Impaired high-density lipoprotein function and endothelial barrier stability in severe anaphylaxis.

BACKGROUND: Growing evidence demonstrates the importance of high- and low-density lipoprotein cholesterol in certain immune and allergy-mediated diseases. OBJECTIVE: This study aimed to evaluate levels of high- and low-density lipoprotein cholesterol and apolipoproteins A1 and B in sera from a cohort of patients presenting with hypersensitivity reactions. We further assessed the function of high-density lipoprotein particles as well as their involvement in the molecular mechanisms of anaphylaxis. METHODS: Lipid profile determination was performed in paired (acute and baseline) serum samples from 153 patients. Thirty-eight experienced a non-anaphylactic reaction and 115 had an anaphylactic reaction (88 moderate and 27 severe). Lecithin cholesterol acyl transferase activity was assessed in patient sera, and we also evaluated macrophage cholesterol efflux in response to the serum samples. Last, the effect of anaphylactic-derived high-density lipoprotein (HDL) particles on the endothelial barrier was studied. Detailed methods are provided in the Methods section in this article's Online Repository available at www.jacionline.org. RESULTS: Serum samples from severe anaphylactic reactions show statistically significant low levels of HDL cholesterol, low-density lipoprotein cholesterol, and apolipoproteins A1 and B, which points to their possible role as biomarkers. Specifically, HDL particles play a protective role in cardiovascular diseases. Using functional human serum cell assays, we observed impaired capacity of apolipoprotein B-depleted serum to induce macrophage cholesterol efflux in severe anaphylactic reactions. In addition, purified HDL particles from human anaphylactic sera failed to stabilize and maintain the endothelial barrier. CONCLUSION: These results encourage further research on HDL functions in severe anaphylaxis, which may lead to new diagnostic and therapeutic strategies.

Bone Marrow-Derived RIPK3 Mediates Kidney Inflammation in Acute Kidney Injury.

BACKGROUND: Receptor-interacting protein kinase 3 (RIPK3), a component of necroptosis pathways, may have an independent role in inflammation. It has been unclear which RIPK3-expressing cells are responsible for the anti-inflammatory effect of overall Ripk3 deficiency and whether Ripk3 deficiency protects against kidney inflammation occurring in the absence of tubular cell death. METHODS: We used chimeric mice with bone marrow from wild-type and Ripk3-knockout mice to explore RIPK3's contribution to kidney inflammation in the presence of folic acid-induced acute kidney injury AKI (FA-AKI) or absence of AKI and kidney cell death (as seen in systemic administration of the cytokine TNF-like weak inducer of apoptosis [TWEAK]). RESULTS: Tubular and interstitial cell RIPK3 expressions were increased in murine AKI. Ripk3 deficiency decreased NF-κB activation and kidney inflammation in FA-AKI but did not prevent kidney failure. In the chimeric mice, RIPK3-expressing bone marrow-derived cells were required for early inflammation in FA-AKI. The NLRP3 inflammasome was not involved in RIPK3's proinflammatory effect. Systemic TWEAK administration induced kidney inflammation in wild-type but not Ripk3-deficient mice. In cell cultures, TWEAK increased RIPK3 expression in bone marrow-derived macrophages and tubular cells. RIPK3 mediated TWEAK-induced NF-κB activation and inflammatory responses in bone marrow-derived macrophages and dendritic cells and in Jurkat T cells; however, in tubular cells, RIPK3 mediated only TWEAK-induced Il-6 expression. Furthermore, conditioned media from TWEAK-exposed wild-type macrophages, but not from Ripk3-deficient macrophages, promoted proinflammatory responses in cultured tubular cells. CONCLUSIONS: RIPK3 mediates kidney inflammation independently from tubular cell death. Specific targeting of bone marrow-derived RIPK3 may limit kidney inflammation without the potential adverse effects of systemic RIPK3 targeting.

Mitochondrial Dysfunction in the Cardio-Renal Axis.

Cardiovascular disease (CVD) frequently complicates chronic kidney disease (CKD). The risk of all-cause mortality increases from 20% to 500% in patients who suffer both conditions; this is referred to as the so-called cardio-renal syndrome (CRS). Preclinical studies have described the key role of mitochondrial dysfunction in cardiovascular and renal diseases, suggesting that maintaining mitochondrial homeostasis is a promising therapeutic strategy for CRS. In this review, we explore the malfunction of mitochondrial homeostasis (mitochondrial biogenesis, dynamics, oxidative stress, and mitophagy) and how it contributes to the development and progression of the main vascular pathologies that could be affected by kidney injury and vice versa, and how this knowledge may guide the development of novel therapeutic strategies in CRS.

