Animal Models of Cardiovascular Diseases

Cardiovascular diseases are the first leading cause of death and morbidity in developed countries. The use of animal models have contributed to increase our knowledge, providing new approaches focused to improve the diagnostic and the treatment of these pathologies. Several models have been developed to address cardiovascular complications, including atherothrombotic and cardiac diseases, and the same pathology have been successfully recreated in different species, including small and big animal models of disease. However, genetic and environmental factors play a significant role in cardiovascular pathophysiology, making difficult to match a particular disease, with a single experimental model. Therefore, no exclusive method perfectly recreates the human complication, and depending on the model, additional considerations of cost, infrastructure, and the requirement for specialized personnel, should also have in mind. Considering all these facts, and depending on the budgets available, models should be selected that best reproduce the disease being investigated. Here we will describe models of atherothrombotic diseases, including expanding and occlusive animal models, as well as models of heart failure. Given the wide range of models available, today it is possible to devise the best strategy, which may help us to find more efficient and reliable solutions against human cardiovascular diseases

Retention and Activation of Blood-Borne Proteases in the Arterial Wall Implications for Atherothrombosis

All forms of atheroma are characterized by a risk of arterial wall rupture leading to clinical complications. This involves medial and adventitial ruptures in abdominal aortic aneurysm (AAA) and intimal cap rupture in vulnerable atherothrombotic plaques. Extracellular proteases, including metalloproteinases, locally generated plasmin, and leukocyte elastase, are important molecular mediators of atheroma progression via their matrix degradation properties. The pathological evolution of AAA is linked to the biology of its associated mural thrombus. Indeed, in aneurysmal segments lined by a thrombus, the wall is thinner, the extracellular matrix more degraded, and the adventitial inflammatory response greater than in segments that are not. Several lines of evidence highlight the role of the thrombus, in AAA, as a reservoir of blood-borne proteases that conveys them from the lumen to the diseased wall. In stenosing atheroma, both previous and recent studies provide evidence that recurrent intraplaque hemorrhages play a dominant role in the evolution of the lesion toward vulnerability. In this review, we draw a parallel between the role of protease conveyance and activation of the mural thrombus in AAA and of intraplaque hemorrhages in stenosing atheroma. We hypothesize that intraplaque hemorrhages convey blood-borne proteases into lesions, where they are retained and activated upon thrombus/hematoma formation, thus contributing significantly to their deleterious action

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

Quantitative HDL Proteomics Identifies Peroxiredoxin-6 as a Biomarker of Human Abdominal Aortic Aneurysm

High-density lipoproteins (HDLs) are complex protein and lipid assemblies whose composition is known to change in diverse pathological situations. Analysis of the HDL proteome can thus provide insight into the main mechanisms underlying abdominal aortic aneurysm (AAA) and potentially detect novel systemic biomarkers. We performed a multiplexed quantitative proteomics analysis of HDLs isolated from plasma of AAA patients (N = 14) and control study participants (N = 7). Validation was performed by western-blot (HDL), immunohistochemistry (tissue), and ELISA (plasma). HDL from AAA patients showed elevated expression of peroxiredoxin-6 (PRDX6), HLA class I histocompatibility antigen (HLA-I), retinol-binding protein 4, and paraoxonase/arylesterase 1 (PON1), whereas alpha-2 macroglobulin and C4b-binding protein were decreased. The main pathways associated with HDL alterations in AAA were oxidative stress and immune-inflammatory responses. In AAA tissue, PRDX6 colocalized with neutrophils, vascular smooth muscle cells, and lipid oxidation. Moreover, plasma PRDX6 was higher in AAA (N = 47) than in controls (N = 27), reflecting increased systemic oxidative stress. Finally, a positive correlation was recorded between PRDX6 and AAA diameter. The analysis of the HDL proteome demonstrates that redox imbalance is a major mechanism in AAA, identifying the antioxidant PRDX6 as a novel systemic biomarker of AAA.We thank Simon Bartlett for language and scientific editing. This study was supported by the Spanish Ministry of Economy and Competitiveness (MINECO) (SAF2016-80843-R, BIO2012-37926 and BIO2015-67580-P), Fondo de Investigaciones Sanitarias ISCiii-FEDER (PRB2) (IPT13/0001, ProteoRed, Redes RIC RD12/0042/00038 and RD12/0042/0056, Biobancos RD09/0076/00101 and CA12/00371), Centro de Investigacion Biomedica en Red de Diabetes y Enfermedades Metabolicas Asociadas (CIBERDEM), and FRIAT. The CNIC is supported by the Spanish Ministry of Economy and Competitiveness (MINECO) and the Pro-CNIC Foundation, and is a Severo Ochoa Center of Excellence (MINECO award SEV-2015-0505).S

