Biomedical subjects

M Hayami

Publications and source records attributed to M Hayami.

At least 127 records · Page 7Linked to original sources

PubMed1992

Biological characterization of human immunodeficiency virus type 1 and type 2 mutants in human peripheral blood mononuclear cells.

Mutants of human immunodeficiency virus type 1 (HIV-1) and type 2 (HIV-2), which have been shown to be infectious in established cell lines, were tested for ability to replicate and induce syncytium formation in human peripheral blood mononuclear cells (PBMC). The vpu mutant of HIV-1 showed depressed kinetics of replication in an established T cell line, as reported previously, but in PBMC, its replication was similar to that of the wild type virus. The vpx gene of HIV-2 was required for efficient virus propagation in PBMC, but not in an established T cell line, as previously reported. However, the growth rates of the vpx mutant in PBMC preparations from two individuals were different. The results of experiments on infection of PBMC with the vif and vpr mutants of HIV-1 and HIV-2 were essentially consistent with previous results of infection of established T cell lines. No negative effect of the nef gene products of HIV-1 and HIV-2 was observed. The abilities of the wild type virus and the mutants of HIV-1 to induce syncytium formation in both PBMC and established cell lines were similar. In contrast, neither the wild type nor any of the mutants of HIV-2 induced syncytium formation in PBMC. These results suggest that the functions of some genes can be detected only in mixed populations or primary cells such as PBMC. Studies on the roles of these genes in PBMC may provide a better understanding of their functions in vivo.

Cells, Cultured↗

PubMed1992

Expression of feline immunodeficiency virus gag gene in Escherichia coli.

The gag gene of a Japanese feline immunodeficiency virus (FIV) isolate, designated as FIV TM 2, was expressed in Escherichia coli as a fusion protein with TrpE. Using this expressed protein, an enzyme-linked immunosorbent assay was developed for detection of antibodies to FIV gag protein in feline sera. With serum samples from a cat experimentally infected with FIV, it was demonstrated that the period of seroconversion detected by this method corresponded to that by Western blotting.

Cloning, Molecular↗

PubMed1992

Functional classification of simian immunodeficiency virus isolated from a chimpanzee by transactivators.

In reporter-based transient expression systems, we characterized simian immunodeficiency virus from a chimpanzee (SIVCPZ), with special reference to the human immunodeficiency virus type 1 (HIV-1). SIVCPZ was not equally activated by tat and rev transactivators derived from representative primate lentiviruses. HIV-1 alone activated SIVCPZ to the full extent in both tat and rev assays. The tat and rev gene products of SIVCPZ, as well as those of HIV-1, efficiently transactivated the other viruses. These results indicate that SIVCPZ is identical to HIV-1 with regard to the compatibility of tat and rev gene activities among four subgroups of primate lentiviruses.

Amino Acid Sequence↗

PubMed1992

Detection of feline immunodeficiency proviral DNA in peripheral blood lymphocytes by the polymerase chain reaction.

Feline immunodeficiency virus (FIV) proviral DNA was detected by the polymerase chain reaction method (PCR). PCR products were detected by gel electrophoresis and ethidium bromide staining. The P-10, P-15 and P-24 regions of the gag gene of FIV were chosen as the target sequences for amplification, and three primer pairs were prepared. The PCR products subjected to amplification with each primer pair were found to possess sites of digestion by a restriction enzyme, as hypothesized. They did not react with feline leukemia virus (FeLV)-infected or feline syncytium-forming virus (FeSFV)-infected cell-derived DNA, and specifically amplified FIV-infected cell-derived DNA. FIV proviral DNA was detected by the PCR method with either primer pair (one-step amplification: single PCR) in DNA derived from peripheral blood lymphocytes (PBL) from 7 of 12 FIV antibody-positive cats. When PCR products in each of the 12 cats were subjected to a second amplification using the same primer pair (two-step amplification: double PCR), FIV proviral DNA was detected in all of the cats. When PBL samples collected from three cats that were negative and three that were positive in the single PCR were cultured for a few weeks in the presence of interleukin 2, FIV proviral DNA was detected in all six cats by the single PCR method. The results suggest that either the use of cultured PBL as the sample or the performance of the double PCR method enables simple and specific detection of FIV proviral DNA in PBL.

Animals↗

PubMed1992

Infection of macaque monkeys with a chimeric human and simian immunodeficiency virus.

