Elastin Content Research Articles

Fabrication and Characterization of Recombinant Silk-Elastin-Like-Protein (SELP) Fiber.

Silk-elastin-like-protein polymers (SELPs) are genetically engineered recombinant protein sequences consisting of repeating units of silk-like and elastin-like blocks. By combining these entities, it is shown that both the characteristic strength of silk and the temperature-dependent responsiveness of elastin can be leveraged to create an enhanced stimuli-responsive material. It is hypothesized that SELP behavior can be influenced by varying the silk-to-elastin ratio. If the responsiveness of the material at different ratios is significantly different, this would allow for the design of materials with specific temperature-based swelling and mechanical properties. This study demonstrates that SELP fiber properties can be controlled via a temperature transition dependent on the ratio of silk-to-elastin in the material. SELP fibers are experimentally wet spun from polymers with different ratios of silk-to-elastin and conditioned in either a below or above transition temperature (T t ) water bath prior to characterization. The fibers with higher elastin content showed more stimuli-responsive behavior compared to the fibers with lower elastin content in the hot (57-60°C) versus cold (4-7°C) environment, both computationally and experimentally. This work builds a foundation for developing SELP materials with well-characterized mechanical properties and responsive features.

Abstract 16752: Aortic Pulsatility, and Not Mean Arterial Pressure, is an Independent Determinant of Left Main Coronary Artery Disease

Introduction: Left main coronary artery(LMCA) disease is prognostically the most important lesion in the coronary tree. LMCA differs from the other coronaries in having high elastin content. As aortic blood pressure (BP) directly affects the heart and emerged as independent predictor of major cardiovascular events, there is heightened interest in exploring its relationship with coronary artery disease (CAD). Aortic pulsatility (AP) measurement eliminates the effect of cardiac output and vascular resistance and is shown to be an independent predictor of cardiovascular events in CAD. Hypothesis: As the left main is an elastic vessel, we hypothesized that pulsatile stress may be an independent determinant of disease in this artery. Methods: This was a prospective cohort study in patients undergoing coronary angiography for suspected CAD between the years 2011 and 2016(n=4633, 25% female) at King Abdul Aziz Cardiac Center, Riyadh, Saudi Arabia. We excluded patients with acute myocardial infarction, cardiogenic shock and significant valvular heart disease. Aortic systolic and diastolic BP were measured in the ascending aorta. MAP was obtained by direct integration of the BP curve. PP was calculated as the difference between systolic and diastolic BP. AP was calculated as PP divided by MAP. CAD was defined as > 50% stenosis in LMCA and the 3 major coronaries (left anterior descending, circumflex and right coronary artery). The AP in patients with LMCA disease was compared to those with non-obstructive, 1, 2 and 3-vd as well with LMCA and 3-vd. Results: Six percent of the study population had LMCA disease (mean age 60±11 years, 25% female). LMCA disease was associated with higher PP (69±22 vs. 58±18, p<0.0001) despite similar MAP (94±16 vs. 94.5±14, p=0.92) compared with non-LMCA disease. AP was significantly higher (0.72±0.30) in LMCA disease compared with; 3-vd( 0.63±0.32); 2-vd(0.61±0.28), 1-vd(0.58±0.31) and non-obstructive CAD (0.52±0.26) (p<0.0001). In a stepwise regression model, AP was an independent predictor of LMCA disease( R 2 =0.68,P<0.0001) when adjusted for potential confounders, including MAP, age and gender. Conclusions: We conclude that LMCA disease is independently associated with high aortic pulsatility compared with disease of other coronary vessels,including 3-vd. Considering aortic pulsatile stress to be an independent cardiovascular prognosticator, stiffness of the LMCA may play an important role in plaque formation, hitherto ignored. More research is warranted to test our proof-of-concept study, including outcome.

