Matrix Elastin Research Articles

Investigating The Role of Elastin and Extracellular Matrix Damage in Cardiovascular Calcification

Although calcification in the cardiovascular system is highly studied, the mechanisms behind it are not well understood. Current proposed mechanisms focus on cellular processes leading to, or controlling the unwanted mineralization in soft tissues. However, extracellular components such as collagen and elastin fundamentally regulate the mechanical properties of heart tissues. Here, we report on a toolkit to control the composition of tissues through the selective digestion of extracellular matrix (ECM) components, which can be used to design disease-specific in vitro models. Using this technique, we show that elastin as well as matrix tissue damage may play major role in cardiovascular calcification. This study highlights a novel approach to understand the role of proteins in soft tissue calcifications and may lead to the development of strategies to treat and prevent these unwanted pathological disorders.

Residual Strain and Joint Pressurization Maintain Collagen Tension for On-Joint Lumbar Facet Capsular Ligaments.

Modeling the lumbar facet capsular ligament's (FCL) mechanical behavior under various physiological motions has often been a challenge due to limited knowledge about the on-joint in situ ligament state arising from attachment to the bone or other internal loads. Building on prior work, this study presents an enhanced computational model of the lumbar facet capsular ligament by incorporating residual strain and joint pressurization strain, factors neglected in prior models. Further, the model can predict strain and stress distribution across the ligament under various spinal motions, highlighting the influence of the ligament's attachment to the bone, internal synovial fluid pressurization, and distribution of collagen fiber alignment on the overall mechanical response of the ligament. Joint space inflation was found to influence the total observed stress and strain fields, both at rest and during motion. A significant portion of the ligament was found to be in tension, even in the absence of external load. Additionally, the model's ability to account for residual strain offers a more realistic portrayal of the collagen fibers and elastin matrix's role in ligament mechanics. We conclude that (1) computational models of the lumbar facet capsular ligament should not assume that the ligament is unloaded when the joint is in its neutral position, and (2) the ligament is nearly always in tension, which may be important in terms of its long-term growth and remodeling.

Impact of Cyclic Strain on Elastin Synthesis in a 3D Human Myometrial Culture Model.

The synthesis and assembly of mature, organized elastic fibers remains a limitation to the clinical use of many engineered tissue replacements. There is a critical need for a more in-depth understanding of elastogenesis regulation for the advancement of methods to induce and guide production of elastic matrix structures in engineered tissues that meet the structural and functional requirements of native tissue. The dramatic increase in elastic fibers through normal pregnancy has led us to explore the potential role of mechanical stretch in combination with pregnancy levels of the steroid hormones 17β-estradiol and progesterone on elastic fiber production by human uterine myometrial smooth muscle cells in a three-dimensional (3D) culture model. Opposed to a single strain regimen, we sought to better understand how the amplitude and frequency parameters of cyclic strain influence elastic fiber production in these myometrial tissue constructs (MTC). Mechanical stretch was applied to MTC at a range of strain amplitudes (5%, 10%, and 15% at 0.5 Hz frequency) and frequencies (0.1 Hz, 0.5 Hz, 1 Hz, and constant 0 Hz at 10% amplitude), with and without pregnancy-level hormones, for 6 days. MTC were assessed for cell proliferation, matrix elastin protein content, and expression of the main elastic fiber genes, tropoelastin (ELN) and fibrillin-1 (FBN1). Significant increases in elastin protein and ELN and FBN1 mRNA were produced from samples subjected to a 0.5 Hz, 10% strain regimen, as well as samples stretched at higher amplitude (15%, 0.5 Hz) and higher frequency (1 Hz, 10%); however, no significant effects because of third-trimester mimetic hormone treatment were determined. These results establish that a minimum level of strain is required to stimulate the synthesis of elastic fiber components in our culture model and show this response can be similarly enhanced by increasing either the amplitude or frequency parameter of applied strain. Further, our results demonstrate strain alone is sufficient to stimulate elastic fiber production and suggest hormones may not be a significant factor in regulating elastin synthesis. This 3D culture model will provide a useful tool to further investigate mechanisms underlying pregnancy-induced de novo elastic fiber synthesis and assembly by uterine smooth muscle cells.