The impact of type 2 immunity and allergic diseases in atherosclerosis.

Allergic diseases are allergen-induced immunological disorders characterized by the development of type 2 immunity and IgE responses. The prevalence of allergic diseases has been on the rise alike cardiovascular disease (CVD), which affects arteries of different organs such as the heart, the kidney and the brain. The underlying cause of CVD is often atherosclerosis, a disease distinguished by endothelial dysfunction, fibrofatty material accumulation in the intima of the artery wall, smooth muscle cell proliferation, and Th1 inflammation. The opposed T-cell identity of allergy and atherosclerosis implies an atheroprotective role for Th2 cells by counteracting Th1 responses. Yet, the clinical association between allergic disease and CVD argues against it. Within, we review different phases of allergic pathology, basic immunological mechanisms of atherosclerosis and the clinical association between allergic diseases (particularly asthma, atopic dermatitis, allergic rhinitis and food allergy) and CVD. Then, we discuss putative atherogenic mechanisms of type 2 immunity and allergic inflammation including acute allergic reactions (IgE, IgG1, mast cells, macrophages and allergic mediators such as vasoactive components, growth factors and those derived from the complement, contact and coagulation systems) and late phase inflammation (Th2 cells, eosinophils, type 2 innate-like lymphoid cells, alarmins, IL-4, IL-5, IL-9, IL-13 and IL-17).

CD163 deficiency increases foam cell formation and plaque progression in atherosclerotic mice.

Atherosclerosis is an inflammatory disease characterized by the accumulation of macrophages in the vessel wall. Macrophages depend on their polarization to exert either pro-inflammatory or anti-inflammatory effects. Macrophages of the anti-inflammatory phenotype express high levels of CD163, a scavenger receptor for the hemoglobin-haptoglobin complex. CD163 can also bind to the pro-inflammatory cytokine TWEAK. Using ApoE-deficient or ApoE/CD163 double-deficient mice we aim to investigate the involvement of CD163 in atherosclerosis development and its capacity to neutralize the TWEAK actions. ApoE/CD163 double-deficient mice displayed a more unstable plaque phenotype characterized by an increased lipid and macrophage content, plaque size, and pro-inflammatory cytokine expression. In vitro experiments demonstrated that the absence of CD163 in M2-type macrophages-induced foam cell formation through upregulation of CD36 expression. Moreover, exogenous TWEAK administration increased atherosclerotic lesion size, lipids, and macrophages content in ApoE-/- /CD163-/- compared with ApoE-/- /CD163+/+ mice. Treatment with recombinant CD163 was able to neutralize the proatherogenic effects of TWEAK in ApoE/CD163 double-deficient mice. Recombinant CD163 abolished the pro-inflammatory actions of TWEAK on vascular smooth muscle cells, decreasing NF-kB activation, cytokines and metalloproteinases expression, and macrophages migration. In conclusion, CD163-expressing macrophages serve as a protective mechanism to prevent the deleterious effects of TWEAK on atherosclerotic plaque development and progression.

Aging-Associated miR-217 Aggravates Atherosclerosis and Promotes Cardiovascular Dysfunction.

OBJECTIVE: microRNAs are master regulators of gene expression with essential roles in virtually all biological processes. miR-217 has been associated with aging and cellular senescence, but its role in vascular disease is not understood. Approach and Results: We have used an inducible endothelium-specific knock-in mouse model to address the role of miR-217 in vascular function and atherosclerosis. miR-217 reduced NO production and promoted endothelial dysfunction, increased blood pressure, and exacerbated atherosclerosis in proatherogenic apoE-/- mice. Moreover, increased endothelial miR-217 expression led to the development of coronary artery disease and altered left ventricular heart function, inducing diastolic and systolic dysfunction. Conversely, inhibition of endogenous vascular miR-217 in apoE-/- mice improved vascular contractility and diminished atherosclerosis. Transcriptome analysis revealed that miR-217 regulates an endothelial signaling hub and downregulates a network of eNOS (endothelial NO synthase) activators, including VEGF (vascular endothelial growth factor) and apelin receptor pathways, resulting in diminished eNOS expression. Further analysis revealed that human plasma miR-217 is a biomarker of vascular aging and cardiovascular risk. CONCLUSIONS: Our results highlight the therapeutic potential of miR-217 inhibitors in aging-related cardiovascular disease.

