Biomedical subjects

H Hengartner

Publications and source records attributed to H Hengartner.

At least 163 records · Page 9Linked to original sources

PubMed1994

Immunization with recombinant protein: conditions for cytotoxic T cell and/or antibody induction.

Safe vaccines should optimally induce both cell-mediated and humoral immunity. Recently, it has been shown that protective cytotoxic T cells (CTLs) can be induced not only with live vaccines, but also with recombinant viral proteins. This report shows in C57BL/6 (H-2b) mice that the recombinant nucleoprotein (N) of vesicular stomatitis virus (VSV) induced protective CTLs but no neutralizing antibodies in mice, whereas the recombinant glycoprotein (G) of VSV alone induced neutralizing antibodies but no CTLs. If the N and G of VSV were coinjected, both CTLs and a long-lasting neutralizing IgG response was measurable, demonstrating that mixed vaccines can be used to induce protective CTLs and antibodies with an efficiency comparable to live virus. In an attempt to define optimal conditions for CTL priming, the intravenous, intraperitoneal and subcutaneous route of injection were compared. Intravenous injection of recombinant VSV-N induced up to 30 times higher responses than the latter two routes. Finally, we tried to define conditions inducing only CTLs and no antibodies binding to the native protein form, or vice versa, only antibodies and no CTLs. Intravenous injection of boiled VSV-N induced a CTL response but no antibodies specific for the native VSV-N, whereas VSV-N injected subcutaneously in incomplete Freund's adjuvant induced high amounts of anti-VSV-N antibodies but virtually no CTLs. The conditions defined here permit vaccines to be designed which would function along selected and defined immunological effector pathways.

Animals↗

PubMed1994

Immunogenicity of a viral model vaccine after different inactivation procedures.

Various strategies for the production of safe vaccines have been used. This study compared three different inactivation procedures, i.e. treatment with formaldehyde, beta-propiolactone or UV-light using vesicular stomatitis virus (VSV) as a model antigen. All three inactivation procedures drastically impaired induction of neutralizing IgG responses, confirming previous observations [Bachmann et al. (1993) J Virol 67:3917-3922]. This reduction could be overcome using higher doses for all three preparations. Both formaldehyde and beta-propiolactone completely abrogated the induction of VSV-specific cytotoxic T cells (CTLs), whereas UV-inactivated virus was able to induce significant and long-lasting CTL responses. These results may be of practical relevance since induction of neutralizing antibodies alone is often not sufficient for protection and sometimes may even enhance immunopathological responses of vaccinees.

Animals↗

PubMed1994

T-cell-mediated immunopathology versus direct cytolysis by virus: implications for HIV and AIDS.

It has been much debated as to whether CD4+ T-cell depletion and the pathogenesis of AIDS is the result of direct cytolytic effects of human immunodeficiency virus (HIV), T-cell apoptosis by nonspecific activation, dysregulation of cytokine production, or autoimmunity. In this context, Rolf Zinkernagel and Hans Hengartner discuss data from model infections with non-cytopathic viruses. They suggest that HIV may cause immunosuppression, not by direct cytolytic effects, but rather by a conventional virus-specific cytotoxic T-cell-mediated immunopathology.

Acquired Immunodeficiency Syndrome↗

PubMed1994

Localization of T helper cell epitopes in the vesicular stomatitis virus: the nucleoprotein is responsible for serotype cross-reactive T help.

The glycoprotein (G) of vesicular stomatitis virus (VSV) is known to contain the biologically relevant sites for virus-neutralizing antibodies as well as T helper (Th) cell epitopes. The capacity of other VSV proteins to elicit potent Th cell responses has not yet been investigated. Additionally, a short-lived cross-reactivity between the two serologically distinct VSV serotypes Indiana (IND) and New Jersey (NJ) on the T helper cell level has been reported. Here we address the question of whether the VSV nucleoprotein (N) or matrix protein (M) can elicit T help to VSV-G-specific B cells and which of the VSV proteins contains the elements responsible for the IND/NJ cross-reactivity. The N, G, and M of the VSV Indiana serotype produced in a recombinant baculovirus system were assayed for the ability to activate VSV-specific Th cells to induce immunoglobulin class switch of neutralizing antibody responses in vivo in C57BL/6 (H-2b) mice. All three VSV-IND proteins helped in the production of neutralizing IgG antibodies to the homologous VSV-Indiana serotype but only VSV-IND N was able to trigger an IgG response to the heterologous VSV-New Jersey serotype. This data suggest that Th epitope(s) in the VSV-IND N are responsible for the observed cross-reactivity of T helper cells.

