Activation of cytosolic phospholipase A2 and 15-lipoxygenase by oxidized low-density lipoproteins in cultured human lung fibroblasts.

In cell cultures of human lung fibroblasts, we found that oxidized LDL (oxLDL), after 24-h treatment, stimulated arachidonic acid release. A putative role for phospholipases A(2) and MAPK activities in this process was postulated. Consequently, we studied the contribution of either Ca(2+)-dependent, cytosolic phospholipase A(2) (cPLA(2)) or Ca(2+)-independent phospholipase A(2) (iPLA(2)), and the role of the MAP kinase family in oxLDL toxicity to fibroblastic cells in vitro. Activation of extracellular signal-regulated kinases ERK1/2, p38 and c-Jun NH(2)-terminal kinase (JNK) was also assessed with Western blotting. Compared with cellular samples untreated or treated with native LDL, treatment with oxLDL (50-100 microM hydroperoxides) for 24 h significantly increased the levels of either cPLA(2) protein expression or constitutively phosphorylated cPLA(2) protein; in addition we observed enzyme translocation to membranes. iPLA(2) activity was not stimulated by oxLDL. Arachidonic acid release appeared to be associated with phosphorylation of ERK1/2 which was significantly enhanced in a dose-dependent manner whereas no activation of p38 and JNKs was found, indicating that these MAPKs are not involved in mediating the maximal oxLDL response. Western blotting on subcellular fractions and confocal microscopy analyses confirmed an increase in 15-lipoxygenase (15-LO) protein expression and translocation upon activation. A significant increase of cyclooxygenase-2 expression into membrane fraction was also found. Collectively, the data presented link the stimulation of ERK-cPLA(2)-15-LO pathway by oxLDL to the prooxidant mechanism of the lipoprotein complex. It may initially stimulate the fibroblast reaction against the oxidation challenge as well as metabolic repair, such as during lung inflammation and pulmonary fibrosis.