Oxidized LDL induced extracellular trap formation in human neutrophils via TLR-PKC-IRAK-MAPK and NADPH-oxidase activation

• Published in: Free radical biology & medicine Vol. 93; pp. 190 - 203
• Main Authors: Awasthi, Deepika, Nagarkoti, Sheela, Kumar, Amit, Dubey, Megha, Singh, Abhishek Kumar, Pathak, Priya, Chandra, Tulika, Barthwal, Manoj Kumar, Dikshit, Madhu
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01-04-2016
• Subjects: Aldehydes - pharmacology; Extracellular Traps - genetics; Extracellular Traps - metabolism; Humans; Interleukin-1 Receptor-Associated Kinases - genetics; Interleukin-1 Receptor-Associated Kinases - metabolism; Lipoproteins, LDL - metabolism; Lipoproteins, LDL - pharmacology; Malondialdehyde - pharmacology; NADPH oxidase; NADPH Oxidases - genetics; NADPH Oxidases - metabolism; Neutrophil extracellular traps; Neutrophils - metabolism; Neutrophils - pathology; Oxidation-Reduction - drug effects; Oxidized low density lipoprotein; Oxidized phospholipids; Protein Kinase C - metabolism; Reactive Oxygen Species - metabolism; Signal Transduction - drug effects; Systemic inflammatory response syndrome; Systemic Inflammatory Response Syndrome - genetics; Systemic Inflammatory Response Syndrome - metabolism; Systemic Inflammatory Response Syndrome - pathology; Toll-Like Receptor 2 - genetics; Toll-Like Receptor 2 - metabolism; DHE; PAPC; TNFα; PMNs; PFA; MDA; MPO; DFP; MNase; BSA; HDL; Systemic inflammatory response syndrome; 4-HNE; LDL; ABAH; RNS; NADPH oxidase; ApoB; PBS; CD; Oxidized low density lipoprotein; OxPAPC; Oxidized phospholipids; PMA; DAPI; PKC; IRAKs; IL-1β; IL-8; MAPK; Mcl-1; NAC; SDS-PAGE; SIRS; NETs; NOX; Neutrophil extracellular traps; ROS; DCF-DA; OxLDL; DPI; LPC; TLRs
• ISSN: 0891-5849; 1873-4596
• DOI: 10.1016/j.freeradbiomed.2016.01.004

Neutrophil extracellular traps (NETs) formation was initially linked with host defence and extracellular killing of pathogens. However, recent studies have highlighted their inflammatory potential. Oxidized low density lipoprotein (oxLDL) has been implicated as an independent risk factor in various acute or chronic inflammatory diseases including systemic inflammatory response syndrome (SIRS). In the present study we investigated effect of oxLDL on NETs formation and elucidated the underlying signalling mechanism. Treatment of oxLDL to adhered PMNs led to a time and concentration dependent ROS generation and NETs formation. OxLDL induced free radical formation and NETs release were significantly prevented in presence of NADPH oxidase (NOX) inhibitors suggesting role of NOX activation in oxLDL induced NETs release. Blocking of both toll like receptor (TLR)-2 and 6 significantly reduced oxLDL induced NETs formation indicating requirement of both the receptors. We further identified Protein kinase C (PKC), Interleukin-1 receptor associated kinase (IRAKs), mitogen-activated protein kinase (MAPK) pathway as downstream intracellular signalling mediators involved in oxLDL induced NETs formation. OxLDL components such as oxidized phospholipids (lysophosphatidylcholine (LPC) and oxidized 1-palmitoyl-2-arachidonyl-sn-glycero-3-phosphorylcholine (oxPAPC)) were most potent NETs inducers and might be crucial for oxLDL mediating NETs release. Other components like, oxysterols, malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE) were however less potent as compared to oxidized phospholipids. This study thus demonstrates for the first time that treatment of human PMNs with oxLDL or its various oxidized phopholipid component mediated NETs release, implying their role in the pathogenesis of inflammatory diseases such as SIRS.
OxLDL induced signalling pathway leading to NETosis in human PMNs.
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•OxLDL mediated the release of neutrophil extracellular traps (NETs) from human PMNs.•OxLDL induced ROS generation and NETs formation was dependent on NADPH oxidase.•TLR-PKC-IRAK-MAPK pathway was found to be involved in oxLDL induced NETs formation.•Oxidized phospholipids component of oxLDL was crucial and most potent NETs inducer.