Targeted gold-coated iron oxide nanoparticles for CD163 detection in atherosclerosis by MRI

CD163 is a membrane receptor expressed by macrophage lineage. Studies performed in atherosclerosis have shown that CD163 expression is increased at inflammatory sites, pointing at the presence of intraplaque hemorrhagic sites or asymptomatic plaques. Hence, imaging of CD163 expressing macrophages is an interesting strategy in order to detect atherosclerotic plaques. We have prepared a targeted probe based on gold-coated iron oxide nanoparticles vectorized with an anti-CD163 antibody for the specific detection of CD163 by MRI. Firstly, the specificity of the targeted probe was validated in vitro by incubation of the probe with CD163(+) or (−) macrophages. The probe was able to selectively detect CD163(+) macrophages both in human and murine cells. Subsequently, the targeted probe was injected in 16 weeks old apoE deficient mice developing atherosclerotic lesions and the pararenal abdominal aorta was imaged by MRI. The accumulation of probe in the site of interest increased over time and the signal intensity decreased significantly 48 hours after the injection. Hence, we have developed a highly sensitive targeted probe capable of detecting CD163-expressing macrophages that could provide useful information about the state of the atheromatous lesionsThis work was funded by Spanish Government through a Plan Nacional (CTQ2011–27268), FEDER funds through the Fondo de Investigación Sanitaria (PI10/00072, PI13/00051, PI13/00395, PI13/00802, PI14/00883 and PI14/00386), CIBERDEM group, RETICS RD12/0042/0038, Programa Miguel Servet (CP10/00479) and cvREMOD CENIT project (CEN-20091044), the Basque Government through Etortek 2011 (IE11–301), and Fundacion Lilly, Spanish Society of Atherosclerosis, Spanish Society of Nephrology and Fundacion Renal Iñigo Alvarez de Toled

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

Metabolomics with LC-QTOF-MS Permits the Prediction of Disease Stage in Aortic Abdominal Aneurysm Based on Plasma Metabolic Fingerprint

Abdominal aortic aneurysm (AAA) is a permanent and localized aortic dilation, defined as aortic diameter ≥3 cm. It is an asymptomatic but potentially fatal condition because progressive enlargement of the abdominal aorta is spontaneously evolving towards rupture

Electrostatic Effects in the Folding of the SH3 Domain of the c-Src Tyrosine Kinase: pH-Dependence in 3D-Domain Swapping and Amyloid Formation

