Sequence analysis of the small RNA segment of guinea pig-passaged Pichinde virus variants.
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The established animal model for Lassa fever is based on the new world arenavirus Pichinde (PIC). Natural isolates of PIC virus are attenuated in guinea pigs, but serial guinea pig passage renders them extremely virulent in that host. We have compared the nucleotide sequences of the small RNA segments of two attenuated, low-passage variants of the PIC virus Munchique strain (CoAn 4763) and two virulent, high-passage derivatives. Missense mutations in the glycoprotein precursor (GPC) gene at codons GPC-119, GPC-140, and GPC-164 and the nucleoprotein gene (NP) codons NP-35 and NP-374 were most closely associated with virulence. Codon GPC-140 is predicted to represent a region of peak hydrophilicity of the glycoprotein 1 (GP1); it is conceivable that mutations at this site could influence virulence by altering B cell epitopes or virus attachment protein conformation. (+info)
Attrition of T cell memory: selective loss of LCMV epitope-specific memory CD8 T cells following infections with heterologous viruses.
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Using a variety of techniques, including limiting dilution assays (LDA), intracellular IFNgamma assays, and Db-IgG1 MHC dimer staining to measure viral peptide-specific T cell number and function, we show here that heterologous virus infections quantitatively delete and qualitatively alter the memory pool of T cells specific to a previously encountered virus. We also show that a prior history of a virus infection can alter the hierarchy of the immunodominant peptide response to a second virus and that virus infections selectively reactivate memory T cells with distinct specificities to earlier viruses. These results are consistent with a model for the immune system that accommodates memory T cell populations for multiple pathogens over the course of a lifetime. (+info)
Virus-induced abrogation of transplantation tolerance induced by donor-specific transfusion and anti-CD154 antibody.
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Treatment with a 2-week course of anti-CD154 antibody and a single transfusion of donor leukocytes (a donor-specific transfusion or DST) permits skin allografts to survive for >100 days in thymectomized mice. As clinical trials of this methodology in humans are contemplated, concern has been expressed that viral infection of graft recipients may disrupt tolerance to the allograft. We report that acute infection with lymphocytic choriomeningitis virus (LCMV) induced allograft rejection in mice treated with DST and anti-CD154 antibody if inoculated shortly after transplantation. Isografts resisted LCMV-induced rejection, and the interferon-inducing agent polyinosinic:polycytidylic acid did not induce allograft rejection, suggesting that the effect of LCMV is not simply a consequence of nonspecific inflammation. Administration of anti-CD8 antibody to engrafted mice delayed LCMV-induced allograft rejection. Pichinde virus also induced acute allograft rejection, but murine cytomegalovirus and vaccinia virus (VV) did not. Injection of LCMV approximately 50 days after tolerance induction and transplantation had minimal effect on subsequent allograft survival. Treatment with DST and anti-CD154 antibody did not interfere with clearance of LCMV, but a normally nonlethal high dose of VV during tolerance induction and transplantation killed graft recipients. We conclude that DST and anti-CD154 antibody induce a tolerant state that can be broken shortly after transplantation by certain viral infections. Clinical application of transplantation tolerance protocols may require patient isolation to facilitate the procedure and to protect recipients. (+info)
Alterations in NF-kappaB and RBP-Jkappa by arenavirus infection of macrophages in vitro and in vivo.
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Pichinde virus is an arenavirus that infects guinea pigs and serves as an animal model for human Lassa fever. An attenuated Pichinde virus variant (P2) and a virulent variant (P18) are being used to delineate pathogenic mechanisms that culminate in shock. In guinea pigs, the infection has been shown to begin in peritoneal macrophages following intraperitoneal inoculation and then spreads to the spleen and other reticuloendothelial organs. We show here that infection of the murine monocytic cell line P388D1 with either Pichinde virus variant resulted in the induction of inflammatory cytokines and effectors, including interleukin-6 and tumor necrosis factor alpha. Since these genes are regulated in part by the cellular transcription factors NF-kappaB and RBP-Jkappa, we compared the activities of NF-kappaB and RBP-Jkappa in P388D1 cells following infection with Pichinde virus. The attenuated P2 virus inhibited NF-kappaB activation and caused a shift in the size of the RBP-Jkappa complex. The virulent P18 virus showed less inhibition of NF-kappaB and failed to alter the size of the RBP-Jkappa complex. Peritoneal cells from P2-infected guinea pigs showed induction of NF-kappaB RelA/p50 heterodimer and p50/p50 homodimer and manifested an increase in the size of RBP-Jkappa. By contrast, P18 induced large amounts of the NF-kappaB p50/p50 dimer but failed to induce RelA/p50 or to cause an increase in the RBP-Jkappa size. Taken together, these changes suggest that the attenuated viral strain induces an "activation" of macrophages, while the virulent form of the virus does not. (+info)
Reassortant analysis of guinea pig virulence of pichinde virus variants.