The TNF-like weak inducer of the apoptosis/fibroblast growth factor-inducible molecule 14 axis mediates histamine and platelet-activating factor-induced subcutaneous vascular leakage and anaphylactic shock.

BACKGROUND: Anaphylaxis includes mast cell (MC) activation, but less is known about downstream mechanisms (ie, vascular permeability controlled by endothelial cells [ECs]). The TNF-like weak inducer of apoptosis (TWEAK) and its sole receptor, fibroblast growth factor-inducible molecule 14 (Fn14), belong to the TNF superfamily and are involved in proinflammatory responses. OBJECTIVE: We sought to investigate the role of TWEAK/Fn14 axis in anaphylaxis. METHODS: In vivo vascular permeability and mouse models of passive systemic anaphylaxis (PSA) and active systemic anaphylaxis were applied to wild-type (WT), TWEAK- and Fn14-deficient mice (TWEAK-/- and Fn14-/-, respectively). Primary bone marrow-derived mast cells (BMMCs) and ECs from WT and Fn14-/- or TWEAK-/- mice were studied. The TWEAK/Fn14 axis was also investigated in human samples. RESULTS: Mice with PSA and active systemic anaphylaxis had increased Fn14 and TWEAK expression in lung tissues and increased serum soluble TWEAK concentrations. TWEAK and Fn14 deficiencies prevent PSA-related symptoms, resulting in resistance to decreased body temperature, less severe reactions, and maintained physical activity. Numbers of MCs after PSA are similar between genotypes in different tissue regions, such as ear skin and the trachea, tongue, peritoneum, lungs, and bone marrow. Moreover, in vitro studies revealed no differences in degranulation or mediator release between WT and Fn14-/- BMMCs after IgE-FcεRI stimulation. In vivo and in vitro histamine and platelet-activating factor administration increases Fn14 receptor expression in lungs and ECs. Moreover, Fn14 deficiency in ECs maintained in vitro impermeability when stimulated by mediators or activated BMMCs but not by TWEAK-/- BMMCs, indicating that Fn14 is crucial for endothelial barrier function. TWEAK/Fn14 deletion or TWEAK-blocking antibody prevented histamine/platelet-activating factor-induced vascular subcutaneous permeability. Circulating soluble TWEAK levels were increased in patients with anaphylaxis, and plasma from those patients increased Fn14 expression in ECs. CONCLUSION: The TWEAK/Fn14 axis participates in anaphylactic reactions. Inhibition of TWEAK/Fn14 interaction could be efficacious in anaphylaxis therapy.

Cellular Crosstalk between Endothelial and Smooth Muscle Cells in Vascular Wall Remodeling.

Pathological vascular wall remodeling refers to the structural and functional changes of the vessel wall that occur in response to injury that eventually leads to cardiovascular disease (CVD). Vessel wall are composed of two major primary cells types, endothelial cells (EC) and vascular smooth muscle cells (VSMCs). The physiological communications between these two cell types (EC-VSMCs) are crucial in the development of the vasculature and in the homeostasis of mature vessels. Moreover, aberrant EC-VSMCs communication has been associated to the promotor of various disease states including vascular wall remodeling. Paracrine regulations by bioactive molecules, communication via direct contact (junctions) or information transfer via extracellular vesicles or extracellular matrix are main crosstalk mechanisms. Identification of the nature of this EC-VSMCs crosstalk may offer strategies to develop new insights for prevention and treatment of disease that curse with vascular remodeling. Here, we will review the molecular mechanisms underlying the interplay between EC and VSMCs. Additionally, we highlight the potential applicable methodologies of the co-culture systems to identify cellular and molecular mechanisms involved in pathological vascular wall remodeling, opening questions about the future research directions.

Regulator of calcineurin 1 modulates vascular contractility and stiffness through the upregulation of COX-2-derived prostanoids.

