represent subtypes of the ATLV family and the macaque virus is apparently not involved in human ATL.

Animals↗