We previously have shown that CPAF+IL-12 vaccinated animals display faster resolution, but not resistance, to genital chlamydial infection suggesting that cellular immunity, not neutralizing antibodies, may be involved in mediating this protective effect. In the current study, we examined the role of CD4+ T cells in CPAF+IL-12 mediated immunity against genital chlamydial infection. CPAF+IL-12 vaccination induced the generation of IFN-γ producing antigen-specific CD4+ T cells and resulted in enhanced resolution of genital chlamydial infection in C57BL/6 mice. Depletion of CD4+ T cells from such immunized animals abrogated the protective effects of vaccination, whereas adoptive transfer of CPAF-specific CD4+ T cells induced enhanced resolution of genital C. muridarum infection.
Splenocytes from CPAF+IL-12, but not mock (PBS) vaccinated animals displayed high levels of IFN-γ and minimal IL-4 production upon in vitro
stimulation with CPAF, demonstrating the induction of an Ag-specific Th1 type cellular response. There is accumulated evidence to suggest that Th1 responses and IFN-γ production are important for optimal resolution of genital chlamydial infection [1
]. To this end, Chlamydia
-specific Th1 clones, but not Th2 clones, have been demonstrated to be capable of adoptively transferring anti-chlamydial immunity [9
]. Conversely, MHC class II deficient, but not MHC class I deficient animals, displayed an inability to resolve a primary chlamydial infection [20
]. Mice with a targeted disruption in IFN-γ production (IFN-γ −/−
mice) have also been shown to display a marked inability to resolve chlamydial infection and prevent bacterial dissemination [21
]. In this regard, the bactericidal effect of IFN-γ on intracellular Chlamydia
in epithelial cell cultures has been shown to occur via the indoleamine-2,3-dioxygenase pathway in human cells versus p47 GTPases in murine cells [22
]. Thus, induction of Th1 responses is an essential component of an effective anti-chlamydial vaccine.
CPAF+IL-12 vaccinated animals exhibited high titers of serum anti-CPAF total Ab and IgG2b, and lower titers of IgG2a and IgG1 at day 40 after immunization. However, the contribution of these antibodies to protective immunity has yet to be characterized. Other studies suggest a possible role for antibodies in immunity against chlamydial infections. B cell deficient (μMT) animals, but not wild type animals displayed a severe inability to resolve chlamydial reinfection in the absence of CD4+
T cells, suggesting an important role for B cells in anti-chlamydial immunity [8
]. Additionally, mice deficient in Fc receptors have been shown to exhibit reduced antigen-specific T cell responses and sub-optimal resolution of infection following rechallenge [24
]. Specifically, Fc receptors of IgG2a (FcγRII) and IgA (Fcα) antibodies has been shown to be important for efficient chlamydial antigen presentation to T cells.
The enhanced resolution of genital chlamydial infection and protection against oviduct dilatation induced by CPAF+IL-12 vaccination was abrogated by depletion of CD4+ T cells. In addition, protective immunity could be adoptively transferred by CPAF-specific CD4+ T cells. Taken together with the fact that CPAF+IL-12 mediated immunity is highly dependent on endogenous IFN-γ production, these results suggest that CPAF induced anti-chlamydial immunity may be highly dependent on IFN-γ secreting Th1 type CD4+ T cells. The findings that CPAF-specific CD4+ T cells are important for induction of protective immunity has bearing on several aspects of chlamydial vaccine development for humans including (a) route of vaccination and type of adjuvant(s) required to elicit the appropriate type of immune response at the site of infection and (b) identification of MHC class II restricted protective epitopes within CPAF for development of subunit vaccines. To this end, we have recently found that CpG deoxynucleotides are also highly effective Th1 adjuvants when used with CPAF for induction of protective immunity against genital chlamydial infection in mice (Cong and Arulanandam, unpublished observations). Furthermore, we have determined that CPAF contains human HLA-DR4 (MHC class II) restricted epitopes that are capable of generating protective immunity against chlamydial infection (Murthy et al., In Press, Infection and Immunity).
Adoptive transfer of CPAF-specific CD4+
T cells induced comparable protective immunity against genital challenge to that induced by transfer of CD4+
T cells primed by C. muridarum
infection suggesting that CPAF may be a dominant chlamydial antigen for induction of protective immune responses. To this end, Chlamydia
sero-positive humans have been shown to exhibit higher titers of serum antibodies against CPAF than the highly expressed surface exposed antigens, such as MOMP and Hsp60 [26
]. Importantly, recombinant CPAF cloned from C. trachomatis
L2 induced protective immunity against C. muridarum
challenge. CPAF derived from these two strains display approximately 82% amino acid identity [15
] with greater homology between human serovars suggesting that the protective CPAF epitopes may be conserved across the species and serovars of Chlamydia
. In addition, monoclonal antibodies raised against serovar L2 CPAF have been shown to recognize CPAF from either serovar L2 or C. muridarum
], indicating a high degree of immunological cross-reactivity. In contrast, the most extensively characterized chlamydial vaccine candidate MOMP is homotypic and does not afford cross-protection [2
]. Additionally, MOMP has been shown to be predominantly a B cell immunogen [1
] and needs refolding before vaccination to induce protective immunity [4
In summary, we have described a defined chlamydial vaccine candidate (CPAF) that induces CD4+ T cell dependent protective immunity. The possibility of using CPAF as an alternative to, or in combination with, MOMP to induce robust anti-chlamydial immunity is currently being explored.