Cyclooxygenase-2 (COX-2) derived-prostanoids participate in the altered vascular function and mechanical properties in cardiovascular diseases. We investigated whether regulator of calcineurin 1 (Rcan1) participates in vascular contractility and stiffness through the regulation of COX-2. For this, wild type (Rcan1+/+) and Rcan1-deficient (Rcan1-/-) mice untreated or treated with the COX-2 inhibitor rofecoxib were used. Vascular function and structure were analysed by myography. COX-2 and phospo-p65 expression were studied by western blotting and immunohistochemistry and TXA2 production by ELISA. We found that Rcan1 deficiency increases COX-2 and IL-6 expression and NF-κB activation in arteries and vascular smooth muscle cells (VSMC). Adenoviral-mediated re-expression of Rcan1.4 in Rcan1-/- VSMC normalized COX-2 expression. Phenylephrine-induced vasoconstrictor responses were greater in aorta from Rcan1-/- compared to Rcan1+/+ mice. This increased response were diminished by etoricoxib, furegrelate, SQ 29548, cyclosporine A and parthenolide, inhibitors of COX-2, TXA2 synthase, TP receptors, calcineurin and NF-κB, respectively. Endothelial removal and NOS inhibition increased phenylephrine responses only in Rcan1+/+ mice. TXA2 levels were greater in Rcan1-/- mice. In small mesenteric arteries, vascular function and structure were similar in both groups of mice; however, vessels from Rcan1-/- mice displayed an increase in vascular stiffness that was diminished by rofecoxib. In conclusion, our results suggest that Rcan1 might act as endogenous negative modulator of COX-2 expression and activity by inhibiting calcineurin and NF-kB pathways to maintain normal contractility and vascular stiffness in aorta and small mesenteric arteries, respectively. Our results uncover a new role for Rcan1 in vascular contractility and mechanical properties.

A Novel Systems-Biology Algorithm for the Analysis of Coordinated Protein Responses Using Quantitative Proteomics.

The coordinated behavior of proteins is central to systems biology. However, the underlying mechanisms are poorly known and methods to analyze coordination by conventional quantitative proteomics are still lacking. We present the Systems Biology Triangle (SBT), a new algorithm that allows the study of protein coordination by pairwise quantitative proteomics. The Systems Biology Triangle detected statistically significant coordination in diverse biological models of very different nature and subjected to different kinds of perturbations. The Systems Biology Triangle also revealed with unprecedented molecular detail an array of coordinated, early protein responses in vascular smooth muscle cells treated at different times with angiotensin-II. These responses included activation of protein synthesis, folding, turnover, and muscle contraction - consistent with a differentiated phenotype-as well as the induction of migration and the repression of cell proliferation and secretion. Remarkably, the majority of the altered functional categories were protein complexes, interaction networks, or metabolic pathways. These changes could not be detected by other algorithms widely used by the proteomics community, and the vast majority of proteins involved have not been described before to be regulated by AngII. The unique capabilities of The Systems Biology Triangle to detect functional protein alterations produced by the coordinated action of proteins in pairwise quantitative proteomics experiments make this algorithm an attractive choice for the biological interpretation of results on a routine basis.

Deficiency of MMP17/MT4-MMP proteolytic activity predisposes to aortic aneurysm in mice.

RATIONALE: Aortic dissection or rupture resulting from aneurysm causes 1% to 2% of deaths in developed countries. These disorders are associated with mutations in genes that affect vascular smooth muscle cell differentiation and contractility or extracellular matrix composition and assembly. However, as many as 75% of patients with a family history of aortic aneurysms do not have an identified genetic syndrome. OBJECTIVE: To determine the role of the protease MMP17/MT4-MMP in the arterial wall and its possible relevance in human aortic pathology. METHODS AND RESULTS: Screening of patients with inherited thoracic aortic aneurysms and dissections identified a missense mutation (R373H) in the MMP17 gene that prevented the expression of the protease in human transfected cells. Using a loss-of-function genetic mouse model, we demonstrated that the lack of Mmp17 resulted in the presence of dysfunctional vascular smooth muscle cells and altered extracellular matrix in the vessel wall; and it led to increased susceptibility to angiotensin-II-induced thoracic aortic aneurysm. We also showed that Mmp17-mediated osteopontin cleavage regulated vascular smooth muscle cell maturation via c-Jun N-terminal kinase signaling during aorta wall development. Some features of the arterial phenotype were prevented by re-expression of catalytically active Mmp17 or the N-terminal osteopontin fragment in Mmp17-null neonates. CONCLUSIONS: Mmp17 proteolytic activity regulates vascular smooth muscle cell phenotype in the arterial vessel wall, and its absence predisposes to thoracic aortic aneurysm in mice. The rescue of part of the vessel-wall phenotype by a lentiviral strategy opens avenues for therapeutic intervention in these life-threatening disorders.