Skin graft on a collagen and elastin matrix fixed with negative pressure in a complicated pediatric dog bite: case report

Animal bites represent a significant source of morbidity in the pediatric population, with dogs being the primary cause. Highlighting the need for innovative solutions to effectively tackle the issue and enhance both functional and aesthetic outcomes. Dog bites, marked by a heightened morbidity rate, present an inherent infection risk owing to the considerable bacterial load present in saliva. The fundamental approach to treatment revolves around preventing infection through meticulous washing, debridement, and the timely implementation of early primary closure. We detail the management of a pediatric patient with a dog bite on the pelvic limb, resulting in a challenging-to-manage wound. This case was successfully treated with a skin graft on a bed of collagen matrix, secured with negative pressure therapy. The choice of the appropriate technique for reconstructing skin defects depends on their depth and size. Consequently, combining these three techniques yielded a favorable functional and aesthetic outcome for the patient. This case report emphasizes the relevance of employing different techniques in managing complicated dog bites.

Inhibition of the histone methyltransferase EZH2 induces vascular stiffness.

Vascular stiffness increases with aging, obesity and hypertension and predicts cardiovascular risk. The levels of histone H3-lysine-27 methylation (H3K27me) and the histone methyltransferase EZH2 both decrease in aging vessels, driving vascular stiffness. The impact of EZH2 inhibitors on vascular stiffness is unknown. We tested the hypothesis that the EZH2 inhibitor GSK126, currently in development for cancer treatment, increases vascular stiffness and explored underlying molecular mechanisms. Young (3 month) and middle-aged (12 month) male mice were treated with GSK126 for 1-2 months and primary human aortic smooth muscle cells (HASMCs) from young male and female donors were treated with GSK126 for 24-48 h. Stiffness was measured in vivo by pulse wave velocity and in vitro by atomic force microscopy (AFM) and vascular structure was quantified histologically. Extracellular matrix proteins were studied by qRT-PCR, immunoblotting, zymography and chromatin immunoprecipitation. GSK126 treatment decreased H3K27 methylation (H3K27me) and increased acetylation (H3K27ac) in mouse vessels and in HASMCs. In GSK126-treated mice, aortic stiffness increased without changes in vascular fibrosis. EZH2 inhibition enhanced elastin fiber degradation and matrix metalloprotease-2 (MMP2) expression. In HASMCs, GSK126 treatment increased synthetic phenotype markers and intrinsic HASMCs stiffness by AFM with altered cytoskeletal structure and increased nuclear actin staining. GSK126 also increased MMP2 protein expression, activity and enrichment of H3K27ac at the MMP2 promoter in HASMCs. GSK126 causes vascular stiffening, inducing MMP2 activity, elastin degradation, and modulation of SMC phenotype and cytoskeletal stiffness. These findings suggest that EZH2 inhibitors used to treat cancer could negatively impact the vasculature by enhancing stiffness and merits examination in human trials.

Outcomes of lower extremity arterial bypass using the Human Acellular Vessel in patients with chronic limb-threatening ischemia

ObjectivePatients with chronic limb-threatening ischemia (CLTI) and no great saphenous vein to use as a conduit for arterial bypass have a high risk for amputation despite advances in medical and endovascular therapies. This report presents findings from a U.S. Food and Drug Administration (FDA) supported study of the Human Acellular Vessel (HAV) (Humacyte Inc.) used as a conduit for arterial bypass in patients with CLTI and inadequate or absent autologous conduit. MethodsThe HAV is a 6-mm, 40-cm vessel created from human vascular smooth muscle cells seeded onto a polyglycolic acid scaffold pulsed in a bioreactor for 8 weeks as cells proliferate and the scaffold dissolves. The resultant vessel is decellularized, creating a nonimmunogenic conduit composed of collagen, elastin, and extracellular matrix. The FDA issued an Investigational New Drug for an intermediate-sized, single-center study of the HAV under the agency's Expanded Access Program in patients with advanced CLTI and inadequate or absent autologous conduit. Technical results and clinical outcomes were analyzed and reported. ResultsBetween March 2021 and July 2023, 29 patients (20 males; mean age, 71 ± 11 years) underwent limb salvage operation using the HAV as a bypass conduit. Most patients had advanced CLTI (Rutherford class 5/6 in 72%; wound, ischemia, and foot infection stage 3/4 in 83%), and 97% had previously failed revascularization(s) of the extremity. Two HAVs were sewn together to attain the needed bypass length in 24 patients (83%). Bypasses were to tibial arteries in 23 patients (79%) and to the popliteal artery in 6 (21%). Technical success was 100%, and the 30-day mortality rate was 7% (2 patients). With 100% follow-up (median, 9.3 months), the limb salvage rate was 86% (25/29 patients). There were 16 reinterventions to restore secondary patency, of which 15 (94%) were successful. Primary and secondary patency of the HAV at 9 months were 59% and 71%, respectively. ConclusionsThe HAV has demonstrated short- to intermediate-term safety and efficacy as an arterial bypass conduit in a complex cohort of patients with limb-threatening ischemia and no autologous options. This experience using the FDA’s Expanded Access Program provides real-world data to inform regulatory deliberations and future trials of the HAV, including the study of the vessel as a first-line bypass conduit in less severe cases of chronic limb ischemia.

