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S Kyuwa

Publications and source records attributed to S Kyuwa.

At least 37 records · Page 2Linked to original sources
PubMed1995

Transcription and translation of proinflammatory cytokines following JHMV infection.

Infection with JHMV results in the transcriptional activation of two host cell genes encoding proinflammatory cytokines, tumor necrosis factor (TNF)-alpha and interleukin (IL)-1 beta. Analysis of irradiated mice showed that IL-1 beta mRNA accumulation in the central nervous system was predominantly derived from the mononuclear infiltrate. By contrast, accumulation of TNF-alpha mRNA was unaffected by immunosuppression, suggesting that resident cells were the source of this cytokine. Infected mice were treated with anti-TNF antibody to determine if TNF-alpha contributed to either the encephalomyelitis or demyelination associated with JHMV infection. Surprisingly, neither the cellular infiltrate nor demyelination were affected. In vitro analysis showed that IL-1 beta but not TNF was secreted from JHMV infected macrophages. The absence of TNF secretion is due to a block in translation of the TNF mRNA which accumulates during infection.

Animals↗
PubMed1995

Induction and natural occurrence of serum nucleosomal DNA in autoimmune MRL/lpr/lpr mice: its relation to apoptosis in the thymus.

Long-term administration of low doses (5 micrograms) of recombinant nucleobindin (rNuc), which is an MRL/lpr/lpr (MRL/l) mouse-derived DNA-binding protein, induces autoimmunity in both young lupus-prone MRL/+/+ (MRL/n) and normal BALB/c mice. In relation to this autoimmunity, using agarose gel electrophoresis we found an approximately 160 bp mono-nucleosomal DNA (nsDNA) in the sera of 6-week-old normal mice 15 h after i.p. injection of 50 micrograms rNuc. Co-injection of rNuc (50 micrograms) and anti-CD3 monoclonal antibody (mAb, 50 micrograms) further accelerated the appearance of nsDNA in the serum together with DNA fragmentation (apoptosis) in the thymus, which had not been so clearly induced by either double amounts of rNuc or anti-CD3 mAb alone. Acceleration of the appearance of nsDNA in the serum by co-injection was also found in age-matched MRL/n and MRL/l mice, indicating the close association of apoptosis in the thymus with the appearance of nsDNA in the serum. Furthermore, we have detected naturally occurring tri-, di- and mono-nsDNAs from immune complexes (IC) of the sera of 20 approximately 22-week-old MRL/l mice, which indicates that apoptosis in the lymphoid tissues, including the thymus, is the source of serum nsDNA that may trigger or continue production of anti-nuclear antibodies. Evidence that clearance of nsDNA from the circulation is retarded in the presence of rNuc in BALB/c mice may give rationale to the induction of autoimmunity in normal mice by long-term administration of even low doses (5 micrograms) of rNuc after all.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Coronavirus translational regulation: leader affects mRNA efficiency.

Cells infected with the murine coronavirus, mouse hepatitis virus (MHV), show decreased host protein synthesis concomitant with an increase in viral protein synthesis. We examined the in vitro translation property of the conserved MHV 5'-leader RNA sequence by constructing chimeric mRNAs in which the 72-nt 5'-leader of M protein mRNA (A59 strain) was positioned upstream of the human alpha-globin coding region in a T7 expression vector. Synthetic 5'-capped transcripts of these mRNA constructs were translated in cell-free extracts prepared from uninfected and MHV-infected murine DBT cells. Nonviral mRNAs translated readily in both uninfected and infected cell-free extracts. By contrast, replacement of the human alpha-globin 5'-untranslated region (UR) with the MHV 5'-leader increased translation ca. three- to fourfold in cell-free extracts from MHV-infected cells versus translation in extracts from uninfected cells. Chimeric globin mRNA containing the reverse complementary sequence of the viral leader RNA in the 5'-UR showed no such increase in translation, indicating sequence specificity for the effect. A 13-nt region (-UCUAAUCCAAACA-) immediately proximal to the start codon was found to be important for the increased translation of the MHV leader-containing mRNAs. These data indicate that the apparent down-regulation of host translation is not primarily due to an inhibition of host translation but also involves a significant stimulation of viral translation in cis by a structural feature of the MHV 5'-leader RNA sequence in conjunction with a virus-specified or virus-induced factor.

