Suppression of HIV-1 by SIL correlates with inhibition of PBMC and CEM growth.

<p><b>A</b>, SIL suppresses growth of PBMC and CEM cells. Cells were treated with SIL as described in the Materials and Methods and cells were counted 24 hours later by hemacytometer and trypan blue. <b>B</b>, Suppression of PHA-stimulated PBMC cell growth. After 24 hours of PHA stimulation, PBMC were placed in culture for 24 or 96 hours and the concentration of live cells per µL of sample was measured by flow cytometry. <b>C</b>, Suppression of CD4+ T cell proliferation measured by CFSE staining. PBMCs were labeled with CFSE and stimulated with PHA (2 µg/ml) for 24 hours prior to incubation with SIL for 4 days. <b>D</b>, SIL suppresses growth of T Cells and B Cells. Three day PHA-stimulated PBMC were treated with or without SIL for 24 hours and analyzed by flow cytometry following staining of T (CD4 or CD8) and B (CD20) cells and labeling of DNA using the Click-iT® EdU labeling kit. Peaks on the left of each graph represent EdU-, non-proliferating cells, while peaks on the right represent EdU+ proliferating cells. Top two rows depict unstimulated PBMC with and without SIL. Bottom two rows depict 72 <i>h</i> PHA and 24 <i>h</i> IL-2 stimulated PBMC with and without SIL. <b>E</b>, the number of EdU+ proliferating cells is inhibited by SIL. Cells were treated as described for panel C following exposure to SIL. The cell concentration is expressed per µl of cell suspension and is determined using counting beads as for panel B. <b>F, G</b>, SIL is not cytotoxic to PBMCs. PBMCs were stimulated with PHA for 3 days and treated with IL-2 in the presence of varying doses of SIL and 24 hours later, stained for CD4+, CD8+ and CD19+ (B cell) immune cell subsets and dead cells (<b>F</b>) as well as Annexin V+ apoptotic cells (<b>G</b>) were quantified.