Contributions of Glycosaminoglycans to Collagen Fiber Recruitment in Constitutive Modeling of Arterial Mechanics

The contribution of glycosaminoglycans (GAGs) to the biological and mechanical functions of biological tissue has emerged as an important area of research. GAGs provide structural basis for the organization and assembly of extracellular matrix (ECM). The mechanics of tissue with low GAG content can be indirectly affected by the interaction of GAGs with collagen fibers, which have long been known to be one of the primary contributors to soft tissue mechanics. Our earlier study showed that enzymatic GAG depletion results in straighter collagen fibers that are recruited at lower levels of stretch, and a corresponding shift in earlier arterial stiffening (Mattson et al., 2016). In this study, the effect of GAGs on collagen fiber recruitment was studied through a structure-based constitutive model. The model incorporates structural information, such as fiber orientation distribution, content, and recruitment of medial elastin, medial collagen, and adventitial collagen fibers. The model was first used to study planar biaxial tensile stress-stretch behavior of porcine descending thoracic aorta. Changes in elastin and collagen fiber orientation distribution, and collagen fiber recruitment were then incorporated into the model in order to predict the stress-stretch behavior of GAG depleted tissue. Our study shows that incorporating early collagen fiber recruitment into the model predicts the stress-stretch response of GAG depleted tissue reasonably well (rms = 0.141); considering further changes of fiber orientation distribution does not improve the predicting capability (rms = 0.149). Our study suggests an important role of GAGs in arterial mechanics that should be considered in developing constitutive models.

Simultaneous Assessment of Cardiac Inflammation and Extracellular Matrix Remodeling after Myocardial Infarction.

Optimal healing of the myocardium following myocardial infarction (MI) requires a suitable degree of inflammation and its timely resolution, together with a well-orchestrated deposition and degradation of extracellular matrix (ECM) proteins. MI and SHAM-operated animals were imaged at 3,7,14 and 21 days with 3T magnetic resonance imaging (MRI) using a 19F/1H surface coil. Mice were injected with 19F-perfluorocarbon (PFC) nanoparticles to study inflammatory cell recruitment, and with a gadolinium-based elastin-binding contrast agent (Gd-ESMA) to evaluate elastin content. 19F MRI signal co-localized with infarction areas, as confirmed by late-gadolinium enhancement, and was highest 7days post-MI, correlating with macrophage content (MAC-3 immunohistochemistry) (ρ=0.89,P<0.0001). 19F quantification with in vivo (MRI) and ex vivo nuclear magnetic resonance (NMR) spectroscopy correlated linearly (ρ=0.58,P=0.020). T1 mapping after Gd-ESMA injection showed increased relaxation rate (R1) in the infarcted regions and was significantly higher at 21days compared with 7days post-MI (R1[s-1]:21days=2.8 [IQR,2.69-3.30] vs 7days=2.3 [IQR,2.12-2.5], P<0.05), which agreed with an increased tropoelastin content (ρ=0.89, P<0.0001). The predictive value of each contrast agent for beneficial remodeling was evaluated in a longitudinal proof-of-principle study. Neither R1 nor 19F at day 7 were significant predictors for beneficial remodeling (P=0.68;P=0.062). However, the combination of both measurements (R1<2.34Hz and 0.55≤19F≤1.85) resulted in an odds ratio of 30.0 (CI95%:1.41-638.15;P=0.029) for favorable post-MI remodeling. Multinuclear 1H/19F MRI allows the simultaneous assessment of inflammation and elastin remodeling in a murine MI model. The interplay of these biological processes affects cardiac outcome and may have potential for improved diagnosis and personalized treatment.

Chronic Nicotine Exposure Induces Murine Aortic Remodeling and Stiffness Segmentation-Implications for Abdominal Aortic Aneurysm Susceptibility.

Aim: Arterial stiffness is a significant risk factor for many cardiovascular diseases, including abdominal aortic aneurysms (AAA). Nicotine, the major active ingredient of e-cigarettes and tobacco smoke, induces acute vasomotor effects that may temporarily increase arterial stiffness. Here, we investigated the effects of long-term nicotine exposure on structural aortic stiffness.Methods: Mice (C57BL/6) were infused with nicotine for 40 days (20 mg/kg/day). Arterial stiffness of the thoracic (TS) and abdominal (AS) aortic segments was analyzed using ultrasound (PWV, pulse wave velocity) and ex vivo pressure myograph measurements. For mechanistic studies, aortic matrix-metalloproteinase (MMP) expression and activity as well as medial elastin architecture were analyzed.Results: Global aortic stiffness increased with nicotine. In particular, local stiffening of the abdominal segment occurred after 10 days, while thoracic aortic stiffness was only increased after 40 days, resulting in aortic stiffness segmentation. Mechanistically, nicotine exposure enhanced expression of MMP-2/-9 and elastolytic activity in both aortic segments. Elastin degradation occurred in both segments; however, basal elastin levels were higher in the thoracic aorta. Finally, MMP-inhibition significantly reduced nicotine-induced MMP activity, elastin destruction, and aortic stiffening.Conclusion: Chronic nicotine exposure induces aortic MMP expression and structural aortic damage (elastin fragmentation), irreversibly increasing aortic stiffness. This process predominantly affects the abdominal aortic segment, presumably due in part to a lower basal elastin content. This novel phenomenon may help to explain the role of nicotine as a major risk factor for AAA formation and has health implications for ECIGs and other modes of nicotine delivery.

