Opposing immunologic requirements in the female genital tract

From the Hladik lab, Vaccine and Infectious Disease Division

The introduction of semen into the female genital tract induces tolerogenic conditions in recipient immune system. This immunomodulation is partially facilitated by extracellular vesicles (EV) in semen, which contain exosomes, carrying proteins, messenger RNA, and non-coding microRNA. These cargos are known to regulate the immune response and may help to blunt the immune response that could interrupt conception. Conversely, appropriate immune responses are necessary to protect from sexually transmitted infections (STIs). As STIs are generally acquired in the presence of semen, otherwise-protective immune responses to pathogens may be sacrificed in favor of conception, presenting a challenge for vaccine design. Researchers in the Hladik lab of the Vaccine and Infectious Disease Division hypothesized that seminal EV (SEV) may act on the adaptive immune memory responses, and addressed this hypothesis in a recent Plos One publication. “It makes evolutionary sense for semen to inhibit immune responses against co-delivered antigens, to support conception and pregnancy,” explained Dr. Lucia Vojtech, the lead investigator of the study. “Knowing that extracellular vesicles (EV) are important modulators of immune responses, we wondered what the very high concentrations of EV in semen might be doing to memory immune responses,” she continued.

Dr. Vojtech explained that they “reasoned that if semen EV inhibit memory immune responses to viruses, this might be an additional hurdle to overcome for vaccine design against sexually transmitted pathogens.” To test this idea, the authors first isolated, purified, and labeled SEV from human semen and cultured them with immune cell subsets and found that antigen presenting cells (APC) such as dendritic cells uptake SEV, while T and B cells do not. This suggests that immune responses to SEV are mediated initially through APC. They then co-cultured human CD4+ and CD8+ T cells with APC and cytomegalovirus (CMV) or Epstein-Barr virus (EBV) in the presence or absence of SEV. Because these cells were taken from individuals who had been previously exposed to CMV and EBV, they should have displayed effective memory T cell responses to pathogens. However, the authors found that SEV presence inhibited production of antiviral cytokines by T cells, especially in CD8+ T cells. Interestingly, this effect was exacerbated when the viral antigen was in the form of a whole protein compared to a peptide, which requires more extensive processing by APC for antigen presentation.

To further understand which T cell pathways are affected by SEV, T cells were next stimulated with a superantigen that induces T cell signaling through the T cell receptor (TCR) or with PMA/ionomycin, which activates T cells in a TCR-independent manner. SEV presence inhibited T cell responses in both cases, although with more pronounced effects in T cells signaling through the TCR, suggesting that SEV conduct the most potent inhibition when T cells are interacting with their antigen-loaded APC. Given that SEV inhibitory activity increased with APC, the authors asked whether SEV act directly on APC instead of T cells. By separately culturing APC, CD4+, or CD8+ T cells with or without SEV and then combining with CMV peptide to assess T cell cytokine production, the authors found that CD8+ T cell cytokine responses were impaired when APCs alone were exposed to SEV, while CD4+ responses were largely not inhibited when either CD4+ T cells or APCs were separately exposed to SEV. In addition to decreasing antigen-specific cytokine production, SEV inhibition also inhibited CD8+ T cell ability to produce cytotoxic granules and to kill infected cells, demonstrating that SEV decrease T cell immune responses through multiple mechanisms. These results suggest that SEV act on CD4+ T cells when they are interacting with APCs, while SEV inhibit CD8+ responses by directly acting on the dendritic cells that present antigen.

 Seminal extracellular vesicles (SEV) modulate adaptive immune responses to viral pathogens in the female genital tract by decreasing antigen presentation to T cells and increasing antigen presenting cell (APC) production of inhibitory enzyme indoleamine 2,3-dioxygenase (IDO).
Seminal extracellular vesicles (SEV) modulate adaptive immune responses to viral pathogens in the female genital tract by decreasing antigen presentation to T cells and increasing antigen presenting cell (APC) production of inhibitory enzyme indoleamine 2,3-dioxygenase (IDO). Image provide by Lucia Vojtech.

Given that SEV-exposed APC were unable to efficiently activate CD8+ T cells, the authors asked which APC mechanisms were affected by SEV. Surprisingly, SEV exposure did not alter any surface expression of classical APC molecules needed for T cell co-stimulation. However, APCs such as dendritic cells also can directly produce immunosuppressive cytokines and molecules, and SEV increased the dendritic cells’ production of indoleamine 2,3-dioxygenase (IDO), an enzyme that suppresses effector T cell response at both the RNA and protein level. These findings suggest a potential mechanism by which APC dampen CD8+ T cell responses when SEV are present.

Dr. Vojtech summarized their findings: “We found that semen EVs impair antigen-presenting cell function, which leads to weaker memory T cell responses to viral antigens.” This work provides important context for future antiviral and vaccine design, as the opposing immune response requirements of pregnancy and viral exposure need to be considered. “This underscores the importance of considering the contribution of semen to infection dynamics and immune responses against STIs,” Dr. Vojtech said. The authors explain that ongoing research is needed to further understand the mechanisms by which semen may alter immune responses in the female genital mucosa so that fertility treatments and STI vaccine design can carefully consider these interactions.

This work was supported by the National Institutes of Health, the University of Washington Royalty Research Grant, the Center for Exposures, Diseases, Genomics, and Environment, and the Burroughs Wellcome Career Award.

UW/Fred Hutch Cancer Consortium members Florian Hladik and Martin Prlic contributed to this work.

Vojtech L, Zhang M, Davé V, Levy C, Hughes SM, Wang R, Calienes F, Prlic M, Nance E, Hladik F. 2020. Extracellular vesicles in human semen modulate antigen-presenting cell function and decrease downstream antiviral T cell responses. Plos One. https://doi.org/10.1371/journal.pone.0223901