In-situ phase separation constructing robust hydrophobic ionogels with multifunction

Ionogels, as the promising alternative to hydrogels, have garnered great attention due to their excellent ionic conductivity, thermal stability, and non-volatility. However, the design and construction of robust hydrophobic ionogel still is a significant challenge at present, especially with a facile method. Herein, inspired by the two-phase structure of the dermis in human skin (stiff collagen fiber scaffold embedded in soft elastin matrix), a one-pot in-situ phase separation design strategy is proposed to construct the robust hydrophobic ionogel through the random copolymerization of hydrophobic monomers with the solubility difference in hydrophobic ionic liquid (IL). The resultant ionogel forms a homogeneously dual phase network containing solvent-rich soft domains and solvent-poor hard domains. The mechanical robustness can be synergistically achieved by the hard domain strong skeleton and the soft domain elastic matrix. Benefiting from multiple reversible interactions including hydrogen bonding, ion–dipole and ion-ion interactions, and physical entanglement, the ionogel shows strong IL retention, excellent self-healing, and regenerating abilities, while can adhere to different substrates. The unique phase separation structure brings temperature-dependent transparency and high-damping features. The ionogel also presents high ionic conductivity due to the abundance of movable ions. As application demonstrations, the ionogel is successfully used for dynamic pattern encryption, shock attenuation, and strain sensing. What’s more, this strategy can be also applied to improve the mechanical properties of hydrophobic ionogels based on other monomers. Hence, this work establishes an effective and general design strategy for hydrophobic ionogels demanding versatility.

C-reactive protein deficiency ameliorates experimental abdominal aortic aneurysms.

C-reactive protein (CRP) levels are elevated in patients with abdominal aortic aneurysms (AAA). However, it has not been investigated whether CRP contributes to AAA pathogenesis. CRP deficient and wild type (WT) male mice were subjected to AAA induction via transient intra-aortic infusion of porcine pancreatic elastase. AAAs were monitored by in situ measurements of maximal infrarenal aortic external diameters immediately prior to and 14 days following elastase infusion. Key AAA pathologies were assessed by histochemical and immunohistochemical staining procedures. The influence of CRP deficiency on macrophage activation was evaluated in peritoneal macrophages in vitro. CRP protein levels were higher in aneurysmal than that in non-aneurysmal aortas. Aneurysmal aortic dilation was markedly suppressed in CRP deficient (aortic diameter: 1.08 ± 0.11mm) as compared to WT (1.21 ± 0.08mm) mice on day 14 after elastase infusion. More medial elastin was retained in CRP deficient than in WT elastase-infused mice. Macrophage accumulation was significantly less in aneurysmal aorta from CRP deficient than that from WT mice. Matrix metalloproteinase 2 expression was also attenuated in CRP deficient as compared to WT aneurysmal aortas. CRP deficiency had no recognizable influence on medial smooth muscle loss, lymphocyte accumulation, aneurysmal angiogenesis, and matrix metalloproteinase 9 expression. In in vitro assays, mRNA levels for tumor necrosis factor α and cyclooxygenase 2 were reduced in lipopolysaccharide activated peritoneal macrophages from CRP deficient as compared to wild type mice. CRP deficiency suppressed experimental AAAs by attenuating aneurysmal elastin destruction, macrophage accumulation and matrix metalloproteinase 2 expression.

