Pretreatment of PBMCs with VLPs inhibit HIV-1 replication without changing in the expression of HIV receptors. (A) PHA-stimulated PBMCs were pretreated with media alone or 1 μg/mL VLPs for 2 or 24 hours and then washed with fresh media to remove unbound VLPs. The pretreated cells were infected with X4 virus for 2 hours and cultured for 7 days without addition of VLPs. Half of the culture supernatants was changed on the 4th day with fresh culture media alone. HIV-1 replication was assessed by p24 antigen capture assay. Data show means ± SDs and are representative of at least 2 independent experiments. (B) PHA-stimulated PBMCs were cultured in the absence (red) or presence (black) of VLPs (1 μg/mL) for 24 hours. The surface expression of CD4, CXCR4, and CCR5 was then assessed by flow cytometry as described in “Materials and methods.” For a positive control of down-regulated receptors, the cells were treated with 100 ng/mL of a mixture of RANTES and MCP-1 for CCR5 or SDF-1 for CXCR4, and then the expression level was analyzed. Left panels show the staining of CD3⁺ T lymphocytes; right panels show CD14⁺ monocytes. Upper, middle, and bottom panels show CD4, CXCR4, and CCR5 staining, respectively. The staining pattern of isotype control antibodies are shown in blue, and the positive control of down-regulation are shown in green. The x-axis and y-axis show florescence intensity and cell count, respectively. The data are representative of 3 independent experiments with similar outcomes.