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The new world arenavirus Pichinde (PIC) is the basis of an accepted small animal model for human Lassa fever. PIC (Munchique strain) variant P2 is attenuated in guinea pigs, whereas variant P18 is extremely virulent. Previous sequence analysis of the S segments of these two viruses indicated a small number of possible virulence markers in the glycoprotein precursor (GPC) and nucleoprotein (NP) genes. In order to determine the role of these S segment genes in guinea pig virulence in this system, we have generated reassortant viruses. When tested in outbred guinea pigs, the reassortant containing the S segment from the virulent parent P18 (S18L2) caused significantly higher morbidity than the reciprocal reassortant. This increased morbidity was associated with higher viral titers in serum and spleen. However, the S18L2 reassortant was not as fully virulent in this system as the P18 parent, indicating a role for L segment genes in virulence. (+info)
Dynamics of memory T cell proliferation under conditions of heterologous immunity and bystander stimulation.
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By examining adoptively transferred CSFE-labeled lymphocytic choriomeningitis virus (LCMV)-immune donor T cells in Thy-1 congenic hosts inoculated with viruses or with the cytokine inducer poly(I:C), strikingly different responses of bona fide memory T cells were found in response to different stimuli. Poly(I:C) (cytokine) stimulation caused a limited synchronized division of memory CD8 T cells specific to each of five LCMV epitopes, with no increase and sometimes a loss in number, and no change in their epitope hierarchy. Homologous LCMV infection caused more than seven divisions of T cells specific for each epitope, with dramatic increases in number and minor changes in hierarchy. Infections with the heterologous viruses Pichinde and vaccinia (VV) caused more than seven divisions and increases in number of T cells specific to some putatively cross-reactive but not other epitopes and resulted in substantial changes in the hierarchy of the LCMV-specific T cells. Hence, there can be memory T cell division without proliferation (i.e., increase in cell number) in the absence of Ag and division with proliferation in the presence of Ag from homologous or heterologous viruses. Heterologous protective immunity between viruses is not necessarily reciprocal, given that LCMV protects against VV but VV does not protect against LCMV. VV elicited proliferation of LCMV-induced CD8 and CD4 T cells, whereas LCMV did not elicit proliferation of VV-induced T cells. Thus, depending on the pathogen and the sequence of infection, a heterologous agent may selectively stimulate the memory pool in patterns consistent with heterologous immunity. (+info)
Direct visualization of cross-reactive effector and memory allo-specific CD8 T cells generated in response to viral infections.
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CD8 T cell cross-reactivity between heterologous viruses has been shown to provide protective immunity, induce immunopathology, influence the immunodominance of epitope-specific T cell responses, and shape the overall memory population. Virus infections also induce cross-reactive allo-specific CTL responses. In this study, we quantified the allo-specific CD8 T cells elicited by infection of C57BL/6 (B6) mice with lymphocytic choriomeningitis virus (LCMV). Cross-reactive LCMV-specific CD8 T cells were directly visualized using LCMV peptide-charged MHC tetramers to costain T cells that were stimulated to produce intracellular IFN-gamma in response to allogeneic target cells. The cross-reactivity between T cells specific for LCMV and allogeneic Ags was broad-based, in that it involved multiple LCMV-derived peptides, but there were distinctive patterns of reactivity against allogeneic cells with different haplotypes. Experiments indicated that this cross-reactivity was not due to the expression of two TCR per cell, and that the patterns of allo-reactivity changed during sequential infection with heterologous viruses. The allo-specific CD8 T cells generated by LCMV infection were maintained at relatively high frequencies in the memory pool, indicating that memory allo-specific CD8 T cell populations can arise as a consequence of viral infections. Mice previously infected with LCMV and harboring allo-specific memory T cells were refractory to the induction of tolerance to allogeneic skin grafts. (+info)
Attrition of virus-specific memory CD8+ T cells during reconstitution of lymphopenic environments.
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Viruses can cause a severe lymphopenia early in infection and a subsequent, lasting loss of pre-existing CD8(+) memory T cells. We therefore questioned how well virus Ag-specific memory CD8(+) T cells could reconstitute mice rendered lymphopenic as a consequence of genetics, irradiation, or viral or poly(I:C)-induced cytokines. In each case, reconstitution of the CD8(+) compartment was associated with limited division of virus-specific memory T cells and a reduction in their proportion. This indicates that foreign Ag-experienced CD44(high)CD8(+) memory T cells may respond differently to homeostatic signals than other CD44(high)CD8(+) cells, and that events inducing lymphopenia may lead to a permanent reduction in T cell memory. (+info)