Pentagalloyl glucose-stabilized decellularized bovine jugular vein valved conduits as pulmonary conduit replacement.

Congenital heart diseases (CHD) are one of the most frequently diagnosed congenital disorders, affecting approximately 40,000 live births annually in the United States. Out of the new patients diagnosed with CHD yearly, an estimated 2,500 patients require a substitute, non-native conduit artery to replace structures congenitally absent or hypoplastic. Devices used for conduit replacement encounter limitations exhibiting varying degrees of stiffness, calcification, susceptibility to infection, thrombosis, and a lack of implant growth capacity. Here, we report the functionality of pentagalloyl glucose (PGG) stabilized decellularized valved bovine jugular vein conduit (PGG-DBJVC). The PGG-DBJVC tissues demonstrated mechanical properties comparable to native and glutaraldehyde fixed tissues, while exhibiting resistance to both collagenase and elastase enzymatic degradation. Subcutaneous implantation of tissues established their biocompatibility and resistance to calcification, while implantation in sheep in the pulmonary position demonstrated adequate implant functionality, and repopulation of host cells, without excessive inflammation. In conclusion, this PGG-DBJVC device could be a favorable replacement option for pediatric patients, reducing the need for reoperations required with current devices. STATEMENT OF SIGNIFICANCE: Congenital Heart Disease (CHD) is a common congenital disorder affecting many newborns in the United States each year. The use of substitute conduit arteries is necessary for some patients with CHD who have missing or underdeveloped structures. Current conduit replacement devices have limitations, including stiffness, susceptibility to infection and thrombosis, and lack of implant growth capacity. Pentagalloyl glucose-stabilized bovine jugular vein valved tissue (PGG-DBJVC) offers a promising solution as it is resistant to calcification, and biocompatible. When implanted in rats and as pulmonary conduit replacement in sheep, the PGG-DBJVC demonstrated cellular infiltration without excessive inflammation, which could lead to remodeling and integration with host tissue and eliminate the need for replacement as the child grows.

Constructing Lipoparticles Capable of Endothelial Cell-Derived Exosome-Mediated Delivery of Anti-miR-33a-5p to Cultured Macrophages.

Atherosclerosis is driven by intimal arterial macrophages accumulating cholesterol. Atherosclerosis also predominantly occurs in areas consisting of proinflammatory arterial endothelial cells. At time of writing, there are no available clinical treatments that precisely remove excess cholesterol from lipid-laden intimal arterial macrophages. Delivery of anti-miR-33a-5p to macrophages has been shown to increase apoAI-mediated cholesterol efflux via ABCA1 upregulation but delivering transgenes to intimal arterial macrophages is challenging due to endothelial cell barrier integrity. In this study, we aimed to test whether lipoparticles targeting proinflammatory endothelial cells can participate in endothelial cell-derived exosome exploitation to facilitate exosome-mediated transgene delivery to macrophages. We constructed lipoparticles that precisely target the proinflammatory endothelium and contain a plasmid that expresses XMOTIF-tagged anti-miR-33a-5p (LP-pXMoAntimiR33a5p), as XMOTIF-tagged small RNA demonstrates the capacity to be selectively shuttled into exosomes. The cultured cells used in our study were immortalized mouse aortic endothelial cells (iMAECs) and RAW 264.7 macrophages. From our results, we observed a significant decrease in miR-33a-5p expression in macrophages treated with exosomes released basolaterally by LPS-challenged iMAECs incubated with LP-pXMoAntimiR33a5p when compared to control macrophages. This decrease in miR-33a-5p expression in the treated macrophages caused ABCA1 upregulation as determined by a significant increase in ABCA1 protein expression in the treated macrophages when compared to the macrophage control group. The increase in ABCA1 protein also simulated ABCA1-dependent cholesterol efflux in treated macrophages-as we observed a significant increase in apoAI-mediated cholesterol efflux-when compared to the control group of macrophages. Based on these findings, strategies that involve combining proinflammatory-targeting lipoparticles and exploitation of endothelial cell-derived exosomes appear to be promising approaches for delivering atheroprotective transgenes to lipid-laden arterial intimal macrophages.

Elastin stabilization prevents impaired biomechanics in human pulmonary arteries and pulmonary hypertension in rats with left heart disease.

