Background
Innate Lymphoid Cells type 3 (ILC3), abundant in the gut, sense the microbiome and present antigens to naive T cells, promoting regulatory T cell (Treg) differentiation over Th17 cells, fostering immune tolerance. Disruption of this balance is linked to inflammatory bowel disease. While HIV alters the Treg/Th17 ratio in the gut, it’s unclear whether it results from the impairment of the ILC3-T cell interaction in the gut of people living with HIV (PLWH).
Methods
Single-cell RNA sequencing was performed on gut tissues from 4 compartments (duodenum, ileum, colon, rectum) of PLWH under ART (n=7) and PWoH (n=2) collected at CIENI (Mexico). To balance the number of participants, we merged the dataset of biopsies from 6 PWoH previously generated by the RID-HIV initiative (HCRC, Belgium) and publicly available gut data (Gut Reference survey). Using the Seurat (v5.0) and related packages in R, cells were processed and corrected for batch effects across datasets. Downstream analyses consisted of differential gene expression, gene set enrichment analysis (GSEA), and cell-cell communication.
Results
There was a reduction in Treg/Th17 ratio and ILC3 frequencies, including RORC expression, in PLWH, suggesting an alteration of the ILC3-T cell interplay that could contribute to the well-established imbalance of the Treg/Th17 ratio and heightened inflammation observed in PLWH. TGFβ expression, critical for Treg differentiation, was lower in PLWH but was the most important signaling pathway upregulated in these cells (Figure). ILC3-Th17 interactions were more frequent than ILC3-Treg in PLWH. Cytokine signaling between ILC3 and Th17 was mainly mediated by TGFβ, targeting JUNB, IRF7 and NFKB2 in Th17. On the other hand, IFN-II signaling, which upregulates MHCII that presents bacterial antigens to naive T cells and promote their differentiation to Tregs, was most important in PWoH. TGFβ alone is insufficient for Treg differentiation, and decreased IFN-II signaling could impair MHCII expression in ILC3 leading to insufficient antigen presentation. Indeed, GSEA showed downregulation of key ILC3 effector functions in PLWH, including antigen processing cross presentation. We cannot rule out the importance of other antigen presenting cells.
Conclusions
Despite increased TGFβ signaling, these data suggest an impaired antigen presentation machinery that can foster Th17 differentiation. This likely contributes to intestinal inflammation and incomplete immune reconstitution in the gut despite ART.