Fig. 5.

Latency-reversal assay in primary CD4+ T-cells shows suppression of HIV latency reversal with Trx/TrxR inhibitors. (A) Primary CD4+ T cells were isolated from fresh human whole blood using negative selection and stimulated with anti-CD3/anti-CD28 antibodies on the same day. After 3 d of stimulation, the cells were infected with concentrated lentivirus containing the full-length HIV-1 genome for 2 h by spinoculation at room temperature (42). Next, CD4+ T cells were sorted 1 d after infection and GFP– cells were seeded into 48-well plates. One day after seeding, cells were treated with drugs of interest. Measurements of GFP fluorescence were carried out using flow cytometry after a 24-h incubation period. (B) Twenty-four-hour treatment of drugs of interest in infected GFP− sorted primary CD4+ T-cells. Sorted primary CD4+ T-cells were stimulated using PMA and Ionomycin. Resulting percent reactivation is normalized with the PMA + ionomycin control. PX12, NSC 401005 (D75), NSC 400938 (D35), and tiopronin treatments reveal suppression of reactivation using different concentrations, consistent with results in the JLat model (Figs. 2–4). (C) All treatments largely left cell viability intact according to PI staining. Significant P values are labeled with asterisks (familywise error rate ≤ 0.1). No data are statistically significant under familywise error rate ≤ 0.05. See SI Appendix, Fig. S9 for raw percent reactivation data without normalization.