Animals↗

PubMed1994

Characterization of T-helper epitopes of the glycoprotein of vesicular stomatitis virus.

The T-helper (Th) cell epitopes in the glycoprotein (GP) of vesicular stomatitis virus serotype Indiana (VSV-IND) were analyzed with a complete panel of overlapping synthetic peptides. Three Th-cell epitopes in C57BL/6 (H-2b) mice and two epitopes in BALB/c (H-2d) mice were defined by their ability to stimulate in vitro proliferation of virus-primed, CD8+ T-cell-depleted spleen cells in a class II-restricted manner. A series of CD4+, I-Ab-restricted T-cell hybridomas from VSV-primed C57BL/6 mice were characterized by their production of interleukin-2 and interleukin-3 upon stimulation with VSV-IND or purified VSV GP in vitro. Of nine hybridomas derived from three independent fusions, five were specific for amino acids (aa) 415 to 433 (p8) of VSV-IND GP, three recognized aa 52 to 71 (p41), and one reacted against aa 316 to 335 (p17). Fluorocytometric analysis of Th hybridomas or VSV-stimulated T-cell lines with monoclonal antibodies specific for the T-cell receptor V beta chain did not reveal obvious correlations between the T-cell receptor V beta gene segment used and the epitope recognized. All three peptides recognized by H-2b mice and both epitopes recognized by H-2d mice which were characterized in primed T-cell populations were capable of activating specific Th cells in vivo as measured by the induction of antibody class switch from immunoglobulin M (IgM) to IgG. Thus, the epitopes are relevant for VSV GP-specific Th response in vivo and are able to provide functional help for the production of anti-VSV-specific neutralizing IgG antibodies.

Amino Acid Sequence↗

PubMed1994

Vesicular stomatitis virus Indiana glycoprotein as a T-cell-dependent and -independent antigen.

The neutralizing immunoglobulin M (IgM) response to vesicular stomatitis virus (VSV) has been shown to be largely T-cell independent in several T-cell-deficient models of mice. By using different antigen froms of VSV, VSV antigen doses could be graded in vivo (infectious > > UV inactivated > formalin inactivated). The present study reveals a T-cell-dependent component of the neutralizing IgM response in nude mice given intravenous injections of low doses of noninfectious UV-inactivated VSV serotype Indiana (VSV-IND) only if the mice are transfused with VSV-IND-specific helper T cells. Instead, nude mice immunized with infectious VSV, which leads to greater antigen doses in vivo, were able to mount an IgM response in the absence of T cells. These results indicate that the IgM response to low doses of VSV-IND glycoprotein (G) is T-cell dependent. Nude mice immunized with infectious VSV also made a variable but low VSV-IND-neutralizing IgG response. A VSV-IND matrix (M)-specific helper T-cell line rendered this response more consistent, much higher, and longer lasting. Thus (i) VSV-G induces a mostly T-cell-independent but partially T-cell-dependent IgM (the latter can be visualized best at low doses of antigen) and (ii) the antibody response to VSV in nude mice proceeds through steps, i.e., IgM and IgG, that are dose dependent. The results suggest that the predominant role of helper T cells may be to expand and maintain the individual steps of differentiating B cells.

Animals↗

PubMed1994

Enhanced establishment of a virus carrier state in adult CD4+ T-cell-deficient mice.