Constituent-based quasi-linear viscoelasticity: a revised quasi-linear modelling framework to capture nonlinear viscoelasticity in arteries

Arteries exhibit fully nonlinear viscoelastic behaviours (i.e. both elastically and viscously nonlinear). While elastically nonlinear arterial models are well established, effective mathematical descriptions of nonlinear viscoelasticity are lacking. Quasi-linear viscoelasticity (QLV) offers a convenient way to mathematically describe viscoelasticity, but its viscous linearity assumption is unsuitable for whole-wall vascular applications. Conversely, application of fully nonlinear viscoelastic models, involving deformation-dependent viscous parameters, to experimental data is impractical and often reduces to identifying specific solutions for each tested loading condition. The present study aims to address this limitation: By applying QLV theory at the wall constituent rather than at the whole-wall level, the deformation-dependent relative contribution of the constituents allows to capture nonlinear viscoelasticity with a unique set of deformation-independent model parameters. Five murine common carotid arteries were subjected to a protocol of quasi-static and harmonic, pseudo-physiological biaxial loading conditions to characterise their viscoelastic behaviour. The arterial wall was modelled as a constrained mixture of an isotropic elastin matrix and four families of collagen fibres. Constituent-based QLV was implemented by assigning different relaxation functions to collagen- and elastin-borne parts of the wall stress. Nonlinearity in viscoelasticity was assessed via the pressure dependency of the dynamic-to-quasi-static stiffness ratio. The experimentally measured ratio increased with pressure, from 1.03 pm 0.03 (mean pm standard deviation) at 80–40 mmHg to 1.58 pm 0.22 at 160–120 mmHg. Constituent-based QLV captured well this trend by attributing the wall viscosity predominantly to collagen fibres, whose recruitment starts at physiological pressures. In conclusion, constituent-based QLV offers a practical and effective solution to model arterial viscoelasticity.

Fluid-structure interaction modeling of compliant aortic valves using the lattice Boltzmann CFD and FEM methods.

The lattice Boltzmann method (LBM) has been increasingly used as a stand-alone CFD solver in various biomechanical applications. This study proposes a new fluid-structure interaction (FSI) co-modeling framework for the hemodynamic-structural analysis of compliant aortic valves. Toward that goal, two commercial software packages are integrated using the lattice Boltzmann (LBM) and finite element (FE) methods. The suitability of the LBM-FE hemodynamic FSI is examined in modeling healthy tricuspid and bicuspid aortic valves (TAV and BAV), respectively. In addition, a multi-scale structural approach that has been employed explicitly recognizes the heterogeneous leaflet tissues and differentiates between the collagen fiber network (CFN) embedded within the elastin matrix of the leaflets. The CFN multi-scale tissue model is inspired by monitoring the distribution of the collagen in 15 porcine leaflets. Different simulations have been examined, and structural stresses and resulting hemodynamics are analyzed. We found that LBM-FE FSI approach can produce good predictions for the flow and structural behaviors of TAV and BAV and correlates well with those reported in the literature. The multi-scale heterogeneous CFN tissue structural model enhances our understanding of the mechanical roles of the CFN and the elastin matrix behaviors. The importance of LBM-FE FSI also emerges in its ability to resolve local hemodynamic and structural behaviors. In particular, the diastolic fluctuating velocity phenomenon near the leaflets is explicitly predicted, providing vital information on the flow transient nature. The full closure of the contacting leaflets in BAV is also demonstrated. Accordingly, good structural kinematics and deformations are captured for the entire cardiac cycle.

Neutrophil Elastase in the Pathogenesis of Chronic Obstructive Pulmonary Disease: A Review