In vitro 3D co-culture model of human endothelial and smooth muscle cells to study pathological vascular remodeling. / Modelo de cocultivo 3D in vitro de células endoteliales y vasculares de músculo liso humanas para el estudio del remodelado vascular patológico.

Pathological vascular remodeling of the vessel wall refers to the structural and functional changes of the vessel wall that occur in response to injury that eventually leads to cardiovascular disease. The vessel wall is composed of two main types of cells, endothelial cells and vascular smooth muscle cells, whose communication is crucial in both the development of the vasculature and the homeostasis of mature vessels. Changes in the dialogue between endothelial cells and vascular smooth muscle cells are associated with various pathological states that triggers remodeling of the vascular wall. For many years, considerable efforts have been made to develop effective diagnoses and treatments for these pathologies by studying their mechanisms in both in vitro and in vivo models. Compared to animal models, in vitro models can provide great opportunities to obtain data in a more homogeneous, economical and massive way, providing an overview of the signaling pathways responsible for these pathologies. The implementation of three-dimensional in vitro co-culture models for the study of other pathologies has been postulated as a potentially applicable methodology, which determines the importance of its application in studies of cardiovascular diseases. In this article we present a method for culturing human endothelial cells and vascular smooth muscle cells, grown under non-adherent conditions, that generate three-dimensional spheroidal structures with greater physiological equivalence to in vivo conditions. This in vitro modeling could be used as a study tool to identify cellular and molecular mechanisms involved in the pathological processes underlying vascular remodeling.

Annexins and cardiovascular diseases: Beyond membrane trafficking and repair.

Cardiovascular diseases (CVD) remain the leading cause of mortality worldwide. The main cause underlying CVD is associated with the pathological remodeling of the vascular wall, involving several cell types, including endothelial cells, vascular smooth muscle cells, and leukocytes. Vascular remodeling is often related with the development of atherosclerotic plaques leading to narrowing of the arteries and reduced blood flow. Atherosclerosis is known to be triggered by high blood cholesterol levels, which in the presence of a dysfunctional endothelium, results in the retention of lipoproteins in the artery wall, leading to an immune-inflammatory response. Continued hypercholesterolemia and inflammation aggravate the progression of atherosclerotic plaque over time, which is often complicated by thrombus development, leading to the possibility of CV events such as myocardial infarction or stroke. Annexins are a family of proteins with high structural homology that bind phospholipids in a calcium-dependent manner. These proteins are involved in several biological functions, from cell structural organization to growth regulation and vesicle trafficking. In vitro gain- or loss-of-function experiments have demonstrated the implication of annexins with a wide variety of cellular processes independent of calcium signaling such as immune-inflammatory response, cell proliferation, migration, differentiation, apoptosis, and membrane repair. In the last years, the use of mice deficient for different annexins has provided insight into additional functions of these proteins in vivo, and their involvement in different pathologies. This review will focus in the role of annexins in CVD, highlighting the mechanisms involved and the potential therapeutic effects of these proteins.

Galectin-1 prevents pathological vascular remodeling in atherosclerosis and abdominal aortic aneurysm.

Pathological vascular remodeling is the underlying cause of atherosclerosis and abdominal aortic aneurysm (AAA). Here, we analyzed the role of galectin-1 (Gal-1), a ß-galactoside-binding protein, as a therapeutic target for atherosclerosis and AAA. Mice lacking Gal-1 (Lgals1-/-) developed severe atherosclerosis induced by pAAV/D377Y-mPCSK9 adenovirus and displayed higher lipid levels and lower expression of contractile markers of vascular smooth muscle cells (VSMCs) in plaques than wild-type mice. Proteomic analysis of Lgals1-/- aortas showed changes in markers of VSMC phenotypic switch and altered composition of mitochondrial proteins. Mechanistically, Gal-1 silencing resulted in increased foam cell formation and mitochondrial dysfunction in VSMCs, while treatment with recombinant Gal-1 (rGal-1) prevented these effects. Furthermore, rGal-1 treatment attenuated atherosclerosis and elastase-induced AAA, leading to higher contractile VSMCs in aortic tissues. Gal-1 expression decreased in human atheroma and AAA compared to control tissue. Thus, Gal-1-driven circuits emerge as potential therapeutic strategies in atherosclerosis and AAA.

