The Role of Inflammation in Cardiovascular Disease.

Atherosclerosis is a chronic inflammatory disease, in which the immune system has a prominent role in its development and progression. Inflammation-induced endothelial dysfunction results in an increased permeability to lipoproteins and their subendothelial accumulation, leukocyte recruitment, and platelets activation. Recruited monocytes differentiate into macrophages which develop pro- or anti-inflammatory properties according to their microenvironment. Atheroma progression or healing is determined by the balance between these functional phenotypes. Macrophages and smooth muscle cells secrete inflammatory cytokines including interleukins IL-1ß, IL-12, and IL-6. Within the arterial wall, low-density lipoprotein cholesterol undergoes an oxidation. Additionally, triglyceride-rich lipoproteins and remnant lipoproteins exert pro-inflammatory effects. Macrophages catabolize the oxidized lipoproteins and coalesce into a lipid-rich necrotic core, encapsulated by a collagen fibrous cap, leading to the formation of fibro-atheroma. In the conditions of chronic inflammation, macrophages exert a catabolic effect on the fibrous cap, resulting in a thin-cap fibro-atheroma which makes the plaque vulnerable. However, their morphology may change over time, shifting from high-risk lesions to more stable calcified plaques. In addition to conventional cardiovascular risk factors, an exposure to acute and chronic psychological stress may increase the risk of cardiovascular disease through inflammation mediated by an increased sympathetic output which results in the release of inflammatory cytokines. Inflammation is also the link between ageing and cardiovascular disease through increased clones of leukocytes in peripheral blood. Anti-inflammatory interventions specifically blocking the cytokine pathways reduce the risk of myocardial infarction and stroke, although they increase the risk of infections.

Mechano-Transduction Boosts the Aging Effects in Human Erythrocytes Submitted to Mechanical Stimulation.

Erythrocytes' aging and mechano-transduction are fundamental cellular pathways that determine the red blood cells' (RBCs) behavior and function. The aging pattern can be influenced, in morphological, biochemical, and metabolic terms by the environmental conditions. In this paper, we studied the effect of a moderate mechanical stimulation applied through external shaking during the RBCs aging and revealed a strong acceleration of the aging pattern induced by such stimulation. Moreover, we evaluated the behavior of the main cellular effectors and resources in the presence of drugs (diamide) or of specific inhibitors of the mechano-transduction (probenecid, carbenoxolone, and glibenclamide). This approach provided the first evidence of a direct cross-correlation between aging and mechano-transduction and permitted an evaluation of the overall metabolic regulation and of the insurgence of specific morphological features, such as micro-vesicles and roughness alterations. Overall, for the first time the present data provided a schematic to understand the integration of distinct complex patterns in a comprehensive view of the cell and of its interactions with the environment. Mechano-transduction produces structural effects that are correlated with the stimulation and the strength of the environmental stimulation is paramount to effectively activate and trigger the biological cascades initiated by the mechano-sensing.

Surprising Structural and Functional Properties of Favism Erythrocytes Are Linked to Special Metabolic Regulation: A Cell Aging Study.

Favism uniquely arises from a genetic defect of the Glucose-6 Phosphate Dehydrogenase (G6PD) enzyme and results in a severe reduction of erythrocytes' (RBCs) reducing power that impairs the cells' ability to respond to oxidative stresses. After exposure to fava beans or a few other drugs, the patients experience acute hemolytic anemia due to RBCs' lysis both intra and extra-vascularly. In the present paper, we compared selected biochemical, biophysical, and ultra-morphological properties of normal RBCs and cells from favism patients measured along cellular aging. Along the aging path, the cells' characteristics change, and their structural and functional properties degrade for both samples, but with different patterns and effectors that have been characterized in biophysical and biochemical terms. In particular, the analysis revealed distinct metabolic regulation in G6DP-deficient cells that determines important peculiarities in the cell properties during aging. Remarkably, the initial higher fragility and occurrence of structural/morphological alterations of favism cells develop, with longer aging times, into a stronger resistance to external stresses and higher general resilience. This surprisingly higher endurance against cell aging has been related to a special mechanism of metabolic regulation that permits lower energy consumption in environmental stress conditions. Our results provided a direct and coherent link between the RBCs' metabolic regulation and the cell properties that would not have been possible to establish without an investigation performed during aging. The consequences of this new knowledge, in particular, can be discussed in a more general context, such as understanding the role of the present findings in determining the characteristics of the favism pathology as a whole.

