Distinct yet complementary T cell subsets in the human cervix

From the Zhu and Corey Lab, Vaccine and Infectious Disease Division

The female reproductive tract (FRT), including the vagina and cervix, is the entry site for many sexually transmitted infections (STIs). Because of the importance of this barrier site and the need for mucosal vaccines against STIs such as HIV, understanding the immune cell subsets within the FRT is crucial. Previous research has shown that effective STI vaccines must elicit tissue-resident memory T cells (Trm)— T cells that are retained in a tissue compartment—in the FRT. However, Trm subsets in the FRT are understudied, and more knowledge is needed to inform mucosal vaccine design.

Herpes simplex virus 2 (HSV-2) is a prevalent STI for which a vaccine is desperately needed. Although it is known that CD4+ and CD8+ Trm are necessary for protection against HSV-2, comprehensive knowledge of FRT T cell phenotypes and T cell receptor (TCR) specificities are lacking. Drs. Tao Peng and Jia Zhu, along with colleagues from the Vaccine and Infectious Disease Division, used single-cell transcriptomic analysis, TCR tracing, and immunofluorescence staining to interrogate the tissue resident immune compartment in normal ectocervix samples from healthy women who are HSV-2-seropositive. They recently published this work in JCI Insight.

To perform unbiased analysis of the phenotypes and TCR specificities of FRT CD8 T cells, the research group first performed four different tissue processing methods to optimally enrich immune cell populations in ectocervical biopsies for 10X single-cell RNA-sequencing. This analysis revealed two distinct CD8 Trm subsets which, among other Trm markers such as CD103 and CD69, were defined by differential expression of two important functional molecules. One group highly expressed interferon-gamma (IFN-g), a critical anti-viral cytokine produced by CD8 and other cells, while the other group highly expressed Granzyme B (GzmB), a cytotoxic molecule used by CD8 T cells to kill infected cells. They next confirmed the presence of these two subsets by performing immunofluorescence staining of GzmB, IFN-g, and CD8 in HSV-2+ ectocervix samples. Not only were they able to confirm GzmB+IFN-g- and GzmB-IFN-g+ subsets at the protein level, but the two subsets were unequally distributed among anatomical compartments: GzmB+ CD8 were largely located in the epithelium, while IFN-g+ CD8 were predominantly found in the stroma. This suggests that distinct CD8 Trm subsets may have unique functions throughout the compartments of the ectocervix.

Graphical abstract from publication.
Graphical abstract from publication. Image from publication, provided by Tao Peng.

In support of putative differential functions in CD8 subsets, they also found that the expanded TCR clonotypes of the two subsets were non-overlapping, demonstrating that GzmB+IFN-g- and GzmB-IFNg+ CD8 subsets in the FRT may have different roles in host defense and different prior histories responsible for their placement. Based on the interesting TCR findings, the authors finished the study by asking if the CD8 Trm ectocervix subsets recognized HSV-2 antigens. Comparing the TCR sequences of FRT CD8 subsets to known HSV-2-specific TCR sequences isolated from blood, they found that both subsets recognized HSV-2 antigens, suggesting that both subsets of CD8 Trm cells are involved in anti-HSV-2 defense in human cervix.

Together, these findings “extend our knowledge about tissue-based immunity in the female genital tract, specifically the ectocervix, the site of ­­many mucosal pathogens such as HSV-2,” explained Dr. Peng. “We identified two major populations of CD8 TRM in the human ectocervix that exhibit distinct gene expression of antiviral defense and tissue residency markers, anatomic locations, and expanded clonotypes that target anatomically relevant viral antigens,” Dr. Peng said. This knowledge could be leveraged in future STI vaccine design, where specific anti-viral CD8 Trm subsets might be targeted to elicit potent immunity. Going forward, more work is needed to understand the distinct roles of antiviral defense by these cytolytic and non-cytolytic subsets of CD8 Trm cells in female FRT and other relevant antigens that these CD8 Trm cells may target to prevent or control HSV-2 infection.  Further, this study emphasizes the importance of considering CD8 Trm function in conjunction with localization for a more complete picture of mucosal immunity.

Peng T, Phasouk K, Bossard E, Klock A, Jin L, Laing KJ, Johnston C, Williams NA, Czartoski JL, Varon D, Long AN, Bielas JH, Snyder TM, Robins H, Koelle DM, McElrath MJ, Wald A, Corey L, Zhu J. Distinct populations of antigen-specific tissue-resident CD8+ T cells in human cervix mucosa. JCI Insight. 2021 Aug 9;6(15):149950. doi: 10.1172/jci.insight.149950.

This work was supported by the National Institutes of Health.

Fred Hutch/UW Cancer Consortium members Larry Corey, Jia Zhu, David Koelle, Julie McElrath, Anna Wald, and Jason Bielas contributed to this work.