FIGURE 1.

FBS activates cPLA2α and induces LD biogenesis in the presence of BAPTA-AM or U73122. A, Ca²⁺ responses of serum-starved, fura-2/AM-loaded CHO-K1 cells under control conditions (continuous line) or after pretreatment with 45 μm BAPTA-AM (circles) or 10 μm U73122 (triangles) for 30 min before fluorescence measurements were started. At the time indicated, 7.5% FBS was added. B and C, Western blots of serum-starved CHO-K1 pretreated with 45 μm BAPTA-AM or 10 μm U73122 during 30 min and then stimulated with 7.5% FBS for 15 min (B) or 6 h (C). D, cPLA₂ activity measured after the radioactivity was released to the medium of serum-starved CHO-K1 cells that had been prelabeled during 24 h with 0.5 μCi/ml [³H]arachidonic acid, then washed, and treated with 45 μm BAPTA-AM, 10 μm U73122, and/or 10 μm MAFP for 30 min before a 15-min stimulation with 7.5% FBS. E, indirect quantification of LD in serum-starved CHO-K1 cells that had been pretreated for 30 min with 10 μm U73122 and/or MAFP and then stimulated with 7.5% FBS for 6 h. Cells were fixed and stained with Nile red to quantify LD by flow cytometry. Fluorescence intensities in FL1 were quantified as the median values of each event distribution. Results in C and D are means ± S.E. of three independent experiments. *, significantly different (p < 0.01) from controls.