AMD3100, but not SDF-1α, completely inhibits HIV-induced L-selectin shedding on resting CD4⁺ T cells. CFSE-labeled human CD4⁺ T cells (2 × 10⁶) were treated with medium, 1 μg/mL AMD3100, 10 μg/mL SDF-1α,or10 μg/mL BSA at 37°C for one hour, and cocultured with 2 × 10⁶ HIV-1 NL4-3–infected or mock-infected Jurkat cells at 37°C for 16 to 20 hours. (A) The expression of L-selectin and CD69 on CFSE⁺ cells was determined by flow cytometry. Results shown are representative of 3 experiments. (B) As in Figure 1A, showing the average ± SEM of 3 independent experiments. *P < .05; **P > .05. (C) Dose response of SDF-1α in induction of L-selectin shedding. CFSE-labeled human CD4⁺ T cells were pretreated with media or with varying concentrations of SDF-1α at 37°C for one hour. After washing, human CD4⁺ T cells were mixed with HIV-1 NL4-3–infected or mock-infected Jurkat cells and cocultured at 37°C for 16 to 20 hours. The expression of L-selectin, CXCR4, and CD4 was analyzed by flow cytometry (gated on CFSE⁺ cells). Thick lines represent the expression of L-selectin, CXCR4, and CD4 on human CD4⁺ T cells cocultured with HIV-1 NL4-3–infected Jurkat cells; thin lines represent the expression of L-selectin, CXCR4, and CD4 on CD4⁺ T cells cocultured with mock-infected Jurkat cells; dotted lines represent the isotype control. The apparent decrease in expression of CXCR4 in the presence of AMD3100 is due to binding of AMD3100 to the same epitope as 12G5, the anti-CXCR4 Ab.²¹  (D) SDF-1α does not induce L-selectin shedding. Human CD4⁺ T cells were treated with media (thin line) or 2.0 μg/mL SDF-1α (thick line) at 37°C. The expression of L-selectin, CD69, CXCR4, and CD4 was analyzed after 4 hours or 24 hours of culture by flow cytometry. Dotted lines represent the isotype control. A representative experiment of 3 is shown.