Effect of PM2.5 environmental pollution on rat lung.

Particulate matter smaller than 2.5μm (PM2.5) is a continuing challenge to pulmonary health. Here, we investigated the mechanisms involved in PM2.5 exposure-induced acute lung injury in rats. We analyzed biochemical and morphological changes following a 2-week "real-world" exposure. And then we found that PM2.5 exposure increased the concentrations of total protein, malondialdehyde, hydrogen peroxide, nitric oxide, and soluble elastin in bronchoalveolar lavage fluid, levels of cytokines in blood, and expression of MMP-9 in airways. Further, alveolar macrophage and neutrophil counts increased following PM2.5 exposure, and edema and lung lesions were observed. Our results suggest that PM2.5 exposure can induce oxidative stress and acute inflammatory responses, which can damage the micro-environment and decrease the repair ability of the lung, resulting in tissue damage.

The tropoelastin and lysyl oxidase treatments increased the content of insoluble elastin in bioprosthetic heart valves.

Valvular heart diseases lead to over 300,000 heart valve replacements worldwide each year. Commercially available bioprosthetic heart valves (BHVs) are mostly made from porcine or bovine pericardiums which were crosslinked by glutaraldehyde (GLUT). However, valve failures can occur within 10 years due to progressive degradation and calcification. GLUT could crosslink collagen but it fails to stabilize elastin. In this current study, we developed a new BHVs preparation strategy named as "GLUT/TE/LOXL/EGCG" that utilizes exogenous tropoelastin (TE)/lysyl oxidase (LOXL) and epigallocatechin gallate (EGCG) to increase the elastin content as well as the stabilization of elastin. The feeding ratios of tropoelastin and lysyl oxidase were optimized. The contents of desmosine and insoluble elastin, biomechanics, cytotoxicity, hemocompatibility, in vivo componential stability and anti-calcification potential were characterized. Pericardiums with increased elastin content had improved the mechanical properties. GLUT/TE/LOXL/EGCG-treated pericardiums had similar cytotoxicity and coagulation properties compared to GLUT and GLUT/EGCG control. We demonstrated that GLUT/TE/LOXL/EGCG-treated pericardiums had high amount of insoluble elastin in 90 days' rat subdermal implantation model, and better resistance for calcification. This new tropoelastin and lysyl oxidase treatments strategy would be a promising method to make BHVs which have better structural stability and anti-calcification properties.

Structural characterization of four different naturally occurring porcine collagen membranes suitable for medical applications.