Two macaque monkeys were inoculated with a chimeric human and simian immunodeficiency virus carrying the tat, rev, vpu and env genes of human immunodeficiency virus type 1. Infectious virus was recovered from one of the monkeys at 2 and 6 weeks post-infection. The hybrid nature of the isolated viruses was verified by Southern and Western blotting analyses. Both of the monkeys infected with the chimera elicited a humoral antibody response against the virus.

Animals↗

PubMed1991

Detection of HTLV-I antigen in peripheral and cord blood lymphocytes from carrier mothers.

Vertical transmission of human T-cell lymphotropic virus type-I (HTLV-I) from an antibody-positive carrier mother to her infant has been reported, but it is not clear when and by what route such transmission occurs. Peripheral blood and cord blood lymphocytes from 40 antibody-positive carrier mothers were cultured for 2 months and HTLV-I antigen was sought by indirect immunofluorescence. Viral antigen was detected in 28 (70%) samples of peripheral blood lymphocytes and in 2 of cord blood lymphocytes, both from mothers positive for antigen in peripheral blood. From these data it is estimated that transplacental infection occurs at a rate of about 7%.

Adult↗

PubMed1991

Compatibility of rev gene activity in the four groups of primate lentiviruses.

The compatibility of rev genes derived from various primate immunodeficiency viruses of all distinct subgroups identified was assessed in three experimental systems: complementation experiments between proviral rev and gag mutants, evaluation of the ability of the rev gene products to activate proviral reporters, and examination of the capacity of various viruses to augment marker gene expression in the infected reporter cell lines. In all systems, human immunodeficiency virus type 1 (HIV-1) rev was not substantially substituted or was extremely poorly substituted by the rev of the other viruses. The rev of simian immunodeficiency virus (SIV) from a mandrill could be exchanged by HIV-1 rev. In contrast, the rev gene products of all viruses efficiently activate HIV-2 and SIV from an African green monkey.

Base Sequence↗

PubMed1991

Functional analysis of long terminal repeats derived from four strains of simian immunodeficiency virus SIVAGM in relation to other primate lentiviruses.

The promoter activity of long terminal repeats (LTRs) of four strains of the simian immunodeficiency virus isolated from African green monkeys (SIVAGM) was compared with those of various LTRs derived from the other representative primate lentiviruses: human immunodeficiency virus type 1 (HIV-1), type 2 (HIV-2), SIV from a rhesus monkey (SIVMAC), and SIV from a mandrill (SIVMND). The expression of the LTRs was evaluated by monitoring chloramphenicol acetyltransferase production after transfection of reporter plasmid clones. In the absence of viral tat, all SIVAGM LTRs acted as much more efficient promoters than any of the other LTRs. When tat gene products were supplied in trans, LTRs of SIVAGM and SIVMND were activated inefficiently relative to high responder LTRs of HIV-2 and SIVMAC. The LTR of HIV-1 was highly activated by HIV-1 tat, but not so much by HIV-2, SIVAGM, and SIVMND tat.

Animals↗

PubMed1991

Protection of mice from a lethal coronavirus infection in the central nervous system by adoptive transfer of virus-specific T cell clones.

The protective effect of a mouse hepatitis virus type-4 (MHV-4)-specific CD8+ cytotoxic T cell clone and a CD4+ helper T cell clone was examined by the adoptive transfer into brains of mice lethally infected with MHV-4. Mice survived acute encephalitis if more than 5 x 10(5) cells of either type of the virus-specific T cell clones had been transferred into H-2-matched recipients by 1 day post-infection. Although the adoptive transfer of both types of the T cell clones suppressed viral growth and viral antigen-positive cells in the brains, a significant inhibition of virus replication by the cytotoxic T cell clone was detected prior to that induced by the helper T cell clone. Histologically, cell destruction was prominent in the brains of mice which received the cytotoxic T cell clone. These results demonstrate that both the CD8+ cytotoxic T cell and the CD4+ helper T cell can protect mice from a lethal MHV-4 infection in the central nervous system.

Animals↗

PubMed1991

Production and characterization of mouse monoclonal antibodies against the transmembrane protein of a human immunodeficiency virus type 2.

We established seven hybridoma clones producing monoclonal antibodies (MAbs) against the envelope transmembrane protein (TMP) of a Ghanian isolate of human immunodeficiency virus type 2 (HIV-2[GH-1]) from mice immunized with the detergent-disrupted purified whole virus. The MAbs were found to react with 35 kilodalton (kD) TMP of the HIV-2[GH-1] virus in a Western blot assay (WB). Two of these MAbs recognized 135 kD proteins in addition to TMP in the lysate of HIV-2[GH-1]-infected cells. Two other MAbs cross-reacted with viral components corresponding to TMPs of HIV-2ROD and SIVMAC isolates in a Western blot. Results of competitive binding assay suggest that there are at least three epitopes on a TMP molecule of the HIV-2[GH-1] isolate. The MAbs did not inhibit syncytium formation between HIV-2[GH-1]-infected MOLT-4 cells and MOLT-4 clone 8 cells, nor virus infection to MOLT-4 clone 8 cells.

