The Yorkie homologues YAP (Yes-associated protein) and TAZ (transcriptional coactivator with PDZ-binding motif, also known as WWTR1), effectors of the Hippo pathway, have been identified as mediators for mechanical stimuli. However, the role of YAP/TAZ in haemodynamics-induced mechanotransduction and pathogenesis of atherosclerosis remains unclear. Here we show that endothelial YAP/TAZ activity is regulated by different patterns of blood flow, and YAP/TAZ inhibition suppresses inflammation and retards atherogenesis. Atheroprone-disturbed flow increases whereas atheroprotective unidirectional shear stress inhibits YAP/TAZ activity. Unidirectional shear stress activates integrin and promotes integrin-Gα interaction, leading to RhoA inhibition and YAP phosphorylation and suppression. YAP/TAZ inhibition suppresses JNK signalling and downregulates pro-inflammatory genes expression, thereby reducing monocyte attachment and infiltration. In vivo endothelial-specific YAP overexpression exacerbates, while CRISPR/Cas9-mediated Yap knockdown in endothelium retards, plaque formation in ApoE mice. We also show several existing anti-atherosclerotic agents such as statins inhibit YAP/TAZ transactivation. On the other hand, simvastatin fails to suppress constitutively active YAP/TAZ-induced pro-inflammatory gene expression in endothelial cells, indicating that YAP/TAZ inhibition could contribute to the anti-inflammatory effect of simvastatin. Furthermore, activation of integrin by oral administration of MnCl reduces plaque formation. Taken together, our results indicate that integrin-Gα-RhoA-YAP pathway holds promise as a novel drug target against atherosclerosis.

Genome-wide association studies (GWAS) have identified a genetic variant at a locus on chromosome 1p13that is associated with reduced risk of myocardial infarction, reduced plasma levels of LDL cholesterol (LDL-C), and markedly increased expression of the gene sortilin-1 (SORT1) in liver. Sortilin is a lysosomal sorting protein that binds ligands both in the Golgi apparatus and at the plasma membrane and traffics them to the lysosome. We previously reported that increased hepatic sortilin expression in mice reduced plasma LDL-C levels. Here we show that increased hepatic sortilin not only reduced hepatic apolipoprotein B (APOB) secretion, but also increased LDL catabolism, and that both effects were dependent on intact lysosomal targeting. Loss-of-function studies demonstrated that sortilin serves as a bona fide receptor for LDL in vivo in mice. Our data are consistent with a model in which increased hepatic sortilin binds intracellular APOB-containing particles in the Golgi apparatus as well as extracellular LDL at the plasma membrane and traffics them to the lysosome for degradation. We thus provide functional evidence that genetically increased hepatic sortilin expression both reduces hepatic APOB secretion and increases LDL catabolism, providing dual mechanisms for the very strong association between increased hepatic sortilin expression and reduced plasma LDL-C levels in humans.

Pathophysiological cardiac hypertrophy is one of the most common causes of heart failure. Epoxyeicosatrienoic acids, hydrolyzed and degraded by soluble epoxide hydrolase (sEH), can function as endothelium-derived hyperpolarizing factors to induce dilation of coronary arteries and thus are cardioprotective. In this study, we investigated the role of sEH in two rodent models of angiotensin II (Ang II)-induced cardiac hypertrophy. The protein level of sEH was elevated in the heart of both spontaneously hypertensive rats and Ang II-infused Wistar rats. Blocking the Ang II type 1 receptor with losartan could abolish this induction. Administration of a potent sEH inhibitor (sEHI) prevented the pathogenesis of the Ang II-induced hypertrophy, as demonstrated by decreased leftventricular hypertrophy assessed by echocardiography, reduced cardiomyocyte size, and attenuated expression of hypertrophy markers, including atrial natriuretic factor and ␤-myosin heavy chain. Because sEH elevation was not observed in exercise-or norepinephrine-induced hypertrophy, the sEH induction was closely associated with Ang II-induced hypertrophy. In vitro, Ang II upregulated sEH and hypertrophy markers in neonatal cardiomyocytes isolated from rat and mouse. Expression of these marker genes was elevated with adenovirus-mediated sEH overexpression but decreased with sEHI treatment. These results were supported by studies in neonatal cardiomyocytes from sEH ؊/؊ mice. Our results suggest that sEH is specifically upregulated by Ang II, which directly mediates Ang II-induced cardiac hypertrophy. Thus, pharmacological inhibition of sEH would be a useful approach to prevent and treat Ang II-induced cardiac hypertrophy. epoxyeicosatrienoic acid ͉ cardiomyocyte ͉ activator protein 1