Metal-based micro and nanosized pollutant in marine organisms: What can we learn from a combined atomic force microscopy-scanning electron microscopy study.

Atomic force microscopy (AFM) is a consolidated technique for the study of biological systems, usually ex vivo or in culture, under different experimental conditions. Yet, the diffusion of the technique in the scientific context of histology is still rather slow and limited. In the present work, we demonstrate the potential of AFM, in terms of morphological and nanomechanical imaging, to study the effects of nano- and micro-sized metallic pollutants in living biological systems. As a model, we investigated marine molluscs (Mytilus galloprovincialis) grown in the Adriatic Sea. We characterized histological sections from two organs (gonads and digestive glands) of molluscs collected during several surveys at different growth time and distance from gas extraction platforms. We evaluated the effects of nano-pollutants mostly on the local tissue structure by combining AFM microscopy with scanning electron microscopy (SEM). Furthermore, the AFM images allowed evidencing the presence of nano- or micro-sized structures that exhibit different nanomechanical properties compared to the rest of the tissue. The results demonstrate how coupling AFM and SEM analysis can provide an effective procedure to evaluate the morphological alterations produced by the exposure to exogenous nano-pollutants in tissue and constitute a promising way to reveal basic mechanisms mediating the cytotoxicity of specific exogenous pollutants ingested by edible organisms.

A perspective view on the nanomotion detection of living organisms and its features.

The insurgence of newly arising, rapidly developing health threats, such as drug-resistant bacteria and cancers, is one of the most urgent public-health issues of modern times. This menace calls for the development of sensitive and reliable diagnostic tools to monitor the response of single cells to chemical or pharmaceutical stimuli. Recently, it has been demonstrated that all living organisms oscillate at a nanometric scale and that these oscillations stop as soon as the organisms die. These nanometric scale oscillations can be detected by depositing living cells onto a micro-fabricated cantilever and by monitoring its displacements with an atomic force microscope-based electronics. Such devices, named nanomotion sensors, have been employed to determine the resistance profiles of life-threatening bacteria within minutes, to evaluate, among others, the effect of chemicals on yeast, neurons, and cancer cells. The data obtained so far demonstrate the advantages of nanomotion sensing devices in rapidly characterizing microorganism susceptibility to pharmaceutical agents. Here, we review the key aspects of this technique, presenting its major applications. and detailing its working protocols.

Modelling the pathogenesis of Myotonic Dystrophy type 1 cardiac phenotype through human iPSC-derived cardiomyocytes.

Myotonic Dystrophy type 1 (DM1) is a multisystemic disease, autosomal dominant, caused by a CTG repeat expansion in DMPK gene. We assessed the appropriateness of patient-specific induced pluripotent stem cell-derived cardiomyocytes (CMs) as a model to recapitulate some aspects of the pathogenetic mechanism involving cardiac manifestations in DM1 patients. Once obtained in vitro, CMs have been characterized for their morphology and their functionality. CMs DM1 show intranuclear foci and transcript markers abnormally spliced respect to WT ones, as well as several irregularities in nuclear morphology, probably caused by an unbalanced lamin A/C ratio. Electrophysiological characterization evidences an abnormal profile only in CMs DM1 such that the administration of antiarrythmic drugs to these cells highlights even more the functional defect linked to the disease. Finally, Atomic Force Measurements reveal differences in the biomechanical behaviour of CMs DM1, in terms of frequencies and synchronicity of the beats. Altogether the complex phenotype described in this work, strongly reproduces some aspects of the human DM1 cardiac phenotype. Therefore, the present study provides an in vitro model suggesting novel insights into the mechanisms leading to the development of arrhythmogenesis and dilatative cardiomyopathy to consider when approaching to DM1 patients, especially for the risk assessment of sudden cardiac death (SCD). These data could be also useful in identifying novel biomarkers effective in clinical settings and patient-tailored therapies.