Annexins: Involvement in cholesterol homeostasis, inflammatory response and atherosclerosis. / Anexinas: implicación en la homeostasis del colesterol, la respuesta inflamatoria y la aterosclerosis.

The annexin superfamily consists of 12 proteins with a highly structural homology that binds to phospholipids depending on the availability of Ca2+-dependent. Different studies of overexpression, inhibition, or using recombinant proteins have linked the main function of these proteins to their dynamic and reversible binding to membranes. Annexins are found in multiple cellular compartments, regulating different functions, such as membrane trafficking, anchoring to the cell cytoskeleton, ion channel regulation, as well as pro- or anti-inflammatory and anticoagulant activities. The use of animals deficient in any of these annexins has established their possible functions in vivo, demonstrating that annexins can participate in relevant functions independent of Ca2+ signalling. This review will focus mainly on the role of different annexins in the pathological vascular remodelling that underlies the formation of the atherosclerotic lesion, as well as in the control of cholesterol homeostasis.

MCP-1 Predicts Recurrent Cardiovascular Events in Patients with Persistent Inflammation.

Clinical data indicate that patients with C-reactive protein (CRP) levels higher than 2 mg per liter suffer from persistent inflammation, which is associated with high risk of cardiovascular disease (CVD). We determined whether a panel of biomarkers associated with CVD could predict recurrent events in patients with low or persistent inflammation and coronary artery disease (CAD). We followed 917 patients with CAD (median 4.59 ± 2.39 years), assessing CRP, galectin-3, monocyte chemoattractant protein-1 (MCP-1), N-terminal fragment of brain natriuretic peptide (NT-proBNP) and troponin-I plasma levels. The primary outcome was the combination of cardiovascular events (acute coronary syndrome, stroke or transient ischemic event, heart failure or death). Patients with persistent inflammation (n = 343) showed higher NT-proBNP and MCP-1 plasma levels compared to patients with CRP < 2 mg/L. Neither MCP-1 nor NT-proBNP was associated with primary outcome in patients with CRP < 2 mg/L. However, NT-proBNP and MCP-1 plasma levels were associated with increased risk of the primary outcome in patients with persistent inflammation. When patients were divided by type of event, MCP-1 was associated with an increased risk of acute ischemic events. A significant interaction between MCP-1 and persistent inflammation was found (synergy index: 6.17 (4.39-7.95)). In conclusion, MCP-1 plasma concentration is associated with recurrent cardiovascular events in patients with persistent inflammation.

Tumor Necrosis Factor-Like Weak Inducer of Apoptosis (TWEAK)/Fibroblast Growth Factor-Inducible 14 (Fn14) Axis in Cardiovascular Diseases: Progress and Challenges.

Cardiovascular diseases (CVD) are the leading cause of mortality in Western countries. CVD include several pathologies, such as coronary artery disease, stroke, peripheral artery disease, and aortic aneurysm, among others. All of them are characterized by a pathological vascular remodeling in which inflammation plays a key role. Interaction between different members of the tumor necrosis factor superfamily and their cognate receptors induce several biological actions that may participate in CVD. The cytokine tumor necrosis factor-like weak inducer of apoptosis (TWEAK) and its functional receptor, fibroblast growth factor-inducible 14 (Fn14), are abundantly expressed during pathological cardiovascular remodeling. The TWEAK/Fn14 axis controls a variety of cellular functions, such as proliferation, differentiation, and apoptosis, and has several biological functions, such as inflammation and fibrosis that are linked to CVD. It has been demonstrated that persistent TWEAK/Fn14 activation is involved in both vessel and heart remodeling associated with acute and chronic CVD. In this review, we summarized the role of the TWEAK/Fn14 axis during pathological cardiovascular remodeling, highlighting the cellular components and the signaling pathways that are involved in these processes.

A major role of TWEAK/Fn14 axis as a therapeutic target for post-angioplasty restenosis.