Abstract: Chronic obstructive pulmonary disease (COPD) is one of the leading causes of mortality globally. It is associated with a low quality of life and socio-economic burden. Airway destruction in COPD pathogenesis is primarily due to the three mechanisms: protease-antiprotease imbalance, chronic airway inflammation, and oxidative stress, which is triggered by exposure to harmful particles, such as cigarette smoking. Neutrophil elastase (NE), a serine protease stored in azurophilic granules of neutrophils, actively participates in airway remodeling and microbiocidal activity. It hydrolyzes elastin, collagen, and other vital extracellular matrix proteins (EMP) in the respiratory tissue. In addition, neutrophil elastase activates other principal proteinases such as matrix metalloprotease (MMP)-2, MMP-9, Cathepsin B, Meprin α protease, and Calpain that amplify EMP degradation. Macrophage, the primary leukocyte, responsible for lung parenchymal inflammation in COPD, is also activated by NE. However, neutrophil elastase level is positively correlated with the degree of airway inflammation and disease severity. Neutrophil elastase activates reactive oxygengenerating systems such as nicotinamide adenine dinucleotide phosphate (NADPH) oxidase and myeloperoxidase and it also generates mitochondrial-derived-reactive oxygen species formation by inducing the secretion of Interleukin (IL)-1 andTumour necrosis factor (TNF)- α. In addition, neutrophil elastase stimulates respiratory cell apoptosis by direct (e.g., activating the caspase-3 pathway) and indirect mechanisms (e.g., by secretion of Neutrophil Extracellular Traps). Surprisingly, neutrophil elastase may have small anti-inflammatory properties. In conclusion, neutrophil elastase is one of the main culprits responsible for COPD pathogenesis by mediating the activation of Triad COPD pathogenesis.

Modeling the degeneration of the collagen architecture in a microstructural model of the human cornea

We propose an enriched micromechanical model of the collagenous reinforcement of the eye stromal tissue. As a departure from an over-simplified model proposed a few years back, where collagen and chemical bonds were modeled as linear-elastic trusses, here we describe the chemical bonds by means of a more realistic generalized Lennard-Jones potential. In keeping with the original model, we disregard the multi-layer nature of the cornea and the continuum nature of the filling elastin matrix. The under-constrained locally orthogonal network of collagen fibrils is stabilized by crosslinks that provide the rigidity of the system and confer the ability to sustain the action of the intraocular pressure. In Ariza-Gracia et al., it has been shown that the weakening and the bulging of the cornea due to ectasia can be ascribed to the reduction of the density of the chemical bonds. The introduction of a pseudo-chemical potential supplies a more realistic model: any mechanical, enzymatic, or chemical cause of the degradation of the tissue observed in ectasia can be effectively introduced in a multi-physic potential, disregarding the adoption of phenomenological models. In numerical calculations, the high non-linearity of the model is suitably controlled by adopting a robust explicit solver based on dynamic relaxation.

Evidence for Continuity of Interstitial Spaces across Tissue and Organ Boundaries in Humans

Bodies have continuous reticular networks, comprising collagens, elastin, glycosaminoglycans, and other extracellular matrix components, within and around all organs. Fibrous tissue coverings of nerves and blood vessels create structural continuity beyond organ boundaries. We recently validated fluid flow through human fibrous tissues, though whether these interstitial spaces are continuous through the body or discontinuous, confined within individual organs, remains unlcear. We investigated continuity of interstitial spaces using two approaches. Non‐biological particles (tattoo pigment, colloidal silver, anthracotic pigment) were tracked within colon, skin, and lung interstitial spaces and into adjacent fascia. Hyaluronic acid, a macromolecular component of interstitial spaces, was also visualized, analyzing skin and subcutaneous fascia, colon and mesenteric fascia, lung and pleura, gynecologic organs and their fascial sheathes, breast, and heart and great vessels. Both techniques demonstrate interstitial space continuity within and between organs including within perineurium and vascular adventitia traversing organs and the spaces between them. We suggest that there is a body‐wide network of fluid‐filled interstitial spaces that has significant implications for molecular signaling, cell trafficking, and the spread of malignant and infectious disease.

In Vivo Study of Nasal Bone Reconstruction with Collagen, Elastin and Chitosan Membranes in Abstainer and Alcoholic Rats.

The aim of the present study was to evaluate the use of collagen, elastin, or chitosan biomaterial for bone reconstruction in rats submitted or not to experimental alcoholism. Wistar male rats were divided into eight groups, submitted to chronic alcohol ingestion (G5 to G8) or not (G1 to G4). Nasal bone defects were filled with clot in animals of G1 and G5 and with collagen, elastin, and chitosan grafts in G2/G6, G3/G7, and G4/G8, respectively. Six weeks after, all specimens underwent radiographic, tomographic, and microscopic evaluations. Bone mineral density was lower in the defect area in alcoholic animals compared to the abstainer animals. Bone neoformation was greater in the abstainer groups receiving the elastin membrane and in abstainer and alcoholic rats receiving the chitosan membrane (15.78 ± 1.19, 27.81 ± 0.91, 47.29 ± 0.97, 42.69 ± 1.52, 13.81 ± 1.60, 18.59 ± 1.37, 16.54 ± 0.89, and 37.06 ± 1.17 in G1 to G8, respectively). In conclusion, osteogenesis and bone density were more expressive after the application of the elastin matrix in abstainer animals and of the chitosan matrix in both abstainer and alcoholic animals. Chronic alcohol ingestion resulted in lower bone formation and greater formation of fibrous connective tissue.