FC_analysis: a tool for investigating atomic force microscopy maps of force curves.

BACKGROUND: The collection and analysis of Atomic Force Microscopy force curves is a well-established procedure to obtain high-resolution information of non-topographic data from any kind of sample, including biological specimens. In particular, these analyses are commonly employed to study elasticity, stiffness or adhesion properties of the samples. Furthermore, the collection of several force curves over an extended area of the specimens allows reconstructing maps, called force volume maps, of the spatial distribution of the mechanical properties. Coupling these maps with the conventional high-resolution topographic reconstruction of the sample's surface, provides a deeper insight on the sample composition from the structural and nanomechanical point of view. RESULTS: In this paper we present the open source software package FC_analysis that automatically analyses single force curves or entire force volume maps to yield the corresponding elasticity and deformability images. The principal characteristic of the FC_analysis is a large adaptability to the various experimental setups and to different analysis methodologies. For instance, the user can provide custom values for the detector sensitivity, scanner-z sensitivity, cantilever's elastic constant and map's acquisition modality and can choose between different analysis methodologies. Furthermore, the software allows the optimization of the fitting parameters and gives direct control on each step of the analysis procedure. Notably, to overcome a limitation common to many other analysis programs, FC_analysis can be applied to a rectangular portion of the image, allowing the analysis of inhomogeneous samples. Finally, the software allows reconstructing a Young's modulus map at different penetration depths, enabling the use of modern investigation tools such as the force tomography. CONCLUSIONS: The FC_analysis software aims to become a useful tool for the analysis of force curves maps collected using custom or commercial Atomic Force Microscopes, and is especially useful in those cases for which the producer doesn't release a dedicated software.

Detecting nanoscale vibrations as signature of life.

The existence of life in extreme conditions, in particular in extraterrestrial environments, is certainly one of the most intriguing scientific questions of our time. In this report, we demonstrate the use of an innovative nanoscale motion sensor in life-searching experiments in Earth-bound and interplanetary missions. This technique exploits the sensitivity of nanomechanical oscillators to transduce the small fluctuations that characterize living systems. The intensity of such movements is an indication of the viability of living specimens and conveys information related to their metabolic activity. Here, we show that the nanomotion detector can assess the viability of a vast range of biological specimens and that it could be the perfect complement to conventional chemical life-detection assays. Indeed, by combining chemical and dynamical measurements, we could achieve an unprecedented depth in the characterization of life in extreme and extraterrestrial environments.

Hyperplectonemes: A Higher Order Compact and Dynamic DNA Self-Organization.

Bacterial chromosome has a compact structure that dynamically changes its shape in response to bacterial growth rate and growth phase. Determining how chromatin remains accessible to DNA binding proteins, and transcription machinery is crucial to understand the link between genetic regulation, DNA structure, and topology. Here, we study very large supercoiled dsDNA using high-resolution characterization, theoretical modeling, and molecular dynamics calculations. We unveil a new type of highly ordered DNA organization forming in the presence of attractive DNA-DNA interactions, which we call hyperplectonemes. We demonstrate that their formation depends on DNA size, supercoiling, and bacterial physiology. We compare structural, nanomechanic, and dynamic properties of hyperplectonemes bound by three highly abundant nucleoid-associated proteins (FIS, H-NS, and HU). In all these cases, the negative supercoiling of DNA determines molecular dynamics, modulating their 3D shape. Overall, our findings provide a mechanistic insight into the critical role of DNA topology in genetic regulation.

Identification of Oxidative Stress in Red Blood Cells with Nanoscale Chemical Resolution by Infrared Nanospectroscopy.

