GSBS Dissertations and Theses

Title

T Cell Receptor-Dependent and Independent Events During Potent Anti-Viral T Cell Responses

Approval Date

2-1-1998

Document Type

Doctoral Dissertation

Department

Graduate School of Biomedical Sciences, Program in Immunology/Virology

Subjects

Receptors, Antigen, T-Cell; T-Lymphocytes; Infection; Academic Dissertations; Dissertations, UMMS

Abstract

The relative contribution of T cell receptor-dependent stimulation versus TcR-independent bystander stimulation in the massive increase in the number of activated proliferating CD8+ T cells seen during acute many acute viral infections is unclear. To determine if this increase was the result of TcR-independent bystander activation and proliferation, anti-viral cytotoxic T lymphocytes were induced in vivo via DNA immunization so that the anti-viral immune response could be examined in the absence of the high levels of cytokines generated during acute infection. After a single immunization with a plasmid encoding the nucleoprotein of the lymphocytic choriomeningitis virus (LCMV) a nearly 2 log10 reduction in viral titers in the spleen was observed 3 days after LCMV infection. After 2 or 3 immunizations a greater that 3 log10 inhibition of viral titers in the spleen was observed, with most animals having no detectable virus. After intracerebral challenge vaccinated animals displayed either protection or enhanced immunopathology leading to accelerated kinetics of death. By limiting dilution analysis LCMV-specific CTL precursors were detected in both the spleen and lymph nodes of vaccinated animals. C57BL/6 mice inoculated with DNA demonstrated an anamnestic CTL response detectable at days 4 after LCMV challenge. However, the numbers of CTL precursors elicited by DNA vaccination was too low to determine if cytokine-mediated TcR-independent bystander activation and proliferation had taken place.

HY-specific TcR-transgenic mice, which have a restricted TcR repertoire, and LCMV-carrier mice, which are tolerant to LCMV, were used to determine the extent of TcR-independent bystander activation and proliferation during acute LCMV infection. LCMV infection of C57BL/6 mice induced CTL that lysed uninfected H-2k and H-2d allogeneic targets, but, LCMV-induced CTL from HY- transgenic mice lysed only the H-2k-expressing cells. The HY-mice generated both anti-H-2k and anti-H-2d CTL in mixed lymphocyte cultures, strongly suggesting that the generation of allospecific CTL during acute LCMV-infection is antigen specific. During the LCMV infection there was blastogenesis of the CDB+ T cell population, but the HY-specific T cells remained small in size, and did not alter their expression of the activation molecules CD44 and MEL-14. In order to examine the potential for bystander stimulation under conditions of a very strong CTL response, T cell chimeras were made between normal and HY-transgenic mice. Even in the context of a normal vicus-induced CTL response, no stimulation of HY -specific T cells was observed, and HY-specific cells were diluted in number by day 9 post-infection. In LCMV-carrier mice in which donor and host T cells could be distinguished by Thy 1 allotypic markers, adoptive transfer of LCMV-immune T cells into LCMV-carrier mice, whose T cells were tolerant to LCMV, resulted in activation and proliferation of donor CDB cells but little or no activation of host CDB+ T cells. These results show that TcR-independent bystander activation of non virus-specific T cells is not a significant component of an anti-viral T cell response and support the hypothesis that the massive polyclonal CTL response to LCMV infection is virus-specific.

T cells activated during potent anti-viral immune responses are sensitized to undergo apoptosis after strong TcR-stimulation in a process known as activation-induced cell death (AICD). To determine if T cells, not participating in the immune response were also subject to AICD, LCMV-carrier mice were used. Using TUNEL flow cytometry, it was shown that after reconstitution of Thy 1.2+ LCMV-carrier mice with spleen cells from Thy 1.1+ LCMV-immune mice, the Thy 1.2+ host T cells which were not specific for the virus and did not proliferate in a bystander fashion, were rendered sensitive to TcR-induced apoptosis in vitro. This bystander sensitization to AICD was shown not to be dependent on the continued presence of activated proliferating donor cells during the in vitro culture period. Bystander sensitization to AICD was not the result of an antigen presenting cell defect, but rather was the result of an in vivo conditioning of the T cells themselves. The mechanism of this sensitization was, at least, partially dependent on the ability of host T cells to respond to IFNγ, and on the expression of Fas ligand on the activated, proliferating donor cells. This bystander sensitization to AICD may explain why memory T cell responses are so poor during acute viral infection and can serve as a potential mechanism for virus-induced immunosuppression.

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