The Lung Elastin Matrix Undergoes Rapid Degradation Upon Adult Loss of Hox5 Function.

Hox genes encode transcription factors that are critical for embryonic skeletal patterning and organogenesis. The Hoxa5, Hoxb5, and Hoxc5 paralogs are expressed in the lung mesenchyme and function redundantly during embryonic lung development. Conditional loss-of-function of these genes during postnatal stages leads to severe defects in alveologenesis, specifically in the generation of the elastin network, and animals display bronchopulmonary dysplasia (BPD) or BPD-like phenotype. Here we show the surprising results that mesenchyme-specific loss of Hox5 function at adult stages leads to rapid disruption of the mature elastin matrix, alveolar enlargement, and an emphysema-like phenotype. As the elastin matrix of the lung is considered highly stable, adult disruption of the matrix was not predicted. Just 2 weeks after deletion, adult Hox5 mutant animals show significant increases in alveolar space and changes in pulmonary function, including reduced elastance and increased compliance. Examination of the extracellular matrix (ECM) of adult Tbx4rtTA; TetOCre; Hox5afafbbcc lungs demonstrates a disruption of the elastin network although the underlying fibronectin, interstitial collagen and basement membrane appear unaffected. An influx of macrophages and increased matrix metalloproteinase 12 (MMP12) are observed in the distal lung 3 days after Hox5 deletion. In culture, fibroblasts from Hox5 mutant lungs exhibit reduced adhesion. These findings establish a novel role for Hox5 transcription factors as critical regulators of lung fibroblasts at adult homeostasis.

Anti-Aging β-Klotho Gene-Activated Scaffold Promotes Rejuvenative Wound Healing Response in Human Adipose-Derived Stem Cells.

Wound healing requires a tight orchestration of complex cellular events. Disruption in the cell-signaling events can severely impair healing. The application of biomaterial scaffolds has shown healing potential; however, the potential is insufficient for optimal wound maturation. This study explored the functional impact of a collagen-chondroitin sulfate scaffold functionalized with nanoparticles carrying an anti-aging gene β-Klotho on human adipose-derived stem cells (ADSCs) for rejuvenative healing applications. We studied the response in the ADSCs in three phases: (1) transcriptional activities of pluripotency factors (Oct-4, Nanog and Sox-2), proliferation marker (Ki-67), wound healing regulators (TGF-β3 and TGF-β1); (2) paracrine bioactivity of the secretome generated by the ADSCs; and (3) regeneration of basement membrane (fibronectin, laminin, and collagen IV proteins) and expression of scar-associated proteins (α-SMA and elastin proteins) towards maturation. Overall, we found that the β-Klotho gene-activated scaffold offers controlled activation of ADSCs’ regenerative abilities. On day 3, the ADSCs on the gene-activated scaffold showed enhanced (2.5-fold) activation of transcription factor Oct-4 that was regulated transiently. This response was accompanied by a 3.6-fold increase in the expression of the anti-fibrotic gene TGF-β3. Through paracrine signaling, the ADSCs-laden gene-activated scaffold also controlled human endothelial angiogenesis and pro-fibrotic response in dermal fibroblasts. Towards maturation, the ADSCs-laden gene-activated scaffold further showed an enhanced regeneration of the basement membrane through increases in laminin (2.1-fold) and collagen IV (8.8-fold) deposition. The ADSCs also expressed 2-fold lower amounts of the scar-associated α-SMA protein with improved qualitative elastin matrix deposition. Collectively, we determined that the β-Klotho gene-activated scaffold possesses tremendous potential for wound healing and could advance stem cell-based therapy for rejuvenative healing applications.

