Investigating Mononuclear Phagocytes in Human Anogenital and Colorectal Tissues: Their Role in the Sexual Transmission of HIV

Abstract

When an invading pathogen penetrates the physical barriers of the human anogenital and colorectal tract the first cells of the immune system it interacts with are mononuclear phagocytes (MNP). In tissue these are comprised of Langerhans cells (LC) dendritic cells (DC) and macrophages. These cells detect and take up foreign antigens via an array of pathogen binding receptors unique to each cell subset and then present antigen fragments to CD4 T cells via MHC-II to elicit an immune response. In the context of HIV this process allows for the transfer of the virus to its primary target CD4 T cells in which the virus efficiently replicates leading to its dissemination throughout the body. Despite this, very little is known about the exact role MNPs play in the early events of sexual transmission of HIV. This is primarily due to the difficulty of access to human tissue, problems with cell isolation protocols and technological limitations in reliably detecting HIV at early time points. By establishing extensive collaborations with surgeons across Sydney, our lab has access to all the human anogenital and colorectal tissues that HIV may encounter during sexual transmission (labia, vagina, cervix, glans penis, foreskin, penile urethra, perineum, anus, rectum and colon) as well as abdominal tissue as means to liberate larger cell yields for functional assays. We have fully optimised our tissue digestion protocols to reliably extract MNPs from anogenital and colorectal tissue in the non-activated, immature state they are in within these tissues when they encounter HIV. We have also optimised highly sensitive HIV detection methods to directly investigate their initial interactions with HIV. LCs were previously thought to be the only MNP to be present in the human epidermis. However, using our optimised tissue processing and isolation protocols, we have identified two previously undescribed MNPs subsets within the epidermis of skin; CD33low MNP and CD11c+ DC. We went on to characterise these cells via RNAseq and functional assays to reveal that CD33low MNP are transcriptionally very similar to LCs while CD11c+ DC are transcriptionally similar to dermal conventional (c) DC2. Furthermore, we showed that, compared to LCs and dermal cDC2, CD11c+ DCs were more efficient at HIV uptake, supported higher levels of productive infection and transferred the virus to CD4 T cells more efficiently. They were also significantly enriched in anogenital tissues and thus are likely to be a preferential HIV target cell that may play a key role in HIV transmission. Finally, we investigated sub-epithelial MNP subsets and identified two further preferential HIV target cells; langerin+ cDC2 and CD14+ CD1c+ MNPs. Using RNAseq we showed that CD14+ CD1c+ MNPs were transcriptionally more similar to DCs than macrophages. We also fully defined the pathogen binding lectin receptor profiles at the transcriptional and cell surface expression levels of all known epithelial and sub-epithelial MNP subsets. Furthermore, we went on to investigate their interactions with HIV using HIV uptake assays, transfer assays and secreted HIV assays highlighting these two cells as key HIV target cells compared to the other sub-epithelial MNP present. Identifying the initial HIV target cells will be essential for guiding the development of a vaccine and better PrEP regimens, to aid in putting an end to the global HIV pandemic.