Collagen is the main structural element of connective tissues, and its favorable properties make it an ideal biomaterial for regenerative medicine. In dental medicine, collagen barrier membranes fabricated from naturally occurring tissues are used for guided bone regeneration. Since the morphological characteristics of collagen membranes play a crucial role in their mechanical properties and affect the cellular behavior at the defect site, in-depth knowledge of the structure is key. As a base for the development of novel collagen membranes, an extensive morphological analysis of four porcine membranes, including centrum tendineum, pericardium, plica venae cavae and small intestinal submucosa, was performed. Native membranes were analyzed in terms of their thickness. Second harmonic generation and two-photon excitation microscopy of the native membranes showed the 3D architecture of the collagen and elastic fibers, as well as a volumetric index of these two membrane components. The surface morphology, fiber arrangement, collagen fibril diameter and D-periodicity of decellularized membranes were investigated by scanning electron microscopy. All the membrane types showed significant differences in thickness. In general, undulating collagen fibers were arranged in stacked layers, which were parallel to the membrane surface. Multiphoton microscopy revealed a conspicuous superficial elastic fiber network, while the elastin content in deeper layers varied. The elastin/collagen volumetric index was very similar in the investigated membranes and indicated that the collagen content was clearly higher than the elastin content. The surface of both the pericardium and plica venae cavae and the cranial surface of the centrum tendineum revealed a smooth, tightly arranged and crumpled morphology. On the caudal face of the centrum tendineum, a compact collagen arrangement was interrupted by clusters of circular discontinuities. In contrast, both surfaces of the small intestinal submucosa were fibrous, fuzzy and irregular. All the membranes consisted of largely uniform fibrils displaying the characteristic D-banding. This study reveals similarities and relevant differences among the investigated porcine membranes, suggesting that each membrane represents a unique biomaterial suitable for specific applications.

A18474 Aortic Pulsatility, And Not Mean Arterial Pressure, Is An Independent Determinant Of Left Main Coronary Artery Disease

Objectives: Left main coronary artery(LMCA) disease is prognostically the most important lesion in the coronary tree.LMCA differs from the other coronaries in having a high elastin content. Aortic pulsatility (AP) eliminates the effect of cardiac output and vascular resistance and is shown to be an independent predictor of cardiovascular events in CAD.As the left main is an elastic vessel,we hypothesized that aortic pulsatile stress may be an independent determinant of disease in this artery,independent of mean arterial pressure(MAP). Methods: This was a prospective cohort study in patients undergoing coronary angiography between the years 2011 and 2016(n = 4633, 25% female) at King Abdul Aziz Cardiac Center, Riyadh, Saudi Arabia. Aortic systolic and diastolic BP were measured in the ascending aorta. MAP was obtained by direct integration of the BP curve.AP was calculated as PP divided by MAP.CAD was defined as > 50% stenosis in atleast one of the major coronary arteries.AP in patients with LMCA disease was compared to those with non-obstructive, 1, 2 and 3-vd as well with LMCA and 3-vd. Results: Six percent of the study population had LMCA disease(mean age 60 ± 11 years,25% female).LMCA disease was associated with higher PP(69 ± 22 vs.58 ± 18, p < 0.0001)despite similar MAP(94 ± 16 vs. 94.5 ± 14, p = 0.92)compared with non-LMCA disease.AP was significantly higher(0.72 ± 0.30) in LMCA disease compared with;3-vd (0.63 ± 0.32);2-vd(2-vd(0.61 ± 0.28),1-vd(0.58 ± 0.31)and non-obstructive CAD (0.52 ± 0.26) (p < 0.0001).In a stepwise regression model,AP was an independent predictor of LMCA disease(R2 = 0.68,P < 0.0001), adjusted for age, gender, BMI and MAP. Conclusion: Stiffness of the LMCA, independent of MAP, may play an important role in the pathophysiology of plaque formation, hitherto overlooked.