Pulmonary hypertension worsens outcome in left heart disease. Stiffening of the pulmonary artery may drive this pathology by increasing right ventricular dysfunction and lung vascular remodeling. Here we show increased stiffness of pulmonary arteries from patients with left heart disease that correlates with impaired pulmonary hemodynamics. Extracellular matrix remodeling in the pulmonary arterial wall, manifested by dysregulated genes implicated in elastin degradation, precedes the onset of pulmonary hypertension. The resulting degradation of elastic fibers is paralleled by an accumulation of fibrillar collagens. Pentagalloyl glucose preserves arterial elastic fibers from elastolysis, reduces inflammation and collagen accumulation, improves pulmonary artery biomechanics, and normalizes right ventricular and pulmonary hemodynamics in a rat model of pulmonary hypertension due to left heart disease. Thus, targeting extracellular matrix remodeling may present a therapeutic approach for pulmonary hypertension due to left heart disease.

Role of elastic fiber degradation in disease pathogenesis.

Elastin is a crucial extracellular matrix protein that provides structural integrity to tissues. Crosslinked elastin and associated microfibrils, named elastic fiber, contribute to biomechanics by providing the elasticity required for proper function. During aging and disease, elastic fiber can be progressively degraded and since there is little elastin synthesis in adults, degraded elastic fiber is not regenerated. There is substantial evidence linking loss or damage of elastic fibers to the clinical manifestation and pathogenesis of a variety of diseases. Disruption of elastic fiber networks by hereditary mutations, aging, or pathogenic stimuli results in systemic ailments associated with the production of elastin degradation products, inflammatory responses, and abnormal physiology. Due to its longevity, unique mechanical properties, and widespread distribution in the body, elastic fiber plays a central role in homeostasis of various physiological systems. While pathogenesis related to elastic fiber degradation has been more thoroughly studied in elastic fiber rich tissues such as the vasculature and the lungs, even tissues containing relatively small quantities of elastic fibers such as the eyes or joints may be severely impacted by elastin degradation. Elastic fiber degradation is a common observation in certain hereditary, age, and specific risk factor exposure induced diseases representing a converging point of pathological clinical phenotypes which may also help explain the appearance of co-morbidities. In this review, we will first cover the role of elastic fiber degradation in the manifestation of hereditary diseases then individually explore the structural role and degradation effects of elastic fibers in various tissues and organ systems. Overall, stabilizing elastic fiber structures and repairing lost elastin may be effective strategies to reverse the effects of these diseases.

Pentagalloyl Glucose (PGG) Prevents and Restores Mechanical Changes Caused by Elastic Fiber Fragmentation in the Mouse Ascending Aorta.

Thoracic aortic aneurysm (TAA) is characterized by dilation of the aorta that can lead to dissection or rupture. Degradation of elastic fibers is a consistent histopathological feature of TAA that likely contributes to disease progression. Pentagalloyl glucose (PGG) shows promise for stabilizing elastic fibers in abdominal aortic aneurysms, but its efficacy and mechanical effects in the thoracic aorta are unknown. We simulated TAAs using elastase (ELA) to degrade elastic fibers in the mouse ascending aorta and determined the preventative and restorative potential of PGG. Biaxial mechanical tests, constitutive model fitting, and multiphoton imaging were performed on untreated (UNT), PGG, ELA, PGG + ELA, and ELA + PGG treated aortas. PGG treatment alone does not significantly alter mechanical properties or wall structure compared to UNT. ELA treatment alone causes an increase in the unloaded diameter and length, decreased compliance, significant changes in the material constants, and separation of the outer layers of the aortic wall compared to UNT. PGG treatment before or after ELA ameliorates the mechanical and structural changes associated with elastic fiber degradation, with preventative PGG treatment being most effective. These results suggest that PGG is a potential pharmaceutical option to stabilize elastic fibers in TAA.

Reversal of elastase-induced abdominal aortic aneurysm following the delivery of nanoparticle-based pentagalloyl glucose (PGG) is associated with reduced inflammatory and immune markers.