During their lifespan, Red blood cells (RBC), due to their inability to self-replicate, undergo an ageing degradation phenomenon. This pathway, both in vitro and in vivo, consists of a series of chemical and morphological modifications, which include deviation from the biconcave cellular shape, oxidative stress, membrane peroxidation, lipid content decrease and uncoupling of the membrane-skeleton from the lipid bilayer. Here, we use the capabilities of atomic force microscopy based infrared nanospectroscopy (AFM-IR) to study and correlate, with nanoscale resolution, the morphological and chemical modifications that occur during the natural degradation of RBCs at the subcellular level. By using the tip of an AFM to detect the photothermal expansion of RBCs, it is possible to obtain nearly two orders of magnitude higher spatial resolution IR spectra, and absorbance images than can be obtained on diffraction-limited commercial Fourier-transform Infrared (FT-IR) microscopes. Using this approach, we demonstrate that we can identify localized sites of oxidative stress and membrane peroxidation on individual RBC, before the occurrence of neat morphological changes in the cellular shape.

Update on clinical evidence (Part II): A summary of the main post market studies.

Bioresorbable vascular scaffolds (BVS, Absorb, Abbott Vascular, Santa Clara, CA) received the CE mark in October 2011, and were approved by the Food and Drug Administration in July 2016. After their introduction in clinical practice a broad amount of post-marketing clinical experience with BVS has been generated so far in Europe and outside the United States. The available BVS registries differ in many aspects, including their being single-center or multicenter, single-arm or controlled, sponsored or investigator-initiated, published or presented at a large-scale international meeting. This article provides an overview of clinical results of the main post-marketing studies of BVS available. © 2016 Wiley Periodicals, Inc.

Everolimus-eluting bioresorbable vascular scaffolds versus second generation drug-eluting stents for percutaneous treatment of chronic total coronary occlusions: Technical and procedural outcomes from the GHOST-CTO registry.

OBJECTIVES: We aimed at comparing the acute performance of bioresorbable scaffolds (BRS) and second-generation drug-eluting stents (DES) for the treatment of chronic total occlusions (CTO). BACKGROUND: There is a lack of knowledge regarding the use of BRS in CTO. METHODS: Key outcomes of interest were technical and procedural success. Technical success was defined as successful stent delivery and implantation, postprocedural residual diameter stenosis <30% within the treated segment, and restoration of thrombolysis in myocardial infarction (TIMI) grade 3 flow. Procedural success was defined as technical success with no in-hospital major adverse cardiac events (MACE). RESULTS: Between May 2013 and May 2014, 32 patients underwent CTO percutaneous coronary intervention (PCI) with the Absorb BRS (Abbott Vascular, Santa Clara, CA) and were compared with a historical control group of 54 patients who had undergone CTO PCI with second-generation DES. Baseline characteristics were similar between the BRS and DES groups, with the exception of a larger mean reference vessel diameter in the BRS group (2.92 ± 0.34 vs 2.50 ± 0.68; P < 0.001). Technical success was less likely to be achieved in the BRS group compared with the DES group (78.1% vs 96.3%, P = 0.012). Procedural success rates were 78.1% and 94.4% in the BRS and DES group, respectively (P = 0.035). CONCLUSIONS: Compared with second-generation DES for PCI of CTO lesions, BRS were associated with lower rates of technical and procedural success. © 2016 Wiley Periodicals, Inc.

Investigation of resins suitable for the preparation of biological sample for 3-D electron microscopy.

In the last two decades, the third-dimension has become a focus of attention in electron microscopy to better understand the interactions within subcellular compartments. Initially, transmission electron tomography (TEM tomography) was introduced to image the cell volume in semi-thin sections (∼ 500 nm). With the introduction of the focused ion beam scanning electron microscope, a new tool, FIB-SEM tomography, became available to image much larger volumes. During TEM tomography and FIB-SEM tomography, the resin section is exposed to a high electron/ion dose such that the stability of the resin embedded biological sample becomes an important issue. The shrinkage of a resin section in each dimension, especially in depth, is a well-known phenomenon. To ensure the dimensional integrity of the final volume of the cell, it is important to assess the properties of the different resins and determine the formulation which has the best stability in the electron/ion beam. Here, eight different resin formulations were examined. The effects of radiation damage were evaluated after different times of TEM irradiation. To get additional information on mass-loss and the physical properties of the resins (stiffness and adhesion), the topography of the irradiated areas was analysed with atomic force microscopy (AFM). Further, the behaviour of the resins was analysed after ion milling of the surface of the sample with different ion currents. In conclusion, two resin formulations, Hard Plus and the mixture of Durcupan/Epon, emerged that were considerably less affected and reasonably stable in the electron/ion beam and thus suitable for the 3-D investigation of biological samples.