endothelial cells ͉ arachidonic acid ͉ AP-1 ͉ promoter ͉ hypertension A rachidonic acid (AA) derived from membrane phospholipids plays a key role in vascular inflammatory and/or antiinflammatory responses. AA can be converted to eicosanoids by three major enzymatic pathways, namely, cyclooxygenase, lipoxygenase, and CYP 450 epoxygenase. Exerting autocrine effects on vascular endothelial cells (ECs), four epoxyeicosatrienoic acids (EETs) regioisomers 5,6-, 8,9-, 11,12-, and 14,15-EET are the major metabolites generated by CYP 450 epoxygenase (1). EETs can be released by ECs to act as paracrine mediators on neighboring cells such as vascular smooth muscle cells (VSMCs) (2). EETs exert membrane potential-independent effects and modulate several signaling cascades that affect EC proliferation and angiogenesis. EETs also function as endothelium-derived hyperpolarizing factors (3). By increasing intracellular Ca 2ϩ concentration, EETs activate large conductance Ca 2ϩ -activated K ϩ channel (BK Ca ) in the smooth muscle. The activation of BK Ca then causes hyperpolarization of VSMCs and subsequent vasodilation, which lowers the blood pressure (4). As well, EETs inhibit cytokine-induced inflammatory responses in ECs (5, 6). Treating ECs with 11,12-EET or overexpression of CYP2J2 attenuated the TNF␣-, IL-1␣-, and LPSinduced expression of adhesion molecules in ECs, thus decreasing leukocyte adhesion to the vascular wall (7).Epoxide hydrolases (EHs) convert epoxides to the corresponding diols. Under physiological conditions, EETs can be enzymatically hydrolysed to dihydroxyeicosatrienoic acids (DHETs) by EHs (1). Two major EHs in the ␣/␤ hydrolase family exist in mammalian cells: soluble EH (sEH), which primarily presents in the cytosol and peroxisomes, and microsomal EH, which binds to the intracellular membranes (8). Highly expressed in the liver, kidney, intestine, and vasculature, sEH is the main enzyme that converts 5,8,11,14,8,11,14, respectively. The mammalian sEH is a homodimer, and each subunit contains a C-and an N-terminal domain. The active site is located in the C-terminal domain in which the residues Asp-333, Asp-495, and His-523 form the catalytic triad (9). DHETs are much more polar than EETs and are generally considered as biologically inactive products of EETs. However, their roles are not fully understood.Angiotensin II (Ang II), a potent vessel constrictor, elevates blood pressure in various animal models. i.p. injection of sEHselective inhibitors to Ang II-infused hypertensive rats greatly increased the level of EETs and lowered systolic blood pressure (10). Thus, augmentation of EET levels with enhanced production by CYP450s or decreased hydrolysis by sEH seems to control blood pressure in vivo. In line with this hypothesis, recent studies demonstrated that the selective sEH inhibitor Ncyclohexyl-N-dodecyl urea reversed the hypertensive phenotype in the spontaneously hypertensive rat (SHR) (11).We have previously shown that laminar shear stress, an atheroprotective flow, decreased the expression of sE...

Rationale: The mechanisms driving athero-thrombotic risk in individuals with JAK2V617F (Jak2VF) positive clonal hematopoiesis (CH) or myeloproliferative neoplasms (MPN) are poorly understood. Objective: The goal of this study was to assess atherosclerosis and underlying mechanisms in hypercholesterolemic mice with hematopoietic Jak2VF expression. Methods and Results: Irradiated low-density lipoprotein receptor knockout (Ldlr−/−) mice were transplanted with bone marrow from WT or Jak2VF mice and fed a high fat high cholesterol Western diet (WD). Hematopoietic functions and atherosclerosis were characterized. After 7 weeks of WD Jak2VF mice showed increased atherosclerosis. Early atherosclerotic lesions showed increased neutrophil adhesion and content, correlating with lesion size. After 12 weeks of WD Jak2VF lesions showed increased complexity, with larger necrotic cores, defective efferocytosis, prominent iron deposition and co-staining of erythrocytes and macrophages suggesting erythrophagocytosis. Jak2VF erythrocytes were more susceptible to phagocytosis by WT macrophages and showed decreased surface expression of CD47, a “don’t eat me” signal. Human JAK2VF erythrocytes were also more susceptible to erythrophagocytosis. Jak2VF macrophages displayed increased expression and production of pro-inflammatory cytokines and chemokines, prominent inflammasome activation, increased p38 MAP kinase signaling and reduced levels of MerTK, a key molecule mediating efferocytosis. Increased erythrophagocytosis also suppressed efferocytosis. Conclusions: Hematopoietic Jak2VF expression promotes early lesion formation and increased complexity in advanced atherosclerosis. In addition to increasing hematopoiesis and neutrophil infiltration in early lesions, Jak2VF caused cellular defects in erythrocytes and macrophages, leading to increased erythrophagocytosis but defective efferocytosis. These changes promote accumulation of iron in plaques and increased necrotic core formation which, together with exacerbated pro-inflammatory responses, likely contribute to plaque instability.