Regional mechanical and biochemical properties of the porcine cortical meninges

The meninges are pivotal in protecting the brain against traumatic brain injury (TBI), an ongoing issue in most mainstream sports. Improved understanding of TBI biomechanics and pathophysiology is desirable to improve preventative measures, such as protective helmets, and advance our TBI diagnostic/prognostic capabilities. This study mechanically characterised the porcine meninges by performing uniaxial tensile testing on the dura mater (DM) tissue adjacent to the frontal, parietal, temporal, and occipital lobes of the cerebellum and superior sagittal sinus region of the DM. Mechanical characterisation revealed a significantly higher elastic modulus for the superior sagittal sinus region when compared to other regions in the DM. The superior sagittal sinus and parietal regions of the DM also displayed local mechanical anisotropy. Further, fatigue was noted in the DM following ten preconditioning cycles, which could have important implications in the context of repetitive TBI. To further understand differences in regional mechanical properties, regional variations in protein content (collagen I, collagen III, fibronectin and elastin) were examined by immunoblot analysis. The superior sagittal sinus was found to have significantly higher collagen I, elastin, and fibronectin content. The frontal region was also identified to have significantly higher collagen I and fibronectin content while the temporal region had increased elastin and fibronectin content. Regional differences in the mechanical and biochemical properties along with regional tissue thickness differences within the DM reveal that the tissue is a non-homogeneous structure. In particular, the potentially influential role of the superior sagittal sinus in TBI biomechanics warrants further investigation. Statement of significanceThis study addresses the lack of regional mechanical analysis of the cortical meninges, particularly the dura mater (DM), with accompanying biochemical analysis. To mechanically characterise the stiffness of the DM by region, uniaxial tensile testing was carried out on the DM tissue adjacent to the frontal, parietal, temporal and occipital lobes along with the DM tissue associated with the superior sagittal sinus. To the best of the authors’ knowledge, the work presented here identifies, for the first time, the heterogeneous nature of the DM’s mechanical stiffness by region. In particular, this study identifies the significant difference in the stiffness of the DM tissue associated with the superior sagittal sinus when compared to the other DM regions. Constitutive modelling was carried out on the regional mechanical testing data for implementation in Finite Element models with improved biofidelity. This work also presents the first biochemical analysis of the collagen I and III, elastin, and fibronectin content within DM tissue by region, providing useful insights into the accompanying macro-scale biomechanical data.

Cell-seeded 3D scaffolds as in vitro models for electroporation

Electroporation of cells is usually studied using cell suspensions or monolayer cultures. 3D scaffolds for cell culture have been recently designed in order to reproduce in vitro the complex and multifactorial environment experimented in vivo by cells. In fact, it is well known that 2D cell cultures are not able to simulate the complex interactions between the cells and their extracellular matrix (ECM). Recently, some examples of 3D models, like spheroids, have been investigated also in the electroporation field. Spheroids have been proposed in electrochemotherapy (ECT) studies to mimic tumor in vivo conditions: they are easy-to-handle 3D models but their sensitivity to electric field pulses depends from their diameter and, more interestingly, despite being relevant for intercellular junctions, they are not so much so for cell-ECM interactions.In this work, we propose a 3D macroscopic myxoid matrix for cell culture that would mimic the in vivo environment of myxoid stroma tumors. The myxoid stroma consists of abundant basic substances with large amounts of glycosaminoglycans (hyaluronic acid) and proteoglycans, poor collagen fibers and no elastin content. In the proposed approach, tumor cells seeded on 3D scaffolds mimic of myxoid stroma can establish both cell-cell and cell-ECM 3D interactions.The MCF7 cells (human breast adenocarcinoma cell line) were seeded in complete culture medium. Cell cultures were incubated at 37 °C for either 24 h, 3 days or 7 day. Some samples were used to assess cell vitality using 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT) test and others for electroporation tests and for histopathological analysis. The electroporation has been verified by the fluorescent dye Propidium cellular uptake.The proposed myxoid stroma scaffold induces cell proliferation and shows fibrous structures produced by cells, the concentration of which increases with culture time. The proposed matrix will be used for further investigations as a new scaffold for cell culture. Tumor cells grown into these new scaffolds will be used to evaluate electroporation including the stroma effect.

Local upregulation of interleukin-1 beta in aortic dissecting aneurysm: correlation with matrix metalloproteinase-2, 9 expression and biomechanical decrease

Our goal was to examine whether interleukin-1 beta (IL-1β) originates locally and its possible relationship with matrix metalloproteinases (MMPs), apoptosis, elastin fibres and biomechanics in aortic dissecting aneurysms (DAs). Aortic DAs were induced in 24 rats with β-aminopropionitrile (BAPN); another 12 rats without BAPN were designated as controls. Then IL-1β levels were measured both in the circulation and in local aortic specimens. The expression of MMP-2 and MMP-9 and Victoria blue and TUNEL staining were also detected. Biomechanical parameters such as the elasticity modulus were used to detect the biomechanical changes in the aortic wall. The correlation of IL-1β, MMP-2, MMP-9, apoptosis and biomechanical properties was analysed. Seventeen rats (17/24, 71%) in the BAPN-treated group died of DA rupture. IL-1β levels were dramatically increased in the DA specimens but not in the circulation. Victoria blue staining confirmed the formation of the DA and the reduction of elastin content after induction by BAPN. The extent of apoptosis in the aortic media was dramatically higher in rats with BAPN-induced DA than that in the control group and that in rats treated with BAPN but without DA. MMP-2 and MMP-9 levels were significantly increased in BAPN-treated rats compared to the controls, but no statistical significance was found between rats with and without DA. There were significant differences in biomechanical parameters, such as the elasticity modulus. Among the 3 groups, IL-1β was positively correlated with MMP-2 and MMP-9 levels and with the elasticity modulus but not with apoptosis. Local IL-1β might participate in the formation of aortic DA through the upregulation of MMP-2 and MMP-9 and the breakage of elastin fibres, which finally weakens the biomechanical properties of the aortic wall.

