Elucidating human female genital tract resident DC response to HIV-1 exposure.

The female genital tract (FGT) represents a complex and dynamic environment with specialized immune mechanisms uniquely designed to maintain a delicate balance between protection against invading pathogens and accommodating the unique physiological changes associated with reproductive function. Dendritic cells (DCs) are critical in shaping mucosal immunity against pathogens and maintaining tissue homeostasis. The unique ability of DCs to recognize invading pathogens through pattern recognition receptors (PRRs), and prime naive T cell function, make DCs ideal targets for vaccination and therapeutic strategies against cancers and infections. However, in HIV pathogenesis, DCs are considered a double-edged sword due to their ability to secrete anti-viral proteins and resist viral replication but capture and transfer active viral particles to target CD4+ T cells. Understanding the heterogeneity of DC populations in the FGT, along with their unique contribution to HIV pathogenesis is key for targeted interventions. Overall design: Hysterectomy samples of endometrium, endocervix and ectocervix of female donor, determined to be healthy according to study criteria, (no cancer or active infections) were enzymatically digested to generate single-cell suspensions. To enrich for myeloid immune cells, the single cell suspensions were filtered to remove cells larger than 30mm, followed by depletion of CD3+ T cells, CD19+ B cells, CD235a+ red blood cells and fibroblasts through magnetic bead selection. Purified cells were incubated with HIV-BaL (R5) isolates (MOI=0.5) or media alone for 30 minutes at 37C, after which the cells were washed to remove excess unbound virus. The cells were subsequently stained with CITE-seq antibodies (AbSeq) and sample multiplexing antibodies (hs_SampleTag). Single-cell suspensions were processed on the BD Rhapsody system according to manufacturer's protocols. cDNA libraires were constructed using BD Rhapsody Whole Transcriptome (WTA) Amplification KIT (BD Biosciences, Catalog# 633801).