Humoral immunity in secondary poxvirus infection
Creators: name: Tahiliani, Vikas
Publisher: name: The Australian National University
Year: 2010
Subjects: subject: QR405.T35 2010
subject: Orthopoxviruses
subject: Host-virus relationships
subject: Immunity

description: The studies described in this thesis have utilised the orthopoxvirus ectromelia (ECTV), a natural mouse pathogen that causes a disease termed mousepox, to address the roles of specific innate cell types and T cell subsets in virus control during a secondary infection and more importantly, whether CD4{u207A} T cell help is important for generation of protective recall antibody responses. There are 3 key reasons why the experiments detailed in this thesis were undertaken. First, much of our current understanding of the induction and maintenance of long-lived antibody responses is based on studies using non-replicating model antigens, however, these may not be relevant to replicating viruses. Second, humoral immunity to smallpox (caused by the orthopoxvirus variola), has been reported to be stable and last longer than memory CD4{u207A} or CD8{u207A} T cell responses. Third, recent studies from the ECTV and monkeypox virus models suggest that T cell responses may be redundant but antibody is necessary and sufficient for recovery from a secondary infection. The current thinking in the poxvirus field is that CD4{u207A} T cell help is not required for recall antibody responses for recovery from secondary poxvirus infection. To study the requirements for protection against secondary infection, a prime-challenge regime was used, in which avirulent ECTV (ECTV-TK{u207B}) was used to prime mice that were then challenged with virulent ectromelia virus (ECTV-WT). In mice primed with ECTV-TK{u207B}, the level of virus-neutralizing Ab returned to minimal levels 35 days after priming. Based on this observation it was assumed that the ECTV-TK{u207B} might not be efficient in inducing a B cell memory response. Results presented in Chapter 3 established that mice primed with ECTV-TK{u207B} were able to induce an efficient memory response through the induction of long-lived ECTV - specific ASCs. Neutrophils, plasmacytoid dendritic cells (pDCs), natural killer (NK) cells, T cell subsets and antibody are critical for recovery of mice from primary ECTV infection. Previous studies have shown that recovery of mice from the acute phase of secondary ECTV infection is strictly dependent on antibody responses and does not require CD4{u207A} T cell help or effector functions of CD8{u207A} cytotoxic T lymphocytes. However, neutrophils, pDC and NK cells could contribute to virus control in the absence of T cell subsets. The results presented in Chapter 4 show that the ability of mice individually depleted of neutrophils, pDC and NK cells subsets with specific monoclonal antibodies to overcome a secondary ECTV challenge was not diminished but concurrent depletion of NK cells, pDCs and neutrophils affected virus control and this was clearly evident when one or more T cell subsets were also depleted. Mice depleted of one or both T cell subsets, NK cells, pDCs and neutrophils were unable to control viral load, had a severe reduction in ECTV -specific ASCs and a reduction in the serum neutralising antibody response. Thus, the antibody response alone is not sufficient to mediate protection during the acute phase of secondary ECTV infection. The roles of CD4{u207A} T cell help for antibody production and effector functions of CD8{u207A} T cells over the longer term, past the acute stage of a secondary infection, were canvassed in Chapter 5. Although CD4{u207A} T cell help was apparently not required during the acute phase of a secondary infection, this subset was clearly important in the maintenance of ECTV-specific long-lived ASCs and in the generation of neutralizing antibody as the infection progressed and the response developed. In the continued absence of CD4{u207A} T cells over 35 days post-challenge (achieved by CD4{u207A} T cell depletion), there was a significant increase in the viral load, which correlated with a reduction in ECTV-specific ASCs in the BM, a significant reduction in splenic GC B cells and a significant decrease in levels of virus-neutralizing antibody by D 35 p.c. This was further confirmed using the HEL-specific B cell receptor transgenic system and recombinant viruses that express a mutant version of the antigen, hen-egg lysozyme (HEL) as a viral envelope protein. In spleens of mice depleted of CD4{u207A} T cells, there was a reduction in SW(HEL) GC B cells and a significant decrease in SWHEL GC B cells by D 35 p.c., which SW(HEL) correlated with the reduction in levels of anti-HEL IgG. The effector function of CD8{u207A} T cells becomes critical for virus control in the absence of CD4{u207A} T cells. In the absence of CD4{u207A} T cell help, the effector molecules perforin and possibly granzymes A and B expressed by CD8{u207A} T cells contribute to viral control during the later phases of secondary ECTV infection. Taken together, the results from experiments reported in this thesis provide key findings on the mechanism(s) of protection during a secondary ECTV infection during the acute phase (6-8 days post-challenge), maintenance phase (9-21 days post-challenge) and the long-term maintenance (22-35 days post-challenge). During the acute phase of a secondary ECTV infection, the NK cells, pDCs, neutrophils, T cells and extrafollicular antibodies are important in mediating protection. As the infection progresses, the extrafollicular response continues to be important in mediating protection during the maintenance phase of infection in the absence of CD4{u207A} T cell help. However as the infection further progresses, CD4{u207A} T cell help becomes important for long-term maintenance of antigen-specific memory B cells and long-lived ASCs. Protection mediated during this phase of infection requires CD4{u207A} T cell helper-dependent production of ECTV-specific antibody and also CD8{u207A} T effector cells.