CD4+ T cells play an important role in regulating the immune response; their contribution to virus clearance is variable. Mice that lack CD4+ T cells (CD4-/- mice) and are therefore unable to produce neutralizing antibodies cleared viscero-lymphotropic lymphocytic choriomeningitis virus (LCMV) strain WE when infected intravenously with a low dose (2 x 10(2) PFU) because of an effective CD8+ cytotoxic T-cell (CTL) response. In contrast, infection with a high dose (2 x 10(6) PFU) of LCMV strain WE led to expansion of antiviral CTL, which disappeared in CD4-/- mice; in contrast, CD4+ T-cell-competent mice developed antiviral memory CTL. This exhaustion of specific CTL caused viral persistence in CD4-/- mice, whereas CD4+ T-cell-competent mice eliminated the virus. After infection of CD4-/- mice with the faster-replicating LCMV strain DOCILE, abrogation of CTL response and establishment of viral persistence developed after infection with a low dose (5 x 10(2) PFU), i.e., an about 100-fold lower dose than in CD(4+)-competent control mice. These results show that absence of T help enhances establishment of an LCMV carrier state in selected situations.

Animals↗

PubMed1994

[Viral antigen induced autoimmunity: an animal model for diabetes mellitus type I].

To address the mechanisms of tolerance to extrathymic proteins, we have generated transgenic mice expressing the lymphocytic choriomeningitis virus (LCMV) glycoprotein (GP) in the beta islet cells of the pancreas. The fate of LCMV-GP-specific T cells was followed by breeding the GP transgenic mice with T cell receptor transgenic mice, specific for LCMV-GP and H-2Db. These studies suggest that "Peripheral tolerance" of self-reactive T cells does not involve clonal deletion, clonal anergy, or a decrease in the density of T cell receptors or accessory molecules. Instead, this model indicates that potentially self-reactive cytotoxic T cells may remain functionally unresponsive, owing to a lack of appropriate T cell activation. Infection of transgenic mice with LCMV readily abolishes peripheral unresponsiveness to the self LCMV-GP antigen, resulting in a CD8+ T cell-mediated diabetes. These data suggest that similar mechanisms may operate in several so called "T cell-mediated autoimmune diseases". A synthetic peptide corresponding to an immunodominant epitope of lymphocytic choriomeningitis virus glycoprotein (LCMV-GP) was used to prime or to tolerize CD8+ T cells in vivo, dependent on the mode of immunization. Peptide-specific tolerance was then examined in transgenic mice expressing LCMV-GP in the beta islet cells of the pancreas; these mice develop CD8+ T cell-mediated diabetes within 8-14 days after LCMV infection. Specific peptide-induced tolerance prevented autoimmune destruction of beta islet cells and diabetes in this transgenic mouse model.

Animals↗

PubMed1993

The influence of antigen organization on B cell responsiveness.

The influence of antigen epitope density and order on B cell induction and antibody production was assessed with the glycoprotein of vesicular stomatitis virus serotype Indiana [VSV-G (IND)]. VSV-G (IND) can be found in a highly repetitive form the envelope of VSV-IND and in a poorly organized form on the surface of infected cells. In VSV-G (IND) transgenic mice, B cells were unresponsive to the poorly organized VSV-G (IND) present as self antigen but responded promptly to the same antigen presented in the highly organized form. Thus, antigen organization influences B cell tolerance.

Animals↗

PubMed1993

Impairment and delay of neutralizing antiviral antibody responses by virus-specific cytotoxic T cells.