Animals↗

PubMed1991

Molecular cloning of a novel isolate of feline immunodeficiency virus biologically and genetically different from the original U.S. isolate.

The Japanese isolate (TM1 strain) of feline immunodeficiency virus (FIV) which replicates in a feline CD4 (fCD4)-positive lymphoblastoid cell line (MYA-1 cells) was molecularly cloned from extrachromosomal closed circular DNA. The restriction map of the clone, termed pFTM 191 complete genome (CG), showed a considerable difference from that of the U.S. isolate (Petaluma strain) of FIV. The sequence homology in the long terminal repeat between the TM1 and Petaluma strain was 82%. The pFTM 191 CG was biologically active after transfection into Crandell feline kidney cells which were permissive for replication of FIV Petaluma. However, the progeny virions could not reinfect fCD4-negative Crandell feline kidney cells but could infect fCD4-positive MYA-1 cells. When a specific-pathogen-free cat was inoculated with the virus derived from the pFTM 191 CG, the cat seroconverted within 8 weeks postinoculation and FIV was reisolated at 4, 8, and 20 weeks postinoculation. These results indicate the infectivity of the pFTM 191 CG in vivo.

Animals↗

PubMed1991

Generation of a chimeric human and simian immunodeficiency virus infectious to monkey peripheral blood mononuclear cells.

We constructed five chimeric clones between human immunodeficiency virus type 1 (HIV-1) and simian immunodeficiency virus (SIVMAC) and four SIVMAC mutants by recombinant DNA techniques. Three chimeric clones and all mutants with an alteration in either the vif, vpx, vpr, or nef gene were infectious to human CD4-positive cell lines. The susceptibility of macaque monkey peripheral blood mononuclear cells (PBMC) to infection by these mutants and chimeras was examined in vitro. Macaque PBMC supported the replication of wild-type and vpx, vpr, and nef mutant SIVMAC strains. A chimera carrying the long terminal repeats (LTRs), gag, pol, vif, and vpx of SIVMAC and tat, rev, vpu, and env of HIV-1 was also replication competent in PBMC. In contrast, HIV-1, the vif mutant of SIVMAC, a chimera containing rev and env of SIVMAC, and a chimera containing vpx, vpr, tat, rev, and env of SIVMAC did not grow in PBMC. Western immunoblotting analysis of the replicating chimera in PBMC confirmed the hybrid nature of the virus. These data strongly suggested that the sequence important for macaque cell tropism lies within the LTR, gag, pol, and/or vif sequences of the SIVMAC genome.

Animals↗

PubMed1991

Chronic actinic dermatitis: a time course study of histopathological changes.

We report herein a case of chronic actinic dermatitis in which histopathological changes were observed on several occasions during the course of 2 years. The findings were quite different among stages of the eruption. At a half year after the appearance of the lesion, histological findings revealed eczematous changes. However, gradually a granulomatous change appeared with histiocytes and giant cells. Although the mechanism of histogenesis is not clear, the appearance of histiocytic giant cells seems to be unusual in chronic actinic dermatitis.

Aged↗

PubMed1990

Simian T cell leukemia virus type-1-specific killer T cells in naturally infected African green monkey carriers.

Studies were made on the cellular immunity of 13 African green monkeys (Cercopithecus aethiops) naturally infected with Simian T cell leukemia virus type 1 (STLV-1), closely related to human T cell leukemia virus type 1. They were classified into 3 groups: 1) progressed carrier, 2) carrier, and 3) normal control. This grouping was made according to their hematologic features, i.e., number of peripheral white blood cells, existence of blastoid cells, presence of STLV-1 Ag on PBL and anti-STLV-1 antibody titers. None of the STLV-1 carriers showed any clinical signs, but STLV-1-specific killer T cells was detected in these PBL without in vitro stimulation. In vitro studies on Ag stimulation showed that the STLV-1-specific killer T cells had been fully activated in vivo, and that no augmentation of in vitro stimulation with STLV-1 Ag was necessary, and that primary in vitro stimulation of normal control PBL was not sufficient to induce specific killer T cells. In addition NK cell activity in PBL of infected monkeys were significantly higher than those in the uninfected.

