Generation of platelet activating factor (PAF) by a human lung epithelial cell line

Abstract

A human lung epithelial cell line (ATC-CCL-185) was cultured in nutrient Ham-F 12 medium. Cells in monolayers were stimulated with either ionophore A23187 (1 μM) or phorbol myristate acetate (PMA, 0.2 μM) for various periods of time. Samples were analysed by HPLC and the presence of platelet activating factor (PAF) was detected by bioassay of the release of [ 3H]serotonin from rabbit platelets undergoing aggregation. The ATC-CCL 185 cells were found to synthesize PAF following activation with either PMA or ionophore. Ionophore at 1 μM was found to be more potent than PMA at 0.2 μM in the induction of PAF synthesis (≅ 80 ng/mg protein). The synthesis of PAF through ionophore stimulation reached a maximum at 5 min, whereas PMA stimulation peaked at 15–20 min. PMA induced approximately one third the level of PAF synthesis by the ionophore. The PAF synthesized by these CCL185 cells was found to be mainly associated with the cell membrane with less than 10% released into the medium. Release of PAF into cell supernatant was dependent on the presence of bovine serum albumin (BSA). In the absence of BSA, a large portion (≈ 90%) of PAF was found to be cell associated, and only 60% when BSA concentration reached ≥0.2%. These results demonstrate the ability of this lung epithelial cell line to synthesis PAF thus, suggesting that epithelial cells might participate in the process of inflammatory lung diseases, through the generation of this important mediator.