The hydrogen-peroxide producing NADPH oxidase 4 does not limit neointima development after vascular injury in mice

OBJECTIVE: The NADPH oxidase Nox4 is an important source of H(2)O(2). Nox4-derived H(2)O(2) limits vascular inflammation and promotes smooth muscle differentiation. On this basis, the role of Nox4 for restenosis development was determined in the mouse carotid artery injury model. METHODS AND RESULTS: Genetic deletion of Nox4 by a tamoxifen-activated Cre-Lox-system did not impact on neointima formation in the carotid artery wire injury model. To understand this unexpected finding, time-resolved single-cell RNA-sequencing (scRNAseq) from injured carotid arteries of control mice and massive-analysis-of-cDNA-ends (MACE)-RNAseq from the neointima harvested by laser capture microdissection of control and Nox4 knockout mice was performed. This revealed that resting smooth muscle cells (SMCs) and fibroblasts exhibit high Nox4 expression, but that the proliferating de-differentiated SMCs, which give rise to the neointima, have low Nox4 expression. In line with this, the first weeks after injury, gene expression was unchanged between the carotid artery neointimas of control and Nox4 knockout mice. CONCLUSION: Upon vascular injury, Nox4 expression is transiently lost in the cells which comprise the neointima. NADPH oxidase 4 therefore does not interfere with restenosis development after wire-induced vascular injury

The polarity protein Scrib limits atherosclerosis development in mice

AIMS: The protein Scrib (Scribble 1) is known to control apico-basal polarity in epithelial cells. The role of polarity proteins in the vascular system remains poorly characterized; however, we previously reported that Scrib maintains the endothelial phenotype and directed migration. On this basis, we hypothesized that Scrib has anti-atherosclerotic functions. METHODS AND RESULTS: Tamoxifen-induced Scrib-knockout mice were crossed with ApoE-/- knockout mice and spontaneous atherosclerosis under high-fat diet, as well as accelerated atherosclerosis in response to partial carotid artery ligation and high-fat diet, was induced. Deletion of Scrib resulted in increased atherosclerosis development in both models. Mechanistically, flow- as well as acetylcholine-induced endothelium-dependent relaxation and AKT phosphorylation was reduced by deletion of Scrib, whereas vascular permeability and leukocyte extravasation were increased after Scrib knockout. Scrib immune pull down in primary carotid endothelial cells and mass spectrometry identified Arhgef7 (Rho Guanine Nucleotide Exchange Factor 7, βPix) as interaction partner. Scrib or Arhgef7 downregulation by siRNA reduced the endothelial barrier function in HUVEC. Gene expression analysis from murine samples and from human biobank material of carotid endarterectomies indicated that loss of Scrib resulted in endothelial dedifferentiation with a decreased expression of endothelial signature genes. CONCLUSIONS: By maintaining a quiescent endothelial phenotype, the polarity protein Scrib elicits antiatherosclerotic functions

Country report : Spain

First published in September 2009; Revised May 2010Research for this EUDO Citizenship Observatory Country Report has been jointly supported by the European Commission grant agreement JLS/2007/IP/CA/009 EUCITAC and by the British Academy Research Project CITMODES (both projects co-directed by the EUI and the University of Edinburgh)