OBJECTIVE: An Abdominal aortic aneurysm (AAA), a deadly disease in elderly population, is featured by expansion of aortic diameter, degradation and weakening of vasculature. Its common and significant characteristics are disarray and inflammation in vasculature. We tested the hypothesis that the reversal of abdominal aortic aneurysm by pentagalloyl glucose-loaded nanoparticles (PGG-NPs) therapy that targets degraded elastin suppresses inflammatory and immune markers to ameliorate the pathophysiology of the disease in advance stage aneurysm in a porcine pancreatic elastase (PPE)-induced mouse model of AAA. METHODS AND RESULTS: After induction of aneurysm in pathogen-free C57BL/6 male mice by applying PPE peri-adventitially to the abdominal aorta, once a week for two doses of intravenous injections of pentagalloyl glucose-loaded nanoparticles (PGG-NPs) conjugated with elastin targeted antibody were used to reverse the aneurysms. We showed that PGG-NPs therapy could suppress infiltration of macrophages, CD8 and CD4 subsets of T cells, matrix metalloproteinases (MMPs), inflammatory cytokines interferon (IFN-γ) and interleukin (IL)-6 at the local and systemic level. Moreover, such PGG-NPs therapy increases the induction of anti-inflammatory cytokines IL-13, IL-27 and IL-10 at the local and systemic level. The therapy also led to remodeling of elastic lamina at the aneurysm site. CONCLUSION: Nanoparticles-loaded pentagalloyl glucose therapy can be an effective treatment option against advanced stage aneurysms to reverse the disease by ameliorating inflammation and restoring arterial homeostasis.

Advancing peptide siRNA-carrier designs through L/D-amino acid stereochemical modifications to enhance gene silencing.

The 599 peptide has been previously shown to effectively deliver small interfering RNAs (siRNAs) to cancer cells, inducing targeted-oncogene silencing, with a consequent inhibition of tumor growth. Although effective, this study was undertaken to advance the 599 peptide siRNA-carrier design through L/D-amino acid stereochemical modifications. Consequently, 599 was modified to generate eight different peptide variants, incorporating either different stereochemical patterns of L/D-amino acids or a specific D-amino acid substitution. Upon analysis of the variants, it was observed that these modifications could, in some instances, increase/decrease the binding, nuclease/serum stability, and complex release of siRNAs, as well as influence the gene-silencing efficiencies of the complex. These modifications were also found to affect cellular uptake and intracellular localization patterns of siRNA cargo, with one particular variant capable of mediating binding of siRNAs to specific cellular projections, identified as filopodia. Interestingly, this variant also exhibited the most enhanced gene silencing in comparison to the parent 599 peptide, thus suggesting a possible connection between filopodia binding and enhanced gene silencing. Together, these data demonstrate the utility of peptide stereochemistry, as well as the importance of a key D-amino acid modification, in advancing the 599 carrier design for the enhancement of gene silencing in cancer cells.

Systemic delivery of targeted nanotherapeutic reverses angiotensin II-induced abdominal aortic aneurysms in mice.

Abdominal aortic aneurysm (AAA) disease causes dilation of the aorta, leading to aortic rupture and death if not treated early. It is the 14th leading cause of death in the U.S. and 10th leading cause of death in men over age 55, affecting thousands of patients. Despite the prevalence of AAA, no safe and efficient pharmacotherapies exist for patients. The deterioration of the elastic lamina in the aneurysmal wall is a consistent feature of AAAs, making it an ideal target for delivering drugs to the AAA site. In this research, we conjugated nanoparticles with an elastin antibody that only targets degraded elastin while sparing healthy elastin. After induction of aneurysm by 4-week infusion of angiotensin II (Ang II), two biweekly intravenous injections of pentagalloyl glucose (PGG)-loaded nanoparticles conjugated with elastin antibody delivered the drug to the aneurysm site. We show that targeted delivery of PGG could reverse the aortic dilation, ameliorate the inflammation, restore the elastic lamina, and improve the mechanical properties of the aorta at the AAA site. Therefore, simple iv therapy of PGG loaded nanoparticles can be an effective treatment option for early to middle stage aneurysms to reverse disease progression and return the aorta to normal homeostasis.

Polyphenol treatments increase elastin and collagen deposition by human dermal fibroblasts; Implications to improve skin health.