A Three-Dimensional Collagen-Elastin Scaffold for Heart Valve Tissue Engineering.

Since most of the body’s extracellular matrix (ECM) is composed of collagen and elastin, we believe the choice of these materials is key for the future and promise of tissue engineering. Once it is known how elastin content of ECM guides cellular behavior (in 2D or 3D), one will be able to harness the power of collagen-elastin microenvironments to design and engineer stimuli-responsive tissues. Moreover, the implementation of such matrices to promote endothelial-mesenchymal transition of primary endothelial cells constitutes a powerful tool to engineer 3D tissues. Here, we design a 3D collagen-elastin scaffold to mimic the native ECM of heart valves, by providing the strength of collagen layers, as well as elasticity. Valve interstitial cells (VICs) were encapsulated in the collagen-elastin hydrogels and valve endothelial cells (VECs) cultured onto the surface to create an in vitro 3D VEC-VIC co-culture. Over a seven-day period, VICs had stable expression levels of integrin β1 and F-actin and continuously proliferated, while cell morphology changed to more elongated. VECs maintained endothelial phenotype up to day five, as indicated by low expression of F-actin and integrin β1, while transformed VECs accounted for less than 7% of the total VECs in culture. On day seven, over 20% VECs were transformed to mesenchymal phenotype, indicated by increased actin filaments and higher expression of integrin β1. These findings demonstrate that our 3D collagen-elastin scaffolds provided a novel tool to study cell-cell or cell-matrix interactions in vitro, promoting advances in the current knowledge of valvular endothelial cell mesenchymal transition.

Upregulation of Skin-Aging Biomarkers in Aged NHDF Cells by a Sucrose Ester Extract from the Agroindustrial Waste of Physalis peruviana Calyces.

As part of a search for new sustainable plant sources of valuable compounds, the EtOAc extract of the discarded calyces of Physalis peruviana fruit was selected for its significant antiaging activity. Eight new sucrose esters (SEs), named peruvioses F-M (1-8), along with three known SEs, peruvioses A (9), peruviose B (10), and nicandrose D (11), were isolated. Their structures were elucidated by comprehensive analyses of their NMR and MS data. A global fragmentation pattern of these SEs was established from their MS data. The SE extract (SEE) at a concentration of 0.5 mg L-1 upregulated multiple skin-aging biomarkers, namely, collagen I, elastin, and fibrillin-1, in aged normal human dermal fibroblast cells. A 36% increase in collagen I was observed. The elastin and fibrillin-1 contents were fully recovered, and an increase of at least 10% in the production of elastin was observed.

Copper as the most likely pathogenic divergence factor between lung fibrosis and emphysema

Although fibrosis and emphysema are in many ways on opposite ends of the pulmonary parenchymal disease spectrum, they seem to share common pathomechanistic steps. This is illustrated by the coexistence of both entities in lungs of individuals with combined pulmonary fibrosis and emphysema. Macroproteins elastin and collagen are major constituents of the pulmonary extracellular matrix. The prevailing paradigm states that emphysema is caused by an imbalance between destructive proteolytic and protective antiproteolytic enzymes leading to accelerated degradation of elastin fibers in the lungs. Rates of elastin breakdown, however, are equally enhanced in patients with idiopathic pulmonary fibrosis (IPF) and emphysema. Excessive accumulation of collagen is a hallmark of IPF. Surprisingly, collagen levels in the lung parenchyma of patients with emphysema are also higher than in controls. The concentration of elastin fibers is elevated in fibrotic lungs, despite accelerated elastinolysis, suggesting that elastin repair is also enhanced in IPF. Since elastin concentrations are reduced in emphysematous lungs, the factor of divergence between emphysema and fibrosis seems to be the degree of elastin repair. Multiple elastin repair steps can be deduced of which tropoelastin synthesis and crosslinking of tropoelastin polymers by the copper dependent enzyme lysyl oxidase seem to be the most important ones. We suspect that the distinction in the pathogeneses of lung fibrosis and emphysema depends on the local availability of copper to activate sufficient lysyl oxidase for elastin crosslinking, and suggest assessing the effects of inhalation therapy with copper plus heparin in emphysema and heparin monotherapy in IPF.

