Figure 2. HIV gp120 interaction with quiescent naïve CD4 T cells triggers LFA-1 activation and supramolecular rearrangement.

(A) Naïve CD4 T cells establish LFA-1-ICAM-1 interaction upon gp120 binding. The naïve CD4 T cells purified by negative selection from ex vivo HIV-seronegative PBMCs were introduced to a bilayer containing Alexa Fluor 488-labeled gp120 and Cy5-labeled ICAM-1, and images were acquired over an hour. Images from one representative cell to show the dynamics of cell interaction with gp120 and ICAM-1 over time are presented. Top panels show gp120 contact the cell made at the indicated time points, middle panels show ICAM-1 contact, and bottom panels display merged images (gp120 in green, ICAM-1 in red). (B) Morphology of ICAM-1 contact areas made upon the interaction of naïve CD4 T cells with gp120 and ICAM-1 on the bilayers. Images of representative cells and the percentages of cells that form symmetrical (top panel) or asymmetrical (bottom panel) ICAM-1 rings are shown. gp120 was added onto the bilayers at a density of 200 to 250 molecules/µm². (C) Naïve T cells were added to the bilayers in the presence of an anti-gp120 mAb that blocks gp120-CD4 interaction (654), an anti-V3 mAb that interferes with gp120 binding to the chemokine receptor (2219), or a mAb to the N-terminus of gp120 (EH21) that does not affect gp120 interaction with its receptors. The percentages of cells making gp120-positive (left) and ICAM-1-positive (right) contacts out of the total number of cells seen in the fields were calculated. The MAbs were used at 20 µg/ml. The graphs represent the averages +/- SEM of three independent experiments. Statistical analysis was done by one-sided Student's t test. (D and E) Morphology of ICAM-1 contact areas made upon the interaction of naïve CD4 T cells with bilayers containing ICAM-1 and monoclonal antibodies to CD4 (OKT4) or CD3 (OKT3). The percentages of cells that form symmetrical (top panel) or asymmetrical (bottom panel) ICAM-1 rings are shown for comparison with those observed in gp120 + ICAM-1 bilayers (B). The densities of OKT4 and OKT3 on the bilayer were 250 molecules/µm².