The SH3 domain of the c-Src tyrosine kinase (c-Src-SH3) aggregates to form intertwined dimers and amyloid fibrils at mild acid pHs. In this work, we show that a single mutation of residue Gln128 of this SH3 domain has a significant effect on: (i) its thermal stability; and (ii) its propensity to form amyloid fibrils. The Gln128Glu mutant forms amyloid fibrils at neutral pH but not at mild acid pH, while Gln128Lys and Gln128Arg mutants do not form these aggregates under any of the conditions assayed. We have also solved the crystallographic structures of the wild-type (WT) and Gln128Glu, Gln128Lys and Gln128Arg mutants from crystals obtained at different pHs. At pH 5.0, crystals belong to the hexagonal space group P6522 and the asymmetric unit is formed by one chain of the protomer of the c-Src-SH3 domain in an open conformation. At pH 7.0, crystals belong to the orthorhombic space group P212121, with two molecules at the asymmetric unit showing the characteristic fold of the SH3 domain. Analysis of these crystallographic structures shows that the residue at position 128 is connected to Glu106 at the diverging β-turn through a cluster of water molecules. Changes in this hydrogen-bond network lead to the displacement of the c-Src-SH3 distal loop, resulting also in conformational changes of Leu100 that might be related to the binding of proline rich motifs. Our findings show that electrostatic interactions and solvation of residues close to the folding nucleation site of the c-Src-SH3 domain might play an important role during the folding reaction and the amyloid fibril formation.This research was funded by the Spanish Ministry of Science and Innovation and Ministry of Economy and Competitiveness and FEDER (EU): BIO2009-13261-C02-01/02 (ACA); BIO2012-39922-C02-01/02 (ACA); CTQ2013-4493 (JLN) and CSD2008-00005 (JLN); Andalusian Regional Government (Spain) and FEDER (EU): P09-CVI-5063 (ACA); and Valentian Regional Government (Spain) and FEDER (EU): Prometeo 2013/018 (JLN). Data collection was supported by European Synchrotron Radiation Facility (ESRF), Grenoble, France: BAG proposals MX-1406 (ACA) and MX-1541 (ACA); and ALBA (Barcelona, Spain) proposals 2012010072 (ACA) and 2012100378 (ACA)

A search for ultra-high-energy photons at the Pierre Auger Observatory exploiting air-shower universality

The Pierre Auger Observatory is the most sensitive detector to primary photons with energies above ∼0.2 EeV. It measures extensive air showers using a hybrid technique that combines a fluorescence detector (FD) with a ground array of particle detectors (SD). The signatures of a photon-induced air shower are a larger atmospheric depth at the shower maximum (X$_{max}$) and a steeper lateral distribution function, along with a lower number of muons with respect to the bulk of hadron-induced background. Using observables measured by the FD and SD, three photon searches in different energy bands are performed. In particular, between threshold energies of 1-10 EeV, a new analysis technique has been developed by combining the FD-based measurement of X$_{max}$ with the SD signal through a parameter related to its muon content, derived from the universality of the air showers. This technique has led to a better photon/hadron separation and, consequently, to a higher search sensitivity, resulting in a tighter upper limit than before. The outcome of this new analysis is presented here, along with previous results in the energy ranges below 1 EeV and above 10 EeV. From the data collected by the Pierre Auger Observatory in about 15 years of operation, the most stringent constraints on the fraction of photons in the cosmic flux are set over almost three decades in energy

Study on multi-ELVES in the Pierre Auger Observatory

Since 2013, the four sites of the Fluorescence Detector (FD) of the Pierre Auger Observatory record ELVES with a dedicated trigger. These UV light emissions are correlated to distant lightning strikes. The length of recorded traces has been increased from 100 μs (2013), to 300 μs (2014-16), to 900 μs (2017-present), to progressively extend the observation of the light emission towards the vertical of the causative lightning and beyond. A large fraction of the observed events shows double ELVES within the time window, and, in some cases, even more complex structures are observed. The nature of the multi-ELVES is not completely understood but may be related to the different types of lightning in which they are originated. For example, it is known that Narrow Bipolar Events can produce double ELVES, and Energetic In-cloud Pulses, occurring between the main negative and upper positive charge layer of clouds, can induce double and even quadruple ELVES in the ionosphere. This report shows the seasonal and daily dependence of the time gap, amplitude ratio, and correlation between the pulse widths of the peaks in a sample of 1000+ multi-ELVES events recorded during the period 2014-20. The events have been compared with data from other satellite and ground-based sensing devices to study the correlation of their properties with lightning observables such as altitude and polarity