NF-κB signaling in endothelial cells:Identification of NIK as a therapeutic target

Angiogenesis is a major contributor to disease progression in several pathologies including chronic inflammatory disease and cancer. Inflammation is a key factor that drives pathological angiogenesis and signaling through conduits such as the canonical NF-kappaB pathway in endothelial cells have been well established to contribute to this. However, the role of the noncanonical NF-kappaB pathway and its main regulator NF-kappaB inducing kinase (NIK) in these processes was hitherto largely unknown. This thesis delves deeper into the importance of NIK in the regulation of pathological angiogenesis in the context of rheumatoid arthritis (RA), atherosclerosis and colorectal cancer by using novel multicellular in vitro culture models, patient cohorts and various molecular tools modulating the expression of components of the NF-kappaB signaling pathways. Through these studies we have established NIK as a central mediator of lymphotoxin beta receptor (LTβR)-induced synovial neovascularization associated with RA, inflammatory and adverse plaque phenotypes in atherosclerosis, and alternative angiogenic pathways linked to metastatic colorectal cancer. Additionally, it was demonstrated that crosstalk between NIK and canonical NF-kappaB signaling is important for a sustained inflammatory response by endothelial cells. Furthermore, blockade of NIK either by siRNA or a pharmacological NIK inhibitor abrogated neovascularization and inflammatory responses in endothelial cells. Therefore, NIK may be a potential therapeutic target in diseases such as RA, atherosclerosis and (colorectal) cancer

Plasma Protein Signatures of a Murine Venous Thrombosis Model and Slc44a2 Knockout Mice Using Quantitative-Targeted Proteomics

The plasma compartment of the blood holds important information on the risk to develop cardiovascular diseases such as venous thrombosis (VT). Mass spectrometry-based targeted proteomics with internal standards quantifies proteins in multiplex allowing generation of signatures associated with a disease or a condition. Here, to demonstrate the method, we investigate the plasma protein signatures in mice following the onset of VT, which was induced by RNA interference targeting the natural anticoagulants antithrombin and protein C. We then study mice lacking Slc44a2 , which was recently characterized as a VT-susceptibility gene in human genome-wide association studies. We use a recently developed panel of 375 multiplexed mouse protein assays measured by mass spectrometry. A strong plasma protein siganture was observed when VT was induced. Discriminators included acute phase response proteins, and proteins related to erythrocyte function. In mice lacking Slc44a2 , protein signature was primarily overruled by the difference between sexes and not by the absent gene. Upon separate analyses for males and females, we were able to establish a signature for Slc44a2 deficiency, in which glycosylation-dependent cell adhesion molecule-1 and thrombospondin-1 were shared by both sexes. The minimal impact of Slc44a2 deficiency on the measured plasma proteins suggests that the main effect of Slc44a2 on VT does not lay ultimately in the plasma compartment. This suggests further investigation into the role of this VT-susceptibility gene should perhaps also question the possible involvement in cellular mechanisms.Proteomic

NF-kappa B-inducing kinase is a key regulator of inflammation-induced and tumour-associated angiogenesis

Angiogenesis is essential during development and in pathological conditions such as chronic inflammation and cancer progression. Inhibition of angiogenesis by targeting vascular endothelial growth factor (VEGF) blocks disease progression, but most patients eventually develop resistance which may result from compensatory signalling pathways. In endothelial cells (ECs), expression of the pro-angiogenic chemokine CXCL12 is regulated by non-canonical nuclear factor (NF)-κB signalling. Here, we report that NF-κB-inducing kinase (NIK) and subsequent non-canonical NF-κB signalling regulate both inflammation-induced and tumour-associated angiogenesis. NIK is highly expressed in endothelial cells (ECs) in tumour tissues and inflamed rheumatoid arthritis synovial tissue. Furthermore, non-canonical NF-κB signalling in human microvascular ECs significantly enhanced vascular tube formation, which was completely blocked by siRNA targeting NIK. Interestingly, Nik(−/−) mice exhibited normal angiogenesis during development and unaltered TNFα- or VEGF-induced angiogenic responses, whereas angiogenesis induced by non-canonical NF-κB stimuli was significantly reduced. In addition, angiogenesis in experimental arthritis and a murine tumour model was severely impaired in these mice. These studies provide evidence for a role of non-canonical NF-κB signalling in pathological angiogenesis, and identify NIK as a potential therapeutic target in chronic inflammatory diseases and tumour neoangiogenesis. © 2014 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland

Characterization of hemostasis in mice lacking the novel thrombosis susceptibility gene Slc44a2

Thrombosis and Hemostasi