Role of NETs in athero-thrombosis in LNK/SH2B3 deficient mice

GWAS have revealed an association of a common SNP(R262W) in LNK/SH2B3 with leukocytosis and thrombocytosis and with coronary artery disease (CAD). Our lab has shown that LNK(R262W) reduces LNK function and that hematopoietic LNK deficiency in hypercholesterolemic (HC) mice promotes platelet activation, neutrophilia and athero-thrombosis. However, the underlying mechanisms are incompletely understood. Recent evidence suggest that neutrophil extracellular traps (NETs) promote atherosclerotic plaque instability, as well as endothelial cell (EC) injury and thrombosis in a model of superficial erosion. My preliminary data shows that NETs are increased in atherosclerotic lesions of HC hematopoietic Lnk-/- mice and that HC-Lnk-/- mice show accelerated arterial thrombosis which is completely reversed by hematopoietic deficiency of Lnk and Pad4, an enzyme critical for NETosis. Mechanistic studies reveal that activated Lnk-/- platelets promote NETosis when incubated with Lnk-/- neutrophils, likely as a result of augmented release of oxidized phospholipids (oxPL). My central hypothesis is that LNK deficiency synergizes with HC to promote NETosis via an oxPL activated pathway, leading to accelerated athero-thrombosis. Aim 1 will assess the impact of NETs in atherogenesis. WT, Lnk-/-, Pad4-/- and Lnk-/-Pad4-/- bone marrow will be transplanted into Ldlr-/- recipients. Similar mice will also be used to assess plaque erosion and thrombosis, in collaborative studies with Dr Libby. Aim 2 will examine the impact of oxPL on NETosis and athero-thrombosis, by transplantation of bone marrow from WT or Lnk-/- mice into Ldlr-/- expressing oxPL-neutralizing antibodies i.e. E06-scFv transgenic mice, in collaboration with Dr Witztum. These studies will define the role of NETosis in athero-thrombosis using novel mouse models relevant to a common human genetic variant that increased CAD risk, and may suggest new therapeutic approaches to preventing CAD in genetically defined human populations. (AHA Program: Postdoctoral Fellowship)