Label-free assessment of carotid artery biochemical composition using fiber-based fluorescence lifetime imaging.

Novel diagnostic tools with the ability to monitor variations in biochemical composition and provide benchmark indicators of vascular tissue maturation are needed to create functional tissue replacements. We investigated the ability of fiber-based, label-free multispectral fluorescent lifetime imaging (FLIm) to quantify the anatomical variations in biochemical composition of native carotid arteries and validated these results against biochemical assays. FLIm-derived parameters in spectral band 415–455 nm correlated with tissue collagen content (R2 = 0.64) and cell number (R2 = 0.61) and in spectral band 465–553 nm strongly correlated with elastin content (R2 = 0.89). These results suggest that FLIm holds great potential for assessing vascular tissue maturation and functional properties based on tissue autofluorescence.

Elastin Content and Distribution in Endothelial Keratoplasty Tissue Determines Direction of Scrolling

Descemet membrane endothelial keratoplasty (DMEK) and pre-Descemet endothelial keratoplasty (PDEK) tissues always scroll with the endothelial cells (EC) outside. We designed a study to understand the reason for this behavior. Experimental study. Elastin content in Descemet membrane (DM), pre-Descemet layer (PDL), central and peripheral stroma, sclera, and trabecular meshwork were measured by the Fastin elastin assay kit. Distribution of elastin in DM, PDL, and anterior lens capsule (ALC) were examined by immunohistology. The effect of recombinant elastase enzyme and the effect of complete removal of EC and epithelial cells on the scrolling of DM and ALC, respectively, were studied. PDL showed the highest elastin content among the different tissues studied. Elastin localized as a distinct anterior band in the DM and was uniformly distributed in the PDL demarcating the latter from corneal stroma. Enzymatic treatment of DM with elastase reversed scrolling and corresponded with degradation or disappearance of elastin. Removal of EC did not affect the direction of scrolling. ALC behaved in the same manner with regard to distribution of elastin, scrolling, and removal of epithelial cells. This pattern of elastin distribution in DM explains why DMEK and PDEK tissues always scroll with the EC outside. This behavior is not influenced by the EC. High elastin content and uniform distribution in the PDL suggest a structural difference from the posterior stroma.

IGF-1 Deficiency Promotes Pathological Remodeling of Cerebral Arteries: A Potential Mechanism Contributing to the Pathogenesis of Intracerebral Hemorrhages in Aging.

Clinical and experimental studies show that age-related decline in circulating insulin-like growth factor-1 (IGF-1) levels promotes the pathogenesis of intracerebral hemorrhages, which critically contribute to the development of vascular cognitive impairment and disability in older adults. Yet, the mechanisms by which IGF-1 deficiency compromises structural integrity of the cerebral vasculature are not completely understood. To determine the role of IGF-1 deficiency in pathological remodeling of middle cerebral arteries (MCAs), we compared alterations in vascular mechanics, morphology, and remodeling-related gene expression profile in mice with liver-specific knockdown of IGF-1 (Igf1f/f + TBG-Cre-AAV8) and control mice with or without hypertension induced by angiotensin-II treatment. We found that IGF-1 deficiency resulted in thinning of the media and decreased wall-to-lumen ratio in MCAs. MCAs of control mice exhibited structural adaptation to hypertension, manifested as a significant increase in wall thickness, vascular smooth muscle cell (VSMC) hypertrophy, decreased internal diameter and up-regulation of extracellular matrix (ECM)-related genes. IGF-1 deficiency impaired hypertension-induced adaptive media hypertrophy and dysregulated ECM remodeling, decreasing elastin content and attenuating adaptive changes in ECM-related gene expression. Thus, circulating IGF-1 plays a critical role in maintenance of the structural integrity of cerebral arteries. Alterations of VSMC phenotype and pathological remodeling of the arterial wall associated with age-related IGF-1 deficiency have important translational relevance for the pathogenesis of intracerebral hemorrhages and vascular cognitive impairment in elderly hypertensive patients.

