A009 Importance of tweak-CD163 system in peripheral artery disease

IntroductionCD163 is a macrophage receptor of haptoglogin/ haemoglobin complexes responsible for clearance of hemogloin. It has been recently suggested to be a potential scavenger receptor for TWEAK (Tumor necrosis factor-like weak inducer of apoptosis). TWEAK levels were reported to be decreased in carotid atherosclerosis. Our hypothesis is that decreased circulating TWEAK could be paralleled by an increased presence of CD163-expressing macrophage in atherosclerotic plaques. Since peripheral artery disease (PAD) is an important manifestation of systemic atherosclerosis, we have assessed the levels of circulating TWEAK-CD163 in PAD.Methods and ResultsPatients with PAD (n=184) had lower TWEAK (169.2±8.3vs 211.9±15.4pg/mL; p<0.05) and higher sCD163 (408.1±14.5vs 317.4±8.4ng/mL; p<0.05) plasma concentration than age-matched controls (n=330). After stratification according to the severity of disease, we observed that TWEAK/sCD163 ratio was significantly decreased in those patients with higher degree of disease (0.39±0.06vs 0.66±0.08, p<0.05) relative to the other groups. Analysis of conditioned medium obtained from cultured human atherosclerotic femoral plaque samples (n=38) and healthy aortas (n=14) revealed that higher amount of sCD163 was released by the atherosclerotic tissue, whereas TWEAK presented the opposite trend.ConclusionsOur results suggest that CD163/TWEAK plasma ratio could be a potential biomarker of clinical peripheral artery disease. We can hypothesized that decreased levels of circulating TWEAK observed in atherosclerosis may be the result of a trapping by plaque macrophages through their CD163

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 ofTWEAK on vascular smoothmuscle cells (VSMCs) is not completely elucidated. Methods: Next generation sequencing-based methodswere performed to identify genes and pathways regulated by TWEAK in VSMCs. Flow-citometry, wound-healing scratch experiments and transwellmigration 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 functionalmodule 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.This work was supported by Instituto de Salud Carlos III (Fondo de Investigaciones Sanitarias ISCiii/FEDER PI13/00395; PI16/01419; PI17/ 01495) and Spanish Biomedical Research Centre in Cardiovascular Disease (CIBERCV) and Metabolic Diseases and Diabetes (CIBERDEM). PM was supported by ISCIII Miguel Servet Program (CP16/00116). CGM was supported by Fundación Conchita Rábago. NMB and VE were supported by the Spanish Ministry of Economy and Competitiveness (Juan de la Cierva IJCI-2016-29630 and Ramón y Ramón Cajal Program RyC-2013-12880, respectively). JMM has been supported a postdoctoral fellowship fromthe American Diabetes Association (Grant 1-15-MI-03) and a postdoctoral fellowship fromthe American Heart Association

Proteomic approach in the search of new cardiovascular biomarkers

Proteomic approach in the search of new cardiovascular biomarkers With the increasing incidence of cardiovascular diseases worldwide, specifically atherosclerosis and heart failure, the search for novel biomarkers remains a priority. As opposed to complex diagnostic techniques that may not be suitable to be applied to the wider population, biomarkers are useful for population screening. The search for novel biomarkers is based on knowledge of the molecular and cellular processes that take place in the development of a specific disease. Atherosclerosis and heart failure are characterized by a long period of silent disease progression, allowing early diagnosis and the potential of early therapeutic intervention. The use of the so-called proteomic techniques allows not only protein identification but partial characterization, which includes expression and also post-translational modification of these proteins. This allows for the discovery of previously unknown proteins involved in cardiovascular diseases, including some that may be suitable to be used as biomarkers. However, to approach this issue, we have to overcome difficulties such as tissue heterogeneity (vessel wall or myocardium) and the lack of fresh human samples. We discuss the proteomic study of human plaques, secreted proteins by pathologic and normal vessel wall, and left ventricular hypertrophy as potential sources of new biologic markers of cardiovascular disease

Combined Immunoglobulin Free Light Chains Are Novel Predictors of Cardiovascular Events in Patients With Abdominal Aortic Aneurysm.