After infection with some viruses that tend to persist, neutralizing antibodies are generated rather late, i.e., after 1 to 3 mo. This has been observed for HIV, Hepatitis B infection in man, or after infection with lymphocytic choriomeningitis virus (LCMV) in mice. In contrast, nonneutralizing antibodies to LCMV are generated by day 7 and reach high titers by day 10. This study attempts to evaluate reasons for late and low titered neutralizing antibody responses. After a primary infection with low doses (10(2) plaque forming units) of the poorly cytopathic LCMV-WE, neutralizing antibodies were rarely produced at detectable levels before days 60 to 120. In vivo depletion of CD8+ CTL led to a marked enhancement and acceleration of kinetics of the neutralizing antibody response. In contrast, nonneutralizing antibodies, including those specific for LCMV-GP carrying the neutralizing determinant, were detectable very early, i.e., 4 to 7 days after infection, with maximum titers usually by day 10 irrespective of the presence or absence of CTL. Mice completely lacking CD8+ T cells because of deletion by homologous recombination (CD8-/-) also exhibited neutralizing antibodies early by day 10 to 20, and by day 120 reached very high titers. The neurotropic isolate LCMV-ARMSTRONG induced low but significant titers of neutralizing antibodies relatively early (i.e., by days 7 to 10), whereas the lympho-viscerotrope mutant virus LCMV-ARMSTRONG Cl-13 did not. Early and effective CTL responses causing immunopathology (and immunosuppression) correlated with the absence of neutralizing antibodies. The discrepancy between the CTL-dependent inhibition of neutralizing versus unimpaired anti-GP ELISA responses cannot be explained by different Ag doses alone. Additionally, it may reflect different kinetics of the responses, whereby the later neutralizing responses possibly requiring IgG affinity maturation may be more susceptible to general immunosuppression; also B cells expressing neutralizing receptors, but not those with antibodies binding GP (or NP), may be actively infected, therefore they present viral peptides on class I MHC Ag and may become targets for anti-LCMV-specific CTL.

Animals↗

PubMed1993

Immune responses in interleukin-2-deficient mice.

The role of costimulatory signals in T cell induction was evaluated in mice lacking the interleukin-2 (IL-2) gene. In vitro secondary antiviral T cell responses were absent unless IL-2 was added, confirming the crucial role of IL-2 in vitro. In vivo, primary and secondary cytotoxic T cell responses against vaccinia and lymphocytic choriomeningitis virus were within normal ranges. B cell reactivity to vesicular stomatitis virus was not impaired. T helper cell responses were delayed but biologically functional. Natural killer cell activity was markedly reduced but inducible. These normal in vivo immune responses in IL-2-deficient mice question the importance of IL-2 as defined by in vitro studies.

Animals↗

PubMed1993

Comparison of the sensitivity of in vivo and in vitro assays for detection of antiviral cytotoxic T cell activity.

Sensitivities of in vitro and in vivo assays for the detection of lymphocytic choriomeningitis virus (LCMV)-specific CTL were compared. Measurement of primary cytotoxicity was the least sensitive of all the tested assays. However, when the same 51Cr release assays were performed after in vitro restimulation, this in vitro method was found to be more sensitive than all of the five in vivo assays tested. Assessment of CTL-mediated protection against LCMV replication in spleens was most sensitive among the in vivo tests, followed by CTL-mediated resistance to intracerebral or intraperitoneal challenge infections with vaccinia-LCMV-recombinant virus. Inhibition of LCMV-induced choriomeningitis and of the footpad swelling reaction were least sensitive in detecting LCMV-specific CTL. As discussed, the presented sensitivity gradient may most probably be generalized.

Animals↗

PubMed1993

Virus-specific cytotoxic T cell-mediated lysis of lymphocytes in vitro and in vivo.

Virus-specific CTL play a major role in early antiviral protection against lymphocytic choriomeningitis virus (LCMV). When mice are infected with high doses of certain LCMV isolates, the initiated CTL response may vanish before the virus is eliminated completely. To evaluate the possibility that this may be because of CTL lysing CTL, we studied the susceptibility to lysis of LCMV-specific CTL clones and of primary immune spleen cells in vitro and in vivo. Confirming earlier reports, CTL were lysed in vitro when incubated with their specific Ag peptide; lysis was MHC restricted because T cell clones derived from allogeneic bone marrow chimeras that do not express the correct restriction element were not susceptible to mutual CTL lysis. The density of CTL cultures correlated with the degree of inactivation of CTL, indicating that CTL lysed each other mutually rather than committed suicide. To assess CTL-mediated lysis of CTL in vivo, mice were infected s.c. into the footpad with LCMV, and shortly before the CTL-dependent footpad swelling developed by day 6 to 7, specific peptide was injected locally. The regional lymph node (LN) was reduced in size, contained about 10 times fewer cells, and only 2% to 3% of the lytic activity when compared with several control LN. This drastic local effect was observed within a few hours and was accompanied by DNA fragmentation. Thus, in this model system, peptide loading of lymph node cells may lead to CTL-mediated cytotoxicity and death of T cells (including CD8+ effector cells and other LN cells) in vivo. These results may suggest that, during overwhelming virus infections, lysis of CTL passively loaded with relevant released peptides possibly may contribute to the impairment of immune responses.