Animals↗

PubMed1990

Intracellular processing and immunogenicity of the envelope proteins of human T-cell leukemia virus type I that are expressed from recombinant vaccinia viruses.

Two types of recombinant vaccinia viruses (VVs) expressing the env gene of the human T-cell leukemia virus type I (HTLV-I) were reported previously. One recombinant VV, WR-proenv1, synthesized the authentic env protein. In the other recombinant VV, WR-env17, the env gene was inserted within the signal sequence of the VV hemagglutinin (HA) gene, so that the reading frame for the env gene was in phase with that for the HA gene. Comparative studies were performed on the mode of expression and processing of the env proteins in relation to their immunogenicity. In WR-env17-infected cells, translation was initiated exclusively from the initiation methionine of the HA to produce nascently the chimeric env protein, including the altered HA signal peptide. Both this altered HA signal peptide and the internalized env signal peptide functioned as insertion signals for the endoplasmic reticulum. Although about half of the nascent chimeric protein was cleaved at the carboxyl terminus of the internalized env signal peptide to produce the authentic env protein, the other half was cleaved at the carboxyl terminus of the altered HA signal peptide alone to synthesize the chimeric protein. These events led to a less efficient transport of the env protein produced by WR-env17 from the rough endoplasmic reticulum to the Golgi apparatus than that of the authentic env protein synthesized by WR-proenv1. The efficiency of the processing and transport of the env protein affected the immunogenicity of these two recombinant VVs.

Animals↗

PubMed1990

Identification and localization of vpr gene product of human immunodeficiency virus type 1.

The entire vpr gene of human immunodeficiency virus type 1 (HIV-1) was cloned into procaryotic and eucaryotic expression vectors. Production of authentic protein encoded by the gene in bacterial and mammalian cells was monitored by Western blotting using guinea pig antisera raised against an N-terminal 14-oligopeptide of the predicted vpr protein. A specific 12-kD protein was clearly detected with these antisera, but not with preimmune sera, in both cell systems, and this binding was blocked by the oligopeptide. These antisera also recognized a protein of the same size in several human T-cell lines infected with HIV-1. Western blotting analysis of subcellular fractions prepared from the cells producing wildtype vpr protein strongly suggested that the protein was membrane associated. A region within the vpr required for the stable expression of vpr product was also suggested by mutational analyses.

Amino Acid Sequence↗

PubMed1990

Molecular cloning of feline leukemia provirus genomes integrated in the feline large granular lymphoma cells.

Feline leukemia virus (FeLV) is a horizontally transmitted agent of the domestic cat which is known to be associated with wide spectrum of diseases of the hematopoietic system. In the present study, proviral DNAs of FeLV proviruses were examined in the tumor cells of natural killer cell lineage which is very rare in cats. In the chromosomal DNA of the tumor cells, 5 distinct bands corresponding to exogenous FeLV provirus genomes were detected by digestion with EcoRI which does not cut most FeLV isolates. Five clones of pLC1, pLC2, pLC3, pLC4, and pLC5 obtained from the 5 respective bands were analysed by restriction endonuclease mapping and Southern blot hybridization using gene-specific probes of FeLV. The results have clearly demonstrated that pLC4 and pLC5 contained large deletions in the pol and part of gag regions, while the full-length proviruses could be observed in pLC1 and pLC2. Furthermore, pLC3 contained part of a variant FeLV genome having an EcoRI site in its gag region. The molecular clones of defective and variant FeLV in this study may be useful for the further examination of tumorigenesis of large granular lymphoma in the cat.

Animals↗

PubMed1990

Evolutionary origin of human and simian immunodeficiency viruses.

From what viruses the human immunodeficiency viruses (HIVs) originated is an extremely controversial question. To address this question, we have analyzed nucleotide sequences of simian immunodeficiency viruses (SIVs) and HIVs by using the techniques for understanding molecular evolution. In particular, we compared the nucleotide sequences of whole genomes, gene region by gene region, between a given pair of viruses, including four types of SIVs--isolated from mandrills (Papio sphinx), African green monkeys (Cercopithecus aethiops), sooty mangabeys (Cercocebus atys), and rhesus macaques (Macaca mulatta)--as well as HIVs. Phylogenetic trees for all gene regions examined showed that the present HIVs may have emerged as different variants of SIVs of Old World monkeys, possibly from recombination between viruses related to SIVs.

Biological Evolution↗