Abdominal aortic aneurysm (AAA) is characterised by the presence of B cells and immunoglobulins in the aortic wall, mainly in the adventitia. Kappa (κ) and lambda (λ) free light chains (FLCs) are produced from B cells during immunoglobulin synthesis. This study investigated the presence and prognostic value of combined FLCs (cFLCs or summed κ and λ) in patients with AAA. cFLCs were analysed by a turbidimetric specific assay in tissue conditioned media from AAA samples (n = 34) compared with healthy aortas (n = 34) from France and in plasma samples from patients with AAA (n = 434) and age matched controls (n = 104) selected from the Viborg Vascular (VIVA) AAA screening trial in Denmark. t test, logistic regression, and Cox regression were used to test whether plasma cFLCs serve as a marker for AAA presence and whether cFLCs were predictive of death, major adverse cardiovascular events (MACE), or major adverse lower limb events (MALE). Increased cFLC levels were detected in the AAA adventitial layer compared with the AAA medial layer and healthy media layer (13.65 ± 3.17 vs. 6.57 ± 1.01 vs. 0.49 ± 0.09 mg/L, respectively, p < .050). The upper tertile of plasma cFLCs was independently associated with AAA presence after correcting for confounders (odds ratio [OR] 7.596, 95% confidence intervals [CI] 3.117 - 18.513; p < .001). Of 434 patients with AAA, 89 (20.5%) died, 104 (24.0%) suffered MACE, and 63 (14.5%) suffered MALE, during a five year follow up. In univariable analysis, the cFLC upper tertile was associated with a higher risk of death, MACE, and MALE (p < .001 for all). After adjustment for confounders, cFLCs remained an independent predictor of all cause mortality (hazard ratio [HR] 4.310, 95% CI 2.157 - 8.609; p < .001), MACE (HR 2.153, 95% CI 1.218 - 3.804; p = .008), or MALE (HR 3.442, 95% CI 1.548 - 7.652; p = .002) for those in the upper tertile. Increased cFLCs are observed in adventitial tissue of patients with AAA, indicating local activation of B cells. Plasma cFLC levels are an independent predictor of death, MACE, and MALE in patients with AAA.This study was funded by the Spanish MINECO (PID2019- 106814RB-I00 and PGC2018-097019-B-I00), la Caixa Foundation (HR17-00247), CAM (S2017/BMD-3673), and Fondo de Investigaciones Sanitarias ISCiii-FEDER (PI19/ 00128).S

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 β-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. Galectin-1 plays an essential role in prevention of atherosclerosis and abdominal aortic aneurysm

Apoptosis, autophagy and ER stress in mevalonate cascade inhibition-induced cell death of human atrial fibroblasts

3-hydroxy-3-methyl-glutaryl-CoA reductase inhibitors (statins) are cholesterol-lowering drugs that exert other cellular effects and underlie their beneficial health effects, including those associated with myocardial remodeling. We recently demonstrated that statins induces apoptosis and autophagy in human lung mesenchymal cells. Here, we extend our knowledge showing that statins simultaneously induces activation of the apoptosis, autophagy and the unfolded protein response (UPR) in primary human atrial fibroblasts (hATF). Thus we tested the degree to which coordination exists between signaling from mitochondria, endoplasmic reticulum and lysosomes during response to simvastatin exposure. Pharmacologic blockade of the activation of ER-dependent cysteine-dependent aspartate-directed protease (caspase)-4 and lysosomal cathepsin-B and -L significantly decreased simvastatin-induced cell death. Simvastatin altered total abundance and the mitochondrial fraction of proapoptotic and antiapoptotic proteins, while c-Jun N-terminal kinase/stress-activated protein kinase mediated effects on B-cell lymphoma 2 expression. Chemical inhibition of autophagy flux with bafilomycin-A1 augmented simvastatin-induced caspase activation, UPR and cell death. In mouse embryonic fibroblasts that are deficient in autophagy protein 5 and refractory to autophagy induction, caspase-7 and UPR were hyper-induced upon treatment with simvastatin. These data demonstrate that mevalonate cascade inhibition-induced death of hATF manifests from a complex mechanism involving co-regulation of apoptosis, autophagy and UPR. Furthermore, autophagy has a crucial role in determining the extent of ER stress, UPR and permissiveness of hATF to cell death induced by statins

Antiretroviral effect of lovastatin on HIV-1-infected individuals without highly active antiretroviral therapy (The LIVE study): a phase-II randomized clinical trial

<p>Abstract</p> <p>Background</p> <p>Highly active antiretroviral therapy produces a significant decrease in HIV-1 replication and allows an increase in the CD4 T-cell count, leading to a decrease in the incidence of opportunistic infections and mortality. However, the cost, side effects and complexity of antiretroviral regimens have underscored the immediate need for additional therapeutic approaches. Statins exert pleiotropic effects through a variety of mechanisms, among which there are several immunoregulatory effects, related and unrelated to their cholesterol-lowering activity that can be useful to control HIV-1 infection.</p> <p>Methods/design</p> <p>Randomized, double-blinded, placebo controlled, single-center, phase-II clinical trial. One hundred and ten chronically HIV-1-infected patients, older than 18 years and naïve for antirretroviral therapy (i.e., without prior or current management with antiretroviral drugs) will be enrolled at the outpatient services from the most important centres for health insurance care in Medellin-Colombia. The interventions will be lovastatin (40 mg/day, orally, for 12 months; 55 patients) or placebo (55 patients). Our primary aim will be to determine the effect of lovastatin on viral replication. The secondary aim will be to determine the effect of lovastatin on CD4+ T-cell count in peripheral blood. As tertiary aims we will explore differences in CD8+ T-cell count, expression of activation markers (CD38 and HLA-DR) on CD4 and CD8 T cells, cholesterol metabolism, LFA-1/ICAM-1 function, Rho GTPases function and clinical evolution between treated and not treated HIV-1-infected individuals.</p> <p>Discussion</p> <p>Preliminary descriptive studies have suggested that statins (lovastatin) may have anti HIV-1 activity and that their administration is safe, with the potential effect of controlling HIV-1 replication in chronically infected individuals who had not received antiretroviral medications. Considering that there is limited clinical data available on this topic, all these findings warrant further evaluation to determine if long-term administration of statins may benefit the virological and immunological evolution in HIV-1-infected individuals before the use of antiretroviral therapy is required.</p> <p>Trial registration</p> <p>Registration number NCT00721305.</p