TRIF adaptor signaling is important in abdominal aortic aneurysm formation

Objective: Abdominal aortic aneurysm (AAA) is characterized by inflammation, loss of smooth muscle cells (SMCs), and degradation of the extracellular matrix in the vessel wall. Innate immune receptors such as Toll-like receptors (TLRs) were recently shown to regulate immunological processes leading to the formation and progression of atherosclerotic plaques as well as to other cardiovascular pathologies. Our aim was to investigate whether blockage of TLR signaling, under the control of TIR domain-containing adaptor protein including IFN-beta (TRIF), could inhibit the inflammatory response and AAA development in mice. Results: In human AAA, an increased TLR3 and TLR4 expression in association with macrophages and T lymphocytes was demonstrated with immunohistochemical analysis. Angiotensin (Ang) II-induced aneurysm formation was significantly reduced by 30% in ApoE(-/-)Trif(-/-) mice compared to ApoE(-/-) mice. Morphologically, AngII-infused ApoE(-/-)Trif(-/-) mice had a more intact cellular and extracellular matrix while ApoE(-/-) mice infused with AngII displayed an increased medial thickness associated with aortic dissection, thrombus formation, and a more disorganized vessel wall. Gene expression analysis of the abdominal aorta revealed a profound decrease of the inflammatory genes CD68 (P <0.05), CD11b (P <0.05), and TNF-alpha (P <0.05) and the protease gene MMP-12 (P <0.01) in ApoE(-/-)Trif(-/-) mice compared to ApoE(-/-) mice infused with AngII. Conclusion: Our results suggest that signaling through TRIF is important for the inflammatory response of AngII-induced AAA and that blockage of the TRIF pathway reduces vascular inflammation and protects against AAA formation. (C) 2015 The Authors. Published by Elsevier Ireland Ltd.Peer reviewe

Desquamation of human coronary artery endothelium by human mast cell proteases: implications for plaque erosion

Objective: endothelial erosion has emerged as an important contributor to the pathogenesis of atherosclerosis and its complications, but the molecular mechanisms have remained unclear. As activated mast cells capable of secreting neutral proteases are present in the intima of eroded coronary plaques, we investigated their potential roles in endothelial erosion. Methods and results: studies involving double immunostaining of mast cells (tryptase(pos) cells) and platelets (CD42b) in human coronary artery specimens indicated that the number of subendothelial mast cells correlated with the number of parietal microthrombi (P=0.001, rs 0.27). The number of parietal microthrombi was significantly higher (P&lt;0.001) in areas of plaques than in areas of healthy intima. Of the microthrombi 86% were in the lesional coronary segments, of all subendothelial mast cells 15% were located under parietal microthrombi, and of all parietal microthrombi 49% were located over subendothelial mast cells. Double immunostaining revealed the mast cell to neutrophil ratio in the human coronary artery intima to be 5 : 1, and that mast cells are a major local source of cathepsin G. Scanning electron and light microscopy indicated that treatment of fresh human coronary arteries intraluminally with recombinant human (rh)-tryptase and rh-chymase induced endothelial damage characterized by retraction of endothelial cells, disruption of endothelial cell to cell adhesions and desquamation of endothelial cells. VE-cadherin and fibronectin, which are necessary for cell-cell interactions and endothelial cell adhesion, were degraded by tryptase and chymase and also by cathepsin G.Conclusions: activated subendothelial mast cells may contribute to endothelial erosion by releasing proteases capable of degrading VE-cadherin and fibronectin

OSBP-related protein 11 (ORP11) dimerizes with ORP9 and localizes at the Golgi-late endosome interface

We characterize here ORP11, a member of the oxysterol-binding protein family. ORP11 is present at highest levels in human ovary, testis, kidney, liver, stomach, brain, and adipose tissue. Immunohistochemistry demonstrates abundant ORP11 in the epithelial cells of kidney tubules, testicular tubules, caecum, and skin. ORP11 in HEK293 cells resides on Golgi complex and LE, co-localizing with GFP-Rab9, TGN46, GFP-Rab7, and a fluorescent medial-trans-Golgi marker. Under electron microscopic observation, cells overexpressing ORP11 displayed lamellar lipid bodies associated with vacuolar structures or the Golgi complex, indicating a disturbance of lipid trafficking. N-terminal fragment of ORP11 (aa 1–292) localized partially to Golgi, but displayed enhanced localization on Rab7- and Rab9-positive LE, while the C-terminal ligand-binding domain (aa 273–747) was cytosolic, demonstrating that the membrane targeting determinants are N-terminal. Yeast two-hybrid screen revealed interaction of ORP11 with the related ORP9. The interacting region was delineated within aa 98–372 of ORP9 and aa 154–292 of ORP11. Overexpressed ORP9 was able to recruit EGFP-ORP11 to membranes, and ORP9 silencing inhibited ORP11 Golgi association. The results identify ORP11 as an OSBP homologue distributing at the Golgi–LE interface and define the ORP9-ORP11 dimer as a functional unit that may act as an intracellular lipid sensor or transporter

Complement regulation in human atherosclerotic coronary lesions - Immunohistochemical evidence that C4b-binding protein negatively regulates the classical complement pathway, and that C5b-9 is formed via the alternative complement pathway

Objective: The complement system is activated in human atherosclerotic lesions and may hence aggravate local inflammation. We studied the presence and localization of C4b-binding protein (C4bp), the major inhibitor of the classical complement pathway, in human atherosclerotic lesions in relation to complement activation products and protein S, which circulates in complex with C4bp. Methods and results: Immunohistochemistry, of human coronary arteries showed C4bp to be virtually absent in normal arteries but present in early and advanced atherosclerotic lesions. In the lesions, C4bp is associated with proteoglycans, and affinity chromatography showed that C4bp interacts with human arterial proteoglycans. Areas containing C4bp also contained IgM and C4 suggesting that C4bp is involved in the regulation of the classical complement pathway. However, C5b-9 was virtually absent in these areas but, instead, colocalized with properdin deeper in the intima, suggesting that C5b-9 is formed by the alternative complement pathway. A fraction of C4bp was associated with protein S and apoptotic cells. Conclusions: The results indicate that C4bp regulates the classical complement pathway in human atherosclerotic lesions. Thus, unlike the alternative pathway, the classical complement pathway does not generate C5b-9, but is likely to be involved in the clean-up of apoptotic cells and cell debris in the arterial intima. (c) 2006 Elsevier Ireland Ltd. All rights reserved