Effect of α-defensin-2 on cell-surface protein expression: induction of CD4 downmodulation. Flow cytometry analysis of the surface expression of CD2, CD4 (Leu3a), CD26, CD45RO, CD46, HLA-DR, CCR5 (2D7), and CXCR4 (44717.111) in PM1 cells. The cells were preincubated for 60 minutes with or without an equimolar mixture of α-defensins 1 and 2 (total concentration, 14.7 μM) in the absence of serum; then the appropriate mAbs were added for cell-surface staining. Solid histograms represent the expression of the indicated markers; empty profiles indicate the background fluorescence signal obtained using irrelevant, isotype-matched mAbs. The numbers denote the MFI for the indicated marker. The histograms shown are representative of 4 to 6 independent experiments with similar results. (B) Time-course analysis of the effect of α-defensin-2 on binding of the anti-CD4 mAb Leu3a and OKT4 to live and formaldehyde-fixed PM1 cells. The cells were incubated without (empty bars) or with α-defensin-2 (14.7 μM) for 20 minutes (light gray bars) or 120 minutes (dark gray bars) and then stained with the mAbs. Fixed cells (treated with 2% paraformaldehyde at 4°C for 30 minutes) were washed twice and incubated for 120 minutes with α-defensins before antibody staining (black bars). Error bars indicate SD of mean values obtained from 3 repeated assays.