Figure 4.

Effects of 11-keto-BAs on the formation of 5-LO metabolites. (a) Freshly isolated PMNL (5 × 10⁶) were resuspended in 1 ml PG buffer containing 1 mM EDTA and preincubated with 30 μM BAPTA/AM for 15 min. Then, cells were stimulated with 20 μM AA alone or together with the indicated BAs (30 μM, each). After 10 min at 37°C, 5-LO products were determined by HPLC. Results are given as mean+s.e., n=4–6. Student's t-test; ^**P<0.01. (b) Dose–response curve of AKBA. PMNL (5 × 10⁶) were resuspended in 1 ml PG buffer containing 1 mM EDTA and preincubated with 30 μM BAPTA/AM for 15 min. Then, cells were stimulated with 20 μM AA alone or together with the indicated amounts of AKBA. After 10 min at 37°C, 5-LO products were determined. Results are given as mean+s.e., n=4–5. (c) Inhibition of 5-LO. PMNL (5 × 10⁶) were resuspended in either 1 ml PGC buffer or PG buffer containing 1 mM EDTA and 30 μM BAPTA/AM. After 15 min at 37°C, AKBA was added, and cells were incubated for another 30 min at 37°C. Then, CaCl₂ was adjusted to 1 mM in all incubations and cells were immediately stimulated with 2.5 μM ionophore and 40 μM AA. After 10 min, 5-LO products were determined. Results are given as mean+s.e., n=3.