Abstract MP21: Transcriptomic Modulation Of Second Heart Field-derived Smooth Muscle Cells In Angiotensin Ii-infused Mice Promotes Aortopathy

Objective: Smooth muscle cells (SMCs) in the ascending aorta are derived from both the cardiac neural crest and second heart field (SHF). The importance of cardiac neural crest-derived SMCs on development of thoracic aortic aneurysms has been reported, while functional roles of SHF-derived SMCs are controversial. The aim of this study was to profile the transcriptome of SHF-derived SMCs in ascending aortas of angiotensin II (AngII)-infused mice by single cell RNA sequencing. Methods and Results: Female ROSA26 mT/mG mice were bred to male mice expressing Cre under the control of the Mef2c promoter (Mef2c Cre). Angiotensin II (AngII, 1,000 ng/kg/day) was infused subcutaneously into Mef2c Cre +/- mice and ascending aortas were harvested at day 3 of AngII infusion (n=5). Ascending aortas without AngII infusion were harvested as control (n=4). SHF-derived cells were sorted based on mGFP signal by FACS and single cell sequencing was performed using SHF-derived cells. Single cell RNA sequencing detected mRNA in multiple cell types, and AngII altered mRNA abundance of 3,658 genes in the SMC cluster. Several SMC contractile-related genes, such as Acta2, Tagln and Cnn1 , were increased by AngII compared to control SHF-derived SMCs. Conversely , Klf4 , a proliferative gene, was decreased in SHF-derived SMCs of AngII-infused mice. In addition, AngII increased multiple extracellular matrix component genes, including fibrillin1 and elastin. AngII also upregulated Serpine1 and Fn1 that are important for extracellular matrix organization. Thus, AngII led to transcriptomic modulations in SHF-derived SMCs. Of note, AngII infusion decreased mRNA abundance of Tgfbr2 and Lrp1 , key regulators of extracellular matrix maturation, in SHF-derived SMCs. To further investigate the importance of SHF-derived SMCs, we deleted either Tgfbr2 or Lrp1 in SHF-derived cells. SHF-specific Tgfrb2 deletion was embryonic lethal with peritoneal hemorrhage and outflow tract dilatation. SHF-specific Lrp1 deletion augmented AngII-induced thoracic aortic rupture and dilatations. Conclusion: SHF-derived SMCs play an important role in maintaining aortic integrity, and exhibit transcriptomic modulations by AngII infusion that may contribute to aortopathies.

Abstract P107: Celastrol Supplementation Profoundly Activates Aortic Mmp-9 And Abolishes Sexual Dimorphism Of Abdominal Aortic Aneurysm In Mice.

Background and Objective: Abdominal Aortic Aneurysms (AAAs) are permanent dilations of the abdominal aorta with greater than 80% mortality after rupture. AAA prevalence is 4-5 times greater in males than females. AAA formation involves a complex process of destruction of aortic media through degradation of extracellular matrix proteins, elastin and collagen. Angiotensin II (AngII) infusion model of AAA in mice recapitulates major features of human AAA mainly male gender specificity. Celastrol, a pentacyclic triterpene from the root extracts of Thunder God Vine ( Tripterygium wilfordii ), strongly suppressed AngII-induced cardiovascular complications in mice. The purpose of this study is to test the effect of Celastrol supplementation on AngII-induced AAAs in mice. Methods and Results: Male and female LDL receptor -/- mice (8 weeks old; n= 12 per group) were fed a fat-enriched diet (21% wt/wt fat; 0.15% wt/wt cholesterol) supplemented with or without Celastrol (10mg/kg/day) for 5 weeks. After 1 week of diet feeding, mice were infused with AngII (500 or 1000 ng/kg/min) for 28 days by osmotic minipumps. Dietary supplementation of celastrol significantly promoted AngII-induced abdominal aortic luminal dilation (Con = 1.25 ± 0.05 versus Celas = 1.55 ± 0.09 mm, P<0.05) and external aortic width (Con= 1.01 ± 0.09 versus Celas = 1.62 ± 0.14 mm, P<0.05) in male mice as measured by ultra-sonography and ex vivo measurement, with 90% incidence (10/11) compared to 36% (4/11) in control group. Interestingly, celastrol supplementation to AAA female mice, resulted in a significant increase in AngII-induced aortic luminal dilation (Con = 1.17 ± 0.30 versus Celas = 1.57 ± 0.21 mm, P<0.001) and AAA formation (Con= 0.94 ± 0.03 versus Celas = 1.36 ± 0.25 mm, P<0.05) mice with 80% incidence (12/15) compared to 6% (1/15) in control group. Celastrol supplementation dramatically increased AngII-induced aortic leukocyte accumulation, MMP-9 activity and medial elastin degradation in both male and female mice compared to saline and AngII controls. Conclusion: These findings demonstrate that celastrol supplementation to LDLr -/- mice ablates sexual dimorphism and promotes AngII-induced AAA formation, which is associated with increased leukocytic infiltration and MMP-9 activation.

