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Fig. 3. Stimulation of erythrocyte annexin-binding by PAF and influence of PAF on cell number. (A) Histograms of FITC-annexin-V-binding as obtained by FACS analysis of a representative experiment of erythrocytes incubated for 24 hours in Ringer solution containing 0.3% ethanol as control (control), in Ringer solution + PAF16 (1 µM PAF16; 3.8 µM PAF16) and in Ringer solution + PAF18 (1 µM PAF18; 3.6 µM PAF18). Numbers depict the percentage of annexin-positive cells. (B) FITC-annexin-V-binding in % of the total population as obtained by FACS analysis of erythrocytes after a 24-hour treatment with Ringer solution containing 0.3% ethanol as control (control) or after incubation with 3.8 µM PAF16 or 3.6 µM PAF18 (arithmetic means±s.e.m., n=8). * indicates significant difference from control (ANOVA, using Dunnett's test as post-hoc test; P $≤$ 0.05). (C) FITC-annexin-V-binding of erythrocytes in % of the total population as obtained by FACS analysis (arithmetic means±s.e.m., n=6-8), treated for 24 hours with different concentrations (0.45-3.8 µM) of PAF16 ( ${blacktriangleup}$ ) or PAF18 ( ${square}$ ). Controls (no PAF) contained appropriate amounts of ethanol. Additionally, erythrocytes were preincubated with 25 µM quinacrine for 3 hours, and PAF16 ( ${blacktriangleup}$ ) was diluted to the appropriate concentrations (125–2000 nM) from 2 mM DMSO stock solutions. FITC-annexin-V-binding in % of the total population as obtained by FACS analysis (arithmetic means ± s.e.m., n=3-5) is depicted in the insert. * indicates significant difference from controls (ANOVA, using Dunnett's test as post-hoc test; P $≤$ 0.05). (D) Number of erythrocytes seeded at 0.3% haematocrit and treated for 24 hours with different concentrations (0.45-7.6 µM) of PAF16 ( ${blacktriangleup}$ ) or PAF18 ( ${square}$ ). The number of erythrocytes (arithmetic means ± s.e.m., n=6-8) after treatment is given in 10⁶ cells/ml. Controls (no PAF) contained appropriate amounts of DMSO vehicle. * indicates significant difference from controls (ANOVA, using Dunnett's test as post-hoc test; P $≤$ 0.05).

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