FIG 5 .

MRP2 and MRP1 promote the secretion of proinflammatory lipids and anti-inflammatory molecules, respectively. (A) Proinflammatory lipids were isolated from apical supernatants of pneumococcus-infected H292 monolayers. HBSS subjected to the apical surface of pneumococcus-infected H292 cells were enriched on C₁₈ columns, which retain the proinflammatory lipids, and eluted in methanol for storage (see Materials and Methods). Yellow triangles represent proinflammatory lipids. (B) Lipids from MRP2-competent (Scrambled control) cells and MRP2-deficient (MRP2 knockdown [MRP2 KD]) cells were enriched and eluted with methanol. This methanol/lipid solution was evaporated under a constant stream of nitrogen and resuspended in HBSS to be used as PMN chemoattractant during a PMN migration assay through a monolayer of naive wild-type H292 cells. (C) Proinflammatory lipids isolated from wild-type cells infected with Streptococcus pneumoniae from panel A were resuspended with either unconditioned media, conditioned media from MRP1-competent scrambled-control cells (Wild-type conditioned media), or conditioned media from MRP1 knockdown cells (MRP1 KD conditioned media) and applied to the apical chamber of naive cells to act as a chemoattractant during a PMN migration assay. HBSS without any lipid acted as a negative control (Buffer Negative control). (D) MRP1-conditioned media failed to inhibit HxA₃-independent PMN-migration produced using formyl-methionyl-leucyl-phenylalanine (fMLP). The values were not significantly different (NS) by two-tailed Student’s t test.