Three-Dimensional Angle Assessment and Plaque Distribution Classification in Left Main Disease: Impact of Geometry on Outcome.

Bifurcation geometry and plaque distribution in a diseased left main artery (LM) have the potential to drive operators' decisions regarding treatment strategies, techniques, and material selection. The three-dimensional (3D) geometry of the LM bifurcation typically results in specific patterns of plaque distribution. Plaque distribution may, in turn, significantly affect the procedural and long-term clinical and angiographic outcomes of LM percutaneous coronary intervention. Each LM bifurcation must be treated according to its unique anatomic and pathologic characteristics. Novel classification schemes of plaque distribution and 3D assessment may be valuable aids to obtaining a working picture of the bifurcation geometry.

Anatomical features and management of bioresorbable vascular scaffolds failure: A case series from the GHOST registry.

The Absorb bioresorbable vascular scaffold (Absorb BVS, Abbott Vascular, Santa Clara, California) promises to address some of the residual shortcomings of existing metallic stents, such as late events induced by permanent caging of the coronary vessel. Scaffold restenosis (ScR) of BVS has been poorly described so far and treatment strategies for this event remain to be codified. We report on a case series of 14 lesions in 12 patients presenting with ScR and discuss their anatomical features and management strategies. © 2015 Wiley Periodicals, Inc.

New insights on acute expansion and longitudinal elongation of bioresorbable vascular scaffolds in vivo and at bench test: a note of caution on reliance to compliance charts and nominal length.

OBJECTIVES: We performed systematic optical coherence tomography (OCT) analyses after bioresorbable vascular scaffolds (BVS) implantation in a "real world" setting aiming at evaluating scaffold expansion and longitudinal integrity. BACKGROUND: a comprehensive elucidation of BVS acute performance in the "real-world" setting is lacking. METHODS: acute BVS expansion compared with compliance chart information and longitudinal integrity were assessed in 29 patients (32 lesions) by OCT. In addition, bench experiments with four scaffolds were performed with different combinations of deployment pressures and tube stiffness. RESULTS: scaffold underexpansion, using compliance chart information as reference, was observed in 97% of OCT cross-sections in vivo; however, only 8.3% of the cross-section analyzed revealed BVS area <5 mm(2) . Calcified plaques were more common in the lowest (9.7%) compared with the mid (8.8%) and highest (6.3%) tertiles of scaffold expansion (P = 0.003 and P = 0.001 for lowest vs. mid, and lowest vs. highest, respectively). Seventeen (54.8%) scaffolds were elongated during implantation, but no signs of scaffold fracture were revealed. Elongation and impaired expansion were reproduced in the bench testing when the scaffold was deployed with high pressure in a hard tube. CONCLUSIONS: compliance chart information should not be used to predict final BVS dimensions in the clinical setting. While BVS expansion could be potentially impaired by calcified plaques, they may elongate during deployment. Bench experiments confirmed the elongation phenomenon when BVS were deployed with high pressure in hard tubes.

Time-lapse AFM imaging of DNA conformational changes induced by daunorubicin.

Cancer is a major health issue that absorbs the attention of a large part of the biomedical research. Intercalating agents bind to DNA molecules and can inhibit their synthesis and transcription; thus, they are increasingly used as drugs to fight cancer. In this work, we show how atomic force microscopy in liquid can characterize, through time-lapse imaging, the dynamical influence of intercalating agents on the supercoiling of DNA, improving our understanding of the drug's effect.

Left ventricular ejection fraction: clinical, pathophysiological, and technical limitations.

