Inflammatory intracellular pathways activated by electronegative LDL in monocytes

Abstract Aims Electronegative LDL (LDL(−)) is a plasma LDL subfraction that induces cytokine release in monocytes through toll-like receptor 4 (TLR4) activation. However, the intracellular pathways induced by LDL(−) downstream TLR4 activation are unknown. We aimed to identify the pathways activated by LDL(−) leading to cytokine release in monocytes. Methods and results We determined LDL(−)-induced activation of several intracellular kinases in protein extracts from monocytes using a multikinase ELISA array. LDL(−) induced higher p38 mitogen-activated protein kinase (MAPK) phosphorylation than native LDL. This was corroborated by a specific cell-based assay and it was dependent on TLR4 and phosphoinositide 3-kinase (PI3k)/Akt pathway. P38 MAPK activation was involved in cytokine release promoted by LDL(−) . A specific ELISA showed that LDL(−) activated cAMP response-element binding (CREB) in a p38 MAPK dependent manner. P38 MAPK was also involved in the nuclear factor kappa-B (NF-kB) and activating protein-1 (AP-1) activation by LDL(−). We found that NF-kB, AP-1 and CREB inhibitors decreased LDL(−)-induced cytokine release, mainly on MCP1, IL6 and IL10 release, respectively. Conclusions LDL(−) promotes p38 MAPK phosphorylation through TLR4 and PI3k/Akt pathways. Phosphorylation of p38 MAPK is involved in NF-kB, AP-1 and CREB activation, leading to LDL(−)-induced cytokine release in monocytes.