Animals↗

PubMed1993

Induction of diabetes is influenced by the infectious virus and local expression of MHC class I and tumor necrosis factor-alpha.

To study self reactivity, a transgenic mouse model has been established in which the lymphocytic choriomeningitis virus (LCMV) glycoprotein (gp) is expressed in the beta-islet cells of the pancreas (rat insulin promoter (RIP)-gp). These mice (H-2b) do not spontaneously develop diabetes; however, infection with the LCMV strain WE rapidly induces hyperglycemia. In this study, comparative analysis of H-2k RIP-gp-transgenic animals demonstrated that the haplotype influences the incidence and kinetics of diabetes and alters the requirement for the CD4+ T cell subset. This study also showed that the properties of the virus expressing the self target Ag determined whether hyperglycemia occurred in RIP-gp-transgenic mice. Various LCMV strains were able to induce diabetes in RIP-gp-transgenic animals, whereas infection with a recombinant vaccinia virus expressing LCMV-gp (vacc-gp) did not induce diabetes. However, vacc-gp could induce diabetes in double (RIP-gp/TCR)-transgenic mice, where the majority of CD8+ T cells expressed a receptor specific for LCMV-gp, suggesting that a critical number of self-reactive T cells must be activated to induce disease. Notably, histologic analysis of pancreata taken various days after LCMV or vacc-gp infections indicated that induction of diabetes coincided with an increase in MHC class I expression on the islets of Langerhans. Additional studies with vacc-gp were done to determine other factors that possibly enhance autoimmune attack. Transgenic mice expressing both LCMV-gp and TNF-alpha under the control of the RIP were infected with vacc-gp, and 50% of RIP-gp/TNF-alpha-transgenic animals became hyperglycemic. These data suggest that the increased local lymphocyte traffic as a result of TNF-alpha expression attracts activated gp-specific T cells, enhancing the possibility of hyperglycemia. Collectively, these results demonstrate that the induction of diabetes in this model is influenced by the MHC haplotype, the infectious agent, TNF-alpha expression, the level of MHC class I expression, and the induction of a threshold number of self-reactive CTL.

Animals↗

PubMed1993

Nonimmunogenic tumor cells may efficiently restimulate tumor antigen-specific cytotoxic T cells.

Induction of immunity to a viral protein that had been transfected into a tumor cell line was studied. The nucleoprotein (NP) of vesicular stomatitis virus (VSV) was used as a model tumor-associated Ag after transfection into EL-4, and H-2b thymoma originating from C57BL/6 mice. The NP-transfected cell line (EL-4NP) was lysed by NP-specific CTL and was found to restimulate NP-specific CTL in vitro as efficiently as did VSV-infected macrophages. Despite both of these in vitro characteristics, C57BL/6 mice inoculated with EL-4NP did not mount a measurable NP-specific CTL response and developed a lethal tumor as rapidly as did mice given control EL-4. This lack of immunogenicity could not be explained by down-regulation of MHC class I molecules or by loss of NP; even EL-4NP cells metastasizing to the spleen kept their high restimulatory capacity and excellent target characteristics. However, once mice were immunized with VSV or with a vaccinia-VSV-NP recombinant virus they were protected against tumor growth of EL-4NP by CD8+ CTL but not by CD4+ T cells. Taken together, the failure of the tumor-associated Ag to induce a protective T cell response in vivo despite its excellent capacity to restimulate CTL in vitro may encourage adjuvant immunotherapy in cancer; even the effects of weakly immunizing tumor vaccines, e.g., recombinant viruses, may be efficiently amplified by tumor cells.

