Figure 4

Envelope-induced cytopathicity in CD4⁺ T cells. Envelope-mediated absolute CD4⁺ T cell depletion. Absolute loss of CD4⁺ T cells in cocultures of Env-expressing HeLa cells and purified CD4⁺ T cells was determined. Single living CD4⁺ T cells were morphologically gated, and the absolute loss of CD4⁺ T cells in the coculture was quantified by the addition of PE-labeled microbeads. *p = 0.022, **p < 0.007 (A). Bystander apoptosis induced by envelopes harbouring changes at position 38 and 140 in gp41. Cocultures of primary CD4⁺ T cells, previously stained with the DDAO cell tracker, and HeLa Env⁺ cells were performed and analyzed after 24 hours by flow cytometry. Single, nonfused CD4⁺ T cells were gated by the forward- vs. side-scatter characteristics and by being positives for the cell tracker. Detection of early (PI^-DIOC^-) and late apoptotic (PI⁺DIOC^-) populations were determined in this gated population for each recombinant Env in the presence or absence of the coreceptor inhibitor JM-2987 (B). Boxes represent median and interquartile range of values. Median values were compared in an inter-patient and in an intra-patient assay, using a Mann-Whitney test. Baseline samples (white boxes) and V38A mutant clones (gray boxes). *^ap = 0.031, *p = 0.0213, **p = 0.0038, and ns non-significant differences.