The majority of HIV infections in women are acquired during sexual intercourse, making it imperative to understand the immune microenvironment of the FGT for developing strategies to prevent infections. A significant effort has been made to understand if and how antibodies can confer protection in the FGT. Given the significant evidence of GI and increased HIV risk, very little is known about how GI can impact the profile and functions of antibodies in the genital tract. Here, we detected that in the presence of pre-existing GI, mucosal IgG subclasses and isotypes were significantly increased in both women who became HIV-infected and in those who remained uninfected.
In this case-control analyses we found significantly higher IgM in cases compared to controls which may indicate that mucosal IgM can be an early marker of pathogen exposure. Circulating IgM are elicited during the early stage in response to any infection including HIV29. Moreover, HIV-specific IgM were functionally effective in neutralizing and reducing HIV infections in cervicovaginal tissue models30. Natural IgM can limit HIV infection by modulating inflammation and T cell activation31,32. In addition, IgM can directly bind to CD4 + T cells and chemokine receptors CCR5, hindering HIV entry33. Together, these data highlight the varied functional capability of pentameric IgM to effectively crosslink and capture virus thereby reducing HIV infection. Upon class switching from IgM, IgG1 predominates during acute and chronic HIV infection11. We also found higher IgG1 in cases compared to controls. HIV-specific IgG1 have been associated with viral control and slower progression to HIV infection34,35. These data suggest that in the cases, at the pre-HIV infection stage, sexual exposure to HIV or other pathogens may have elicited increased mucosal IgG1 and IgM titres. However, whether these antibodies are functionally competent and HIV-specific IgG or IgM, remains undefined.
To determine genital antibody profiles in the presence of GI, women were stratified for the presence or absence of GI. IgM, IgG1, IgG3 and IgG4 were significantly higher in women with GI compared to women without GI. Both in vivo and in vitro studies suggested that IgM increases as a compensatory drive to resolve inflammation32,36. Therefore, increased mucosal IgM among women with GI may be in response to the inflammation. Studies have demonstrated cytokine-induced enhancement of IgG subclasses and isotypes37,38 explaining in part the increased antibodies with GI. Alternatively, compromised barrier integrity may also account for the increased antibody transudate in the presence of GI. Inflammation is strongly associated with the glycosylation of the fragment crystallizable (Fc) region of IgG subclasses particularly39. This reduces antibody affinities for Fcγ receptors decreasing functional activities40,41. Besides glycosylation, increased proteases have been associated with elevated inflammatory cytokines in the FGT7 which can lead to non-specific proteolytic antibody degradation compromising the function of the antibodies24. Therefore, the impact of GI on antibody functions in the genital tract need to be investigated in future studies. Nevertheless, the findings suggest that inflammatory environment in the FGT may be responsible for maintaining specific antibody responses, but this needs further investigation in large cohort studies.
Cytokines have been shown to influence Ig class switching and subclass synthesis38,42. In this study we demonstrated a link between genital inflammatory cytokines and mucosal antibodies. We found that prior to HIV infection, mucosal IgG subclasses and isotypes were positively and significantly associated with several inflammatory cytokines: TNF-α, IL-1α, IL-1β, IL-10 and IL-6; adaptive cytokines: IL-13, IL-4 and IFN-γ and chemotactic and growth factors cytokines: MIP-1α, MIP-1β, MCP-1, IL-8, CTACK, MIG after adjusting for potential confounders. Positive associations were observed between IL-13 and IgG4 and IgM. These data corroborate previous findings of IL-13 induced synthesis of IgM and IgG4 in human B cells in the presence of activated T cells43. Furthermore, IL-6 along with IFN-γ can inhibit IgG1 and enhance IgG2 production in stimulated PBMCs demonstrating the differential regulation of cytokines on human IgG subclass production38. The significantly direct association between IL-7 and IgG4 is also supported by a study showing IL-7 driven class switching to IgG444. Collectively, these preliminary findings suggest that the local isotypes and IgG subclasses may be influenced, in part, by pre-existing GI.
The strength of the study is the evaluation of rare pre-HIV infection mucosal samples from high-risk South African women in two clinical trials investigating mucosal Ig isotype/subclasses in relation to cytokines. Mucosal antibody profiling may be an additional predictor of pre-existing GI. However, these studies would indeed require large cohorts evaluating both mucosal cytokines and antibodies longitudinally and in parallel to establish their validity as an additional surrogate for GI. Understanding the association between GI and antibodies can provide further insight for developing effective HIV prevention strategies for at risk populations with underlying susceptibilities to GI. This study however has several limitations. We did not measure pathogen-specific antibodies, an important surrogate indicator of bacterial and viral pathogen exposure. All the participants were on some form of hormonal contraceptives, a mandatory inclusion criterion for trial participation. Therefore, we could not account for phase of the menstrual cycle, known to affect inflammatory cytokines and antibodies45,46. Despite these shortcomings, our data provides an enhanced dissection of how GI can affect the quantities of mucosal antibodies and, potentially impact their functions through the predominance of certain isotypes or subclasses over the other.
Our findings suggest that GI can shape or reconfigure the mucosal Ig subclass and isotype signature. However, further investigation is required to verify a plausible link between the local inflammatory milieu and the effect on genital antibodies and their functions. These data may be important for future HIV vaccine efficacy studies.