BACKGROUND: Skin aging is marked by progressive loss in elastin and collagen that causes wrinkling and sagging of skin. Tropoelastin (TE) is the precursor monomer of elastin secreted by cells that cross-links extracellularly to create functional elastic fibers. Cells maintain the capacity to make TE during the aging process. However, the process of extracellular tropoelastin cross-linking diminishes with age. Others have shown that TE production by cells increases with UV exposure. OBJECTIVE: We hypothesize that polyphenols may help coacervate cell secreted TE due to its elastin binding property and increase insoluble elastin in human dermal fibroblasts (HDFs). Increase in TE production by short term UV exposure may further improve elastin deposition by polyphenols. METHODS: We treated HDFs with polyphenols viz epigallocatechin gallate (EGCG) and pentagalloyl glucose (PGG) either with or without intermittent (UVA, 12 min three times a week) exposure for 3, 7, and 14 days. RESULTS: Polyphenols increased insoluble elastin deposition several folds as compared to control untreated cells. Furthermore, short UVA light exposure led to several-fold increased TE production in HDFs. Still, UVA exposure alone was unable to increase insoluble elastic fibers. When polyphenols were introduced with UVA exposure, insoluble elastin deposition was further enhanced in HDFs (30-45-fold increase). Polyphenol treatments with UVA exposure also led to increased collagen deposition in cell cultures. Polyphenols also prevented cell oxidation during UVA exposure. CONCLUSIONS: Polyphenols in combination with short exposure to UVA light increase extracellular matrix deposition of elastin and collagen and may improve skin properties.

Targeted Gold Nanoparticles as an Indicator of Mechanical Damage in an Elastase Model of Aortic Aneurysm.

Elastin is a key structural protein and its pathological degradation deterministic in aortic aneurysm (AA) outcomes. Unfortunately, using current diagnostic and clinical surveillance techniques the integrity of the elastic fiber network can only be assessed invasively. To address this, we employed fragmented elastin-targeting gold nanoparticles (EL-AuNPs) as a diagnostic tool for the evaluation of unruptured AAs. Electron dense EL-AuNPs were visualized within AAs using micro-computed tomography (micro-CT) and the corresponding Gold-to-Tissue volume ratios quantified. The Gold-to-Tissue volume ratios correlated strongly with the concentration (0, 0.5, or 10 U/mL) of infused porcine pancreatic elastase and therefore the degree of elastin damage. Hyperspectral mapping confirmed the spatial targeting of the EL-AuNPs to the sites of damaged elastin. Nonparametric Spearman's rank correlation indicated that the micro-CT-based Gold-to-Tissue volume ratios had a strong correlation with loaded (ρ = 0.867, p-val = 0.015) and unloaded (ρ = 0.830, p-val = 0.005) vessel diameter, percent dilation (ρ = 0.976, p-val = 0.015), circumferential stress (ρ = 0.673, p-val = 0.007), loaded (ρ = - 0.673, p-val = 0.017) and unloaded (ρ = - 0.697, p-val = 0.031) wall thicknesses, circumferential stretch (ρ = - 0.7234, p-val = 0.018), and lumen area compliance (ρ = - 0.831, p-val = 0.003). Likewise, in terms of axial force and axial stress vs. stretch, the post-elastase vessels were stiffer. Collectively, these findings suggest that, when combined with CT imaging, EL-AuNPs can be used as a powerful tool in the non-destructive estimation of mechanical and geometric features of AAs.

Nanoparticle-based targeted delivery of pentagalloyl glucose reverses elastase-induced abdominal aortic aneurysm and restores aorta to the healthy state in mice.

AIM: Abdominal aortic aneurysms (AAA) is a life-threatening weakening and expansion of the abdominal aorta due to inflammatory cell infiltration and gradual degeneration of extracellular matrix (ECM). There are no pharmacological therapies to treat AAA. We tested the hypothesis that nanoparticle (NP) therapy that targets degraded elastin and delivers anti-inflammatory, anti-oxidative, and ECM stabilizing agent, pentagalloyl glucose (PGG) will reverse advance stage aneurysm in an elastase-induced mouse model of AAA. METHOD AND RESULTS: Porcine pancreatic elastase (PPE) was applied