BACKGROUND: Tumor necrosis factor-like weak inducer of apoptosis (Tnfsf12; TWEAK) and its receptor Fibroblast growth factor-inducible 14 (Tnfrsf12a; Fn14) participate in the inflammatory response associated with vascular remodeling. However, the functional effect of TWEAK on vascular smooth muscle cells (VSMCs) is not completely elucidated. METHODS: Next generation sequencing-based methods were performed to identify genes and pathways regulated by TWEAK in VSMCs. Flow-citometry, wound-healing scratch experiments and transwell migration assays were used to analyze VSMCs proliferation and migration. Mouse wire injury model was done to evaluate the role of TWEAK/Fn14 during neointimal hyperplasia. FINDINGS: TWEAK up-regulated 1611 and down-regulated 1091 genes in VSMCs. Using a gene-set enrichment method, we found a functional module involved in cell proliferation defined as the minimal network connecting top TWEAK up-regulated genes. In vitro experiments in wild-type or Tnfrsf12a deficient VSMCs demonstrated that TWEAK increased cell proliferation, VSMCs motility and migration. Mechanistically, TWEAK increased cyclins (cyclinD1), cyclin-dependent kinases (CDK4, CDK6) and decreased cyclin-dependent kinase inhibitors (p15lNK4B) mRNA and protein expression. Downregulation of p15INK4B induced by TWEAK was mediated by mitogen-activated protein kinase ERK and Akt activation. Tnfrsf12a or Tnfsf12 genetic depletion and pharmacological intervention with TWEAK blocking antibody reduced neointimal formation, decreasing cell proliferation, cyclin D1 and CDK4/6 expression, and increasing p15INK4B expression compared with wild type or IgG-treated mice in wire-injured femoral arteries. Finally, immunohistochemistry in human coronary arteries with stenosis or in-stent restenosis revealed high levels of Fn14, TWEAK and PCNA in VSMCs enriched areas of the neointima as compared with healthy coronary arteries. INTERPRETATION: Our data define a major role of TWEAK/Fn14 in the control of VSMCs proliferation and migration during neointimal hyperplasia after wire injury in mice, and identify TWEAK/Fn14 as a potential target for treating in-stent restenosis. FUND: ISCiii-FEDER, CIBERCV and CIBERDEM.

Pathophisiology of abdominal aortic aneurysm: biomarkers and novel therapeutic targets. / Fisiopatología del aneurisma de aorta abdominal: biomarcadores y nuevas dianas terapéuticas.

Abdominal aortic aneurysm (AAA) is a vascular pathology with a high rate of morbidity and mortality and a prevalence that, in men over 65 years, can reach around 8%. In this disease, usually asymptomatic, there is a progressive dilatation of the vascular wall that can lead to its rupture, a fatal phenomenon in more than 80% of cases. The treatment of patients with asymptomatic aneurysms is limited to periodic monitoring with imaging tests, control of cardiovascular risk factors and treatment with statins and antiplatelet therapy. There is no effective pharmacological treatment capable of limiting AAA progression or avoiding their rupture. At present, the aortic diameter is the only marker of risk of rupture and determines the need for surgical repair when it reaches values greater than 5.5cm. This review addresses the main aspects related to epidemiology, risk factors, diagnosis and clinical management of AAA, exposes the difficulties to have good biomarkers of this pathology and describes the strategies for the identification of new therapeutic targets and biomarkers in AAA.

Conditional deletion of Rcan1 predisposes to hypertension-mediated intramural hematoma and subsequent aneurysm and aortic rupture.

Aortic intramural hematoma (IMH) can evolve toward reabsorption, dissection or aneurysm. Hypertension is the most common predisposing factor in IMH and aneurysm patients, and the hypertensive mediator angiotensin-II induces both in mice. We have previously shown that constitutive deletion of Rcan1 isoforms prevents Angiotensin II-induced aneurysm in mice. Here we generate mice conditionally lacking each isoform or all isoforms in vascular smooth muscle cells, endothelial cells, or ubiquitously, to determine the contribution to aneurysm development of Rcan1 isoforms in vascular cells. Surprisingly, conditional Rcan1 deletion in either vascular cell-type induces a hypercontractile phenotype and aortic medial layer disorganization, predisposing to hypertension-mediated aortic rupture, IMH, and aneurysm. These processes are blocked by ROCK inhibition. We find that Rcan1 associates with GSK-3ß, whose inhibition decreases myosin activation. Our results identify potential therapeutic targets for intervention in IMH and aneurysm and call for caution when interpreting phenotypes of constitutively and inducibly deficient mice.