Relaxin receptor deficiency promotes vascular inflammation and impairs outward remodeling in arteriovenous fistulas.

The pathophysiology of arteriovenous fistula (AVF) maturation failure is not completely understood but impaired outward remodeling (OR) and intimal hyperplasia are thought to be contributors. This adverse vascular response after AVF surgery results from interplay between vascular smooth muscle cells (VSMCs), the extracellular matrix (ECM), and inflammatory cells. Relaxin (RLN) is a hormone that acts on the vasculature via interaction with RLN/insulin-like peptide family receptor 1 (RXFP1), resulting in vasodilatation, ECM remodeling, and decreased inflammation. In the present study, we evaluated the consequences of RXFP1 knockout ( Rxfp1-/-) on AVF maturation in a murine model of AVF failure. Rxfp1-/- mice showed a 22% decrease in vessel size at the venous outflow tract 14 d after AVF surgery. Furthermore, a 43% increase in elastin content was observed in the lesions of Rxfp1-/- mice and coincided with a 41% reduction in elastase activity. In addition, Rxfp1-/- mice displayed a 6-fold increase in CD45+ leukocytes, along with a 2-fold increase in monocyte chemoattractant protein 1 (MCP1) levels, when compared with wild-type mice. In vitro, VSMCs from Rxfp1-/- mice exhibited a synthetic phenotype, as illustrated by augmentation of collagen, fibronectin, TGF-β, and platelet-derived growth factor mRNA. In addition, VSMCs derived from Rxfp1-/- mice showed a 5-fold increase in cell migration. Finally, RXFP1 and RLN expression levels were increased in human AVFs when compared with unoperated cephalic veins. In conclusion, RXFP1 deficiency hampers elastin degradation and results in induced vascular inflammation after AVF surgery. These processes impair OR in murine AVF, suggesting that the RLN axis could be a potential therapeutic target for promoting AVF maturation.-Bezhaeva, T., de Vries, M. R., Geelhoed, W. J., van der Veer, E. P., Versteeg, S., van Alem, C. M. A., Voorzaat, B. M., Eijkelkamp, N., van der Bogt, K. E., Agoulnik, A. I., van Zonneveld, A.-J., Quax, P. H. A., Rotmans, J. I. Relaxin receptor deficiency promotes vascular inflammation and impairs outward remodeling in arteriovenous fistulas.

Differential Stiffening between the Abdominal and Thoracic Aorta: Effect of Salt Loading in Stroke-Prone Hypertensive Rats

Central artery stiffening is recognized as a cardiovascular risk. The effects of hypertension and aging have been shown in human and animal models but the effect of salt is still controversial. We studied the effect of a high-salt diet on aortic stiffness in salt-sensitive spontaneously hypersensitive stroke-prone rats (SHRSP). Distensibility, distension, and β-stiffness were measured at thoracic and abdominal aortic sites in the same rats, using echotracking recording of the aortic diameter coupled with blood pressure (BP), in SHRSP-salt (5% salted diet, 5 weeks), SHRSP, and normotensive Wistar-Kyoto (WKY) rats. Hemodynamic parameters were measured at BP matched to that of WKY. Histological staining and immunohistochemistry were used for structural analysis. Hemodynamic isobaric parameters in SHRSP did not differ from WKY and only those from the abdominal aorta of SHRSP-salt presented decreased distensibility and increased stiffness compared with WKY and SHRSP. The abdominal and thoracic aortas presented similar thickening, increased fibrosis, and remodeling with no change in collagen content. SHRSP-salt presented a specific increased elastin disarray at the abdominal aorta level but a decrease in elastin content in the thoracic aorta. This study demonstrates the pro-stiffening effect of salt in addition to hypertension; it shows that only the abdominal aorta presents a specific pressure-independent stiffening, in which elastin disarray is likely a key mechanism.