Animals↗

PubMed1993

T cell-dependent IFN-gamma exerts an antiviral effect in the central nervous system but not in peripheral solid organs.

The antiviral relevance of soluble mediators that may operate in the vicinity of virus specific effector T cells was investigated. Mice were immunized with vesicular stomatitis virus (VSV) wild type (wt) and subsequently challenged with a mixture of two vaccinia recombinant viruses, one expressing the nucleoprotein of VSV (vacc-VSV-NP) the other expressing the glycoprotein of lymphocytic choriomeningitis virus (vacc-LCMV-GP). It was determined whether or not the VSV wt-induced memory T cell response that is protective against vacc-VSV-NP would inhibit growth of the nonrecognized vacc-LCMV-GP. In ovaries and testes replication of vacc-LCMV-GP was not inhibited. In contrast, the T cell response against vacc-VSV-NP nonspecifically inhibited growth of the non-recognized vacc-LCMV-GP in the central nervous system. This inhibiting effect was partly abrogated by treatment with anti-IFN-gamma antiserum but not by an anti-TNF-alpha antiserum. Similar results were obtained in VSV wt-immune H-2b mice, which eliminate vacc-VSV-NP by CD8+ T cells and in H-2k mice, which eliminate vacc-VSV-NP by a CD4+ T cell-dependent mechanism. These data suggest that a protective bystander effect mediated by soluble CD8+ and CD4+ T cell-dependent factors may be demonstrated against vaccinia virus only in an organ such as the central nervous system in which the blood-brain barrier inhibits diffusion and draining of the soluble antiviral factors released by specific effector T cells.

Animals↗

PubMed1993

Tolerance induction by clonal deletion of CD4+8+ thymocytes in vitro does not require dedicated antigen-presenting cells.

The cellular requirements of T cell tolerance induction in the thymus by clonal deletion was investigated by using an in vitro assay: thymocytes from mice expressing a transgenic TcR specific for lymphocytic choriomeningitis virus (LCMV) and H-2Db were co-cultured with various H-2b cell types as antigen-presenting cells in the presence of the antigenic LCMV peptide. The results revealed that all cell lines examined including embryonic and transformed fibroblasts, melanoma cells, cortical thymic epithelial cells, lymphomas and neuronal cells induced an antigen dose-dependent deletion of CD4+8+ thymocytes. Similarly, highly enriched accessory cell populations from thymus and spleen (macrophages, dendritic and cortical epithelial cells, i.e. thymic nurse cells) could induce antigen-specific depletion of immature CD4+8+ thymocytes. Depending on the cell type examined micromolar to picomolar concentration of LCMV peptide were required to induce deletion. The effectiveness of deletion by the different cell types did not correlate with their major histocompatibility class I expression level; it was, however, influenced by the presence of ICAM-1 adhesion molecules.

Animals↗

PubMed1993

Alloreactive cytotoxic T cells can develop and function in mice lacking both CD4 and CD8.

Using the technique of homologous recombination in embryonic stem cells, a mouse strain without functional CD4 and CD8 genes has been generated. Surprisingly, these mice contain significant numbers of alpha beta T cells. Although mice deficient for CD8 only do not show any cytotoxic response when their T cells are stimulated with either alloantigen or viral antigen, the CD4-8- mice do generate alloreactive cytotoxic T cells. These cytotoxic T cells bear the alpha beta T cell receptor and recognize major histocompatibility complex class I antigens. In addition fully allogeneic skin transplants were rejected but skin transplants expressing only minor transplantation antigens were not. Virus-specific cytotoxic T lymphocytes were also not detected. It seems that alloreactive cytotoxic T cells can be induced and exert their effector function in vitro and in vivo in the absence of CD8, and that they can develop and mature in vivo without the CD8 molecule or the signals it might provide.

Animals↗