HIV Latency Is Established Directly and Early in Both Resting and Activated Primary CD4 T Cells
Fig 2
Primary CD4+ T cells transitioning from an activated state back to a resting state are more likely to become latently infected.
(A) Schematic of experimental procedure. Primary CD4+ T cells were isolated from uninfected donor blood and stimulated with αCD3/αCD28 activating beads in the presence of IL-2 for 72 h and were then allowed to return to a resting state over 20 days in the presence of IL-2. Cells were infected at peak activation (day 4) and every 5 days thereafter as they returned to a resting state. (B) Expression of activation markers CD69 and CD25 as the cells transition from an activated state to a resting state. Flow cytometry was performed 72 h post activation and every 5 days after the activation beads were removed. Data shown are from a single donor, but representative of three separate donors. Percentage of live cells is calculated from the live gate (forward vs side scatter plots) in the FACS analysis and represents the average of three donors. (C) Quantified values of the cells’ activation status from panel B. Data represents the average of three donors. (D) Infection profiles of primary CD4+ T cells as they transition back to a resting state. Cells were spinoculated with HIV Duo-Fluo I 72 h after activation and every 5 days after the activation beads were removed. Infection was analyzed by flow cytometry 72 h post-infection. Data shown are from a single donor, but representative of three separate donors. (E) Quantified values of latent infection and productive infection from panel D. Data represents the average of three donors. (F) Ratios of latent infection to productive infection were calculated using data from panel E. Data represents the average of three donors. *, P < 0.05; **, P < 0.01; ns, non-significant.