Figure 3. EP3 is responsible for PGE₂-increased apoptosis.

A. PGE₂-mediated increase in mast cell death evaluated by 7-AAD staining was determined for BMMCs lacking each of the four PGE₂ receptor. Data are from 5 independent experiments using 2 cultures of each genotype. B. PGE₂ mediated increase in apoptosis in mPGES1−/− BMMC. Dead cells numbers were determined by 7-AAD staining. Data are from 3 independent experiments and 1 culture of BMMC per genotype. C. Decreased mitochondrial membrane potential in WT and EP3 −/− BMMC measured as a relative ratio of red and green fluorescence of JC-1 dye. Data are from 5 independent experiments using 1 culture of WT and EP3 −/− BMMC. D. Activity of caspase-3 in WT, EP2, EP3, and EP4 deficient BMMC identified by specific antibody to cleaved p17 caspase-3 fragment. Data are representative of 3 experiments for EP2−/− and EP4−/− and 4 experiments for WT and EP3−/− using 1 culture of each genotype. E. Staining by caspase 3 specific FLICA; example of FACS histogram (upper panel) and its quantitative analysis (lower panel). Data are from 4 independent experiments of 1 culture of WT BMMC and EP3−/−. BMMC were cultured in complete medium (CTRL or +), medium without cytokines (no cyt. or -) and medium without cytokines in the presence 1×10⁻⁶ M PGE₂ (no cyt. + PGE₂, or P) for 16 h. Student's two-tailed t test was used to evaluate statistical differences between cytokine deprive mast cells and cytokine deprived mast cells treated with PGE₂ in A, C, and E. Statistical significance: * = P<0.05, ** = P<0.01. F. Increase in mast cell death after 20 min treatment with various concentration of sulprostone. Cells cultured for 16 h in complete medium (CTRL), medium without cytokines (no cyt.) and stain with 7AAD. Data from 3 independent experiments of 1 culture of WT BMMC.