High p21 expression in HIC CD4⁺ T cells is not directly involved in HIV restriction. (A) Levels of p21 mRNA in MDMs from HICs and HDs at the time of infection (after 7-10 days of differentiation from monocytes). Each symbol represents one person. Median values for each group are shown. (B) Levels of p21 mRNA in nonactivated CD4⁺ T cells from HICs, HDs, viremic HIV-1–infected patients (VIR) and patients on effective HAART (ART; left). Levels of p21 mRNA in CD4⁺ T cells from HICs, HDs, VIRs, and ARTs after 5 days of activation with anti-CD3 (right). (A-B) The results are expressed as relative expression levels compared with cDNA in CD4⁺ T cells from one HD who was used as a common reference throughout the study. (C) Levels of HIV-1 replication 7 days after infection with replicative HIV-1 BaL (■) in CD4⁺ T cells from 2 HIC, 1 VIR, and 1 HD (top), and levels of p21 protein at the time of infectious challenge (bottom). The ratio of the optical densities of the p21 and GAPDH signals were calculated for each sample with Photoshop CS3 software (Adobe), and reported in the graph () relative to the HD control. (D) Levels of p21 mRNA in anti–CD3-activated CD4⁺ T cells from one HD and one HIC 24 hours after nucleofection of irrelevant and p21-specific siRNAs (top). Luciferase activity 48 hours after challenge of nucleofected cells with single-cycle HIV-1_VSV-G (bottom). Cells were challenged 24 hours after nucleofection. Means ± SDs of 3 independent measures are shown.