Recombinant Interleukin-19 Suppresses the Formation and Progression of Experimental Abdominal Aortic Aneurysms.

BackgroundInterleukin‐19 is an immunosuppressive cytokine produced by immune and nonimmune cells, but its role in abdominal aortic aneurysm (AAA) pathogenesis is not known. This study aimed to investigate interleukin‐19 expression in, and influences on, the formation and progression of experimental AAAs.Methods and ResultsHuman specimens were obtained at aneurysm repair surgery or from transplant donors. Experimental AAAs were created in 10‐ to 12‐week‐old male mice via intra‐aortic elastase infusion. Influence and potential mechanisms of interleukin‐19 treatment on AAAs were assessed via ultrasonography, histopathology, flow cytometry, and gene expression profiling. Immunohistochemistry revealed augmented interleukin‐19 expression in both human and experimental AAAs. In mice, interleukin‐19 treatment before AAA initiation via elastase infusion suppressed aneurysm formation and progression, with attenuation of medial elastin degradation, smooth‐muscle depletion, leukocyte infiltration, neoangiogenesis, and matrix metalloproteinase 2 and 9 expression. Initiation of interleukin‐19 treatment after AAA creation limited further aneurysmal degeneration. In additional experiments, interleukin‐19 treatment inhibited murine macrophage recruitment following intraperitoneal thioglycolate injection. In classically or alternatively activated macrophages in vitro, interleukin‐19 downregulated mRNA expression of inducible nitric oxide synthase, chemokine C‐C motif ligand 2, and metalloproteinases 2 and 9 without apparent effect on cytokine‐expressing helper or cytotoxic T‐cell differentiation, nor regulatory T cellularity, in the aneurysmal aorta or spleen of interleukin‐19–treated mice. Interleukin‐19 also suppressed AAAs created via angiotensin II infusion in hyperlipidemic mice.ConclusionsBased on human evidence and experimental modeling observations, interleukin‐19 may influence the development and progression of AAAs.

In Vitro Antiaging Evaluation of Sunscreen Formulated from Nanostructured Lipid Carrier and Tocotrienol-Rich Fraction

Chronic exposure to ultraviolet (UV) radiation leads to photoaging. There is a tremendous rise in products having a dual activity of photoprotection and antiaging. In vitro analysis in dermal fibroblasts and their biological mechanisms involved are critical to determine antiaging potential. The study aimed to investigate the antiaging potential of sunscreen formulated from nanostructured lipid carrier and tocotrienol-rich fraction (NLC-TRF sunscreen). The antioxidant activity of the NLC-TRF sunscreen was evaluated by radical scavenging and hydrogen peroxide inhibition properties. Also, collagenase, elastase and matrix metalloproteinase-1 (MMP-1) inhibition activities, and type I collagen and elastin protein expression were studied. Quantitative real-time polymerase chain reaction (qPCR) was used to evaluate the mRNA expression of fibroblast growth factor (FGF), vascular endothelial growth factor (VEGF), transforming growth factor-β1 (TGF-β1), type I collagen (COL1A1), elastin (ELN), MMP-1, MMP-2, and tissue inhibitor matrix metalloproteinase-1 (TIMP-1). The results suggested that NLC-TRF sunscreen is effective in radical, anti-hydrogen peroxide, and collagenase, elastase and MMP-1 inhibition activities. Besides, a significant increase for type I collagen (3.47-fold) and elastin (2.16-fold) protein and fibroblast regeneration genes (FGF (2.12-fold), VEGF (1.91-fold), TGF-β1 (2.84-fold), TIMP-1 (1.42-fold), ELN (2.13-fold)) were observed after sample treatment. These findings support the therapeutic potential of NLC-TRF sunscreen in antiaging.