Risk stratification of cardiovascular death and treatment strategies in patients with heart failure (HF), the optimal timing for valve replacement, and the selection of patients for implantable cardioverter defibrillators are based on an echocardiographic calculation of left ventricular ejection fraction (LVEF) in most guidelines. As a marker of systolic function, LVEF has important limitations being affected by loading conditions and cavity geometry, as well as image quality, thus impacting inter- and intra-observer measurement variability. LVEF is a product of shortening of the three components of myocardial fibres: longitudinal, circumferential, and oblique. It is therefore a marker of global ejection performance based on cavity volume changes, rather than directly reflecting myocardial contractile function, hence may be normal even when myofibril's systolic function is impaired. Sub-endocardial longitudinal fibers are the most sensitive layers to ischemia, so when dysfunctional, the circumferential fibers may compensate for it and maintain the overall LVEF. Likewise, in patients with HF, LVEF is used to stratify subgroups, an approach that has prognostic implications but without a direct relationship. HF is a dynamic disease that may worsen or improve over time according to the underlying pathology. Such dynamicity impacts LVEF and its use to guide treatment. The same applies to changes in LVEF following interventional procedures. In this review, we analyze the clinical, pathophysiological, and technical limitations of LVEF across a wide range of cardiovascular pathologies.

Recurrent gastric antral vascular ectasia: a single center experience.

Introduction: Gastric antral vascular ectasia (GAVE) is a rare cause of chronic or acute gastrointestinal bleeding. This condition accounts for ∼4% of upper gastrointestinal bleeding cases. This disease is often associated with systemic diseases, such as liver cirrhosis, chronic kidney failure, autoimmune conditions, diabetes mellitus, hypothyroidism, and cardiovascular diseases. However, its etiopathogenesis remains controversial. Materials and method: We retrospectively reviewed the cases of GAVE treated at our digestive surgery unit. A total of nine patients were identified with a male/female ratio of 1.25:1 and an average age of 75.51 years (SD ± 9.85). All patients underwent endoscopic argon plasma coagulation (APC) treatment. At the time of the review, data on eight patients were available after 36 months of follow-up. Results: APC appears to be safe and effective for hemostasis of bleeding vascular ectasia. Only one (11.1%) patient required surgical intervention due to hemodynamic instability after multiple unsuccessful endoscopic treatments. No intraoperative and postoperative complication or bleeding relapse was experienced. Discussion: Based on our findings, we concluded that endoscopic APC is technically simple, but requires multiple re-interventions due to the incidence of relapses. Furthermore, larger randomized studies should be conducted to assess the role of elective surgery as the first intervention in stable patients with severe pathology and the timing of surgery after failed endoscopic treatment.

Correlating nanoscale motion and ATP production in healthy and favism erythrocytes: a real-time nanomotion sensor study.

Introduction: Red blood cells (RBCs) are among the simplest, yet physiologically relevant biological specimens, due to their peculiarities, such as their lack of nucleus and simplified metabolism. Indeed, erythrocytes can be seen as biochemical machines, capable of performing a limited number of metabolic pathways. Along the aging path, the cells' characteristics change as they accumulate oxidative and non-oxidative damages, and their structural and functional properties degrade. Methods: In this work, we have studied RBCs and the activation of their ATP-producing metabolism using a real-time nanomotion sensor. This device allowed time-resolved analyses of the activation of this biochemical pathway, measuring the characteristics and the timing of the response at different points of their aging and the differences observed in favism erythrocytes in terms of the cellular reactivity and resilience to aging. Favism is a genetic defect of erythrocytes, which affects their ability to respond to oxidative stresses but that also determines differences in the metabolic and structural characteristic of the cells. Results: Our work shows that RBCs from favism patients exhibit a different response to the forced activation of the ATP synthesis compared to healthy cells. In particular, the favism cells, compared to healthy erythrocytes, show a greater resilience to the aging-related insults which was in good accord with the collected biochemical data on ATP consumption and reload. Conclusion: This surprisingly higher endurance against cell aging can be addressed to a special mechanism of metabolic regulation that permits lower energy consumption in environmental stress conditions.