Fig. 6.

Effect of temperature, human serum, and pretreatment of cytokines or LPS on the loss of AV⁺ cells. Eighteen-hour-aged neutrophils were treated with 0 or 5 μM LL-37 for 5 min. (a) At 37°C or at 4°C, stained with FITC-AV and PI and analyzed by flow cytomtery. Note the absence of AV⁺ cells after treatment with LL-37. Eighteen-hour-aged neutrophils were treated with 0 or 5 μM LL-37 for 5 min. (b) Corrected AV⁻PI⁻ neutrophil counts. (c) Corrected AV⁺PI⁻ neutrophil counts (see Fig. 1 for details of the calculation). Pretreatments at 1 h as shown on x-axis. (a–c) n = 3; **, P < 0.01; ***, P < 0.001, versus untreated control. When the 18-h-aged neutrophils were incubated in different amounts of FBS (0–20%) in the presence or absence of LL-37, there was no inhibition of the conversion of AV⁺PI⁻ to AV⁺PI⁺ cells (d). However, human serum (0–20%) afforded a modest degree of inhibition (e). (d and e) n = 3; *, P < 0.05, versus 0% human serum + LL-37 5 μM.