Germinal center T follicular helper cells are highly permissive to HIV-1 and alter their phenotype during virus replication

HIV-1 replication is concentrated within CD4⁺ T cells in B cell follicles of secondary lymphoid tissues during asymptomatic disease. Limited data suggest that a subset of T follicular helper cells (T_FH) within germinal centers (GC) is highly permissive to HIV-1. Whether GC T_FH are the major HIV-1 virus-producing cells in vivo has not been established. In this study, we investigated T_FH permissivity to HIV-1 ex vivo by spinoculating and culturing tonsil cells with HIV-1 GFP reporter viruses. Using flow cytometry, higher percentages of GC T_FH (CXCR5^highPD-1^high) and CXCR5⁺programmed cell death-1 (PD-1)^low cells were GFP⁺ than non-GC T_FH (CXCR5⁺PD-1^intermediate) or extrafollicular (EF) (CXCR5^-) cells. When sorted prior to spinoculation, however, GC T_FH were substantially more permissive than CXCR5⁺PD-1^low or EF cells, suggesting that many GC T_FH transition to a CXCR5⁺PD-1^low phenotype during productive infection. In situ hybridization on inguinal lymph node sections from untreated HIV-1-infected individuals without AIDS revealed higher frequencies of HIV-1 RNA⁺ cells in GC than non-GC regions of follicle or EF regions. Superinfection of HIV-1-infected individuals' lymph node cells with GFP reporter virus confirmed the permissivity of follicular cells ex vivo. Lymph node immunostaining revealed 96% of CXCR5⁺CD4⁺ cells were located in follicles. Within sorted lymph node cells from four HIV-infected individuals, CXCR5⁺ subsets harbored 11-66-fold more HIV-1 RNA than CXCR5^- subsets, as determined by RT PCR. Thus, GC T_FH are ^highly permissive to HIV-1, but downregulate PD-1 and, to a lesser extent, CXCR5 during HIV-1 replication. These data further implicate GC T_FH as the major HIV-1-producing cells in chronic asymptomatic HIV-1 infection.