Animals↗
PubMed1994

Modulation of cellular macromolecular synthesis by coronavirus: implication for pathogenesis.

Infection with the murine coronavirus strain JHM decreases cell surface expression of major histocompatibility complex class I antigens. Northern blots showed that JHM virus infection rapidly reduced the level of actin mRNA, whereas the levels of major histocompatibility complex class I and tubulin mRNAs were reduced only slightly. By contrast, the mRNA levels of interleukin 1 beta, colony-stimulating factor 1 receptor, and tumor necrosis factor alpha increased following infection.

Actins↗
PubMed1994

Apoptosis induced in mouse hepatitis virus-infected cells by a virus-specific CD8+ cytotoxic T-lymphocyte clone.

Morphological changes of mouse hepatitis virus-infected J774.1 cells cocultured with cloned mouse hepatitis virus-specific CD8+ cytotoxic T lymphocytes were examined by electron microscopy. Condensation and margination of chromatin, cellular shrinkage with severe vacuolar degeneration, and blebbing were observed. In addition, fragmentation of cellular DNA was observed, and a decrease in virus titer was accompanied by those changes. These findings show that the cloned cytotoxic T lymphocytes induce in the target cell an internal degradation program termed apoptosis, which results in virus clearance.

Animals↗
PubMed1993

Effects of mouse hepatitis virus infection on host cell metabolism.

A time dependent decrease in cell surface expression of major histocompatibility complex (MHC) class 1 proteins was found during JHMV infection of the mouse macrophage J774.1 cells line by radioimmunoassay. MHC class I, actin and CSF-1 receptor mRNA levels were also found to decrease during infection. Surprisingly, not all host cell mRNA were similarly affected, suggesting that the apparent MHV-induced translational shut off of host cell protein synthesis during infection was specific for only some host cell mRNAs. Interestingly, two mRNAs found to be refractory to JHMV infection encode monokines, suggesting a role in pathogenesis. To understand the mechanism(s) of this preferential mRNA stability and the apparent shut off of host cell mRNA, translation lysates were prepared from infected and uninfected cells. Translation of host mRNAs in these extracts showed no apparent loss of translational ability in the infected cells vs. the uninfected cells; however, viral mRNAs were preferentially translated in the lysates from the infected cells. Chimeric mRNAs containing the MHV leader upstream of a globin reporter gene showed that preferential translation was a property of the MHV leader RNA. Deletional analysis showed that the sequences responsible for this cis translational augmentation are in a 12 nucleotide (nt) tract at the 3' end of the leader. The previously reported interaction of the nucleocapsid protein with these nts suggest that it may play a role in translational augmentation of MHV mRNAs.

Animals↗
PubMed1993

JHM virus-specific cytotoxic T cells derived from the central nervous system.

Spleen cells cultured from Balb/c mice immunized with the JHM strain of mouse hepatitis virus (JHMV) have CD8+ cytotoxic T cells (CTL) specific for both the S and N proteins, but not the M or HE proteins. T cell lines were established from the brains of Balb/c mice infected with JHMV. The majority of the lines (20 of 22) were specific for JHMV. Analysis of the viral structural proteins which served as target structures indicate that most (15 of 20) were specific for the N protein. One line was specific for the S protein and four lines were specific for JHMV but the protein recognized could not be determined. These data suggest that early during infection there is a preferential recruitment of N protein specific CTL into the CNS of infected mice.

Animals↗
PubMed1993

Characterization of mouse hepatitis virus-specific cytotoxic T cells derived from the central nervous system of mice infected with the JHM strain.

The cytotoxic T lymphocyte (CTL) activity of spleen cells from BALB/c (H-2d) mice immunized with the neurotropic JHM strain of mouse hepatitis virus (JHMV) was stimulated in vitro for 7 days. CTL were tested for recognition of target cells infected with either JHMV or vaccinia virus recombinants expressing the four virus structural proteins. Only target cells infected with either JHMV or the vaccinia virus recombinant expressing the JHMV nucleocapsid protein were recognized. Cytotoxic T cell lines were established by limiting dilution from the brains of mice undergoing acute demyelinating encephalomyelitis after infection with JHMV. Twenty of the 22 lines recognized JHMV-infected but not uninfected syngeneic target cells, indicating that they are specific for JHMV. All T-cell lines except one were CD8+. The specificity of the CTL lines was examined by using target cells infected with vaccinia virus recombinants expressing the JHMV nucleocapsid, spike, membrane, and hemagglutinin-esterase structural proteins. Seventeen lines recognized target cells expressing the nucleocapsid protein. Three of the JHMV-specific T-cell lines were unable to recognize target cells expressing any of the JHMV structural proteins, indicating that they are specific for an epitope of a nonstructural protein(s) of JHMV. These data indicate that the nucleocapsid protein induces an immunodominant CTL response. However, no CTL activity specific for the nucleocapsid protein could be detected in either the spleens or cervical lymph nodes of mice 4, 5, 6, or 7 days after intracranial infection, suggesting that the CTL response to JHMV infection within the central nervous system may be induced or expanded locally.

Amino Acid Sequence↗
PubMed1992

Mouse hepatitis virus nucleocapsid protein-specific cytotoxic T lymphocytes are Ld restricted and specific for the carboxy terminus.

Infection of mice with the JHM strain of mouse hepatitis virus (MHV) results in an acute encephalomyelitis associated with primary demyelination of the central nervous system. Efforts at understanding the components of the immune response in the development of chronic MHV-induced demyelination have implicated the antibody response and both the CD4+ and CD8+ T cell responses. In this report, we demonstrate that Balb/c (H-2d) mice immunized with the JHM (JHMV) strain of MHV develop a CD8+ cytotoxic T lymphocyte (CTL) response. One population of these virus-specific CTL recognize the nucleocapsid (N) protein. Recombinant vaccinia viruses expressing either the entire N protein or carboxy-terminal deletions were used to determine the number and location of the epitope(s) recognized. The CTLs were found to recognize a peptide contained within the carboxy-terminal 149 amino acids of the N protein. Analysis of infected cell lines expressing transfected major histocompatibility genes demonstrated that the anti-N protein CTLs were restricted exclusively to the Ld molecule. These data provide the first definition of a MHV-specific CTL response directed to a viral protein and suggest that the anti-N protein CTL response is one potential mechanism used by the host to clear JHMV from the central nervous system.

Animals↗
PubMed1992

Acute and late disease induced by murine coronavirus, strain JHM, in a series of recombinant inbred strains between BALB/cHeA and STS/A mice.

To examine the genetic control of acute and late disease induced by a murine coronavirus, strain JHM (JHMV), BALB/cHeA, STS/A, F1 hybrids and 13 recombinant inbred (RI) strains between BALB/cHeA and STS/A mouse strains were inoculated intracerebrally with 100 pfu of JHMV. All the BALB/cHeA mice died within 2 weeks from acute encephalitis. In contrast, STS/A mice were shown to be partially resistant, with a mortality rate of 30%, longer survival times and lower rates of viral production. The mortality rates, survival times and viral titers of F1 hybrids and the RI strains varied, suggesting involvement of multiple genes. STS/A, F1 hybrid and RI mice surviving the acute infection occasionally developed severe paraparesis about 1 month post-infection. In these mice, vacuolar degeneration, astrocytosis, the absence of perivascular cuffing and minimal demyelination were found in the central nervous system. No infectious virus could be recovered from these mice. Although the paralysis of delayed onset was limited to STS/A, F1 hybrid and eight of the 13 RI strains, the incidence varied significantly among the RI strains. These results may suggest that JHMV-induced late disease is also under multifactorial control. The pathogenesis of JHMV infection is discussed.

Acute Disease↗
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

Induction of self-reactive T cells after murine coronavirus infection.

We studied the mechanism of in vitro spontaneous lymphokine production by spleen cells from mice injected intraperitoneally with murine coronavirus stain JHM 1 month after infection, when infectious virus had already been cleared from the spleens. Removal of either CD4+ T cells or Ia+ antigen-presenting cells (APC) from the spleen cells abrogated interleukin-2 (IL-2) production. Addition of anti-CD4 or anti-Iad monoclonal antibodies to the culture suppressed IL-2 production. These results suggest that the response involved typical receptor-mediated activation of T cells. Surprisingly, reciprocal mixing experiments with a coculture of T cells from infected mice and APC from either infected or naive mice resulted in the production of IL-2. The absence of viral antigens in spleen cells 1 month after infection, as indicated by their inability to induce the proliferation of T-cell clones specific for the viral antigens, suggest that the T cells from mice 1 month after infection were not responding to the viral antigens. The inoculum components other than the virus did not induce this immune response. We also found that the frequency of self-reactive but not alloreactive IL-2-producing T cells in the spleens of infected mice was 3- to 10-fold higher than that in naive mice. These findings suggest that an increased frequency of self-reactive T cells which secrete IL-2 occurs following murine coronavirus infection. This may have important implications in the development of autoimmunelike phenomena following murine coronavirus infection.

Animals↗
PubMed1989

Immunopathology of chronic progressive hepatitis in nude mice infected with low-virulent mouse hepatitis virus.

Progressive hepatitis in athymic nude (nu/nu) mice due to a low-virulent mouse hepatitis virus, MHV-2 cc, was examined for involvement of immunocytes and serum antibodies. At 3 to 6 weeks postinoculation (p.i.) a considerable number of Mac 1- and asialo GM1-positive cells were accumulated in the affected liver and spleen. There were also some Thy-1-positive cells. Later than 2 weeks p.i., serum IgG and IgM antibodies were detected in parallel with virus-neutralizing activity, while the IgG levels were lower than those of infected euthymic (nu/+) littermates. By transfer of the infected nu/nu mouse serum, the recipient euthymic mice acquired resistance to lethal challenge infection with a virulent virus, MHV-2.

Animals↗
PubMed1989

T-cell-mediated clearance of mouse hepatitis virus strain JHM from the central nervous system.

Clearance of the neurotropic JHM strain of mouse hepatitis virus from the central nervous system was examined by the transfer of spleen cells from immunized donors. A T cell with the surface phenotype of Thy1.2+ CD4+ CD8- asialo-GM1+ Mac-1- was found to be necessary for viral clearance. The surface phenotype and adherence to nylon wool suggest that these cells are activated helper-inducer T cells. Adoptive transfer to congenic histocompatibility strains demonstrated the necessity for compatibility at the D locus of the major histocompatibility complex. The expression of the CD4 surface marker and the requirement for major histocompatibility complex class I were further studied by the transfer of cells to recipients treated with anti-CD4 or anti-CD8 monoclonal antibodies. Treatment of recipients with either the anti-CD8 or the anti-CD4 antibodies inhibited virus clearance from the central nervous system. This suggests that the CD4+ cell acts as a helper and that virus is cleared from the central nervous system. This suggests that the CD4+ cell acts as a helper and that virus is cleared from the central nervous system by CD8+ cells that recognize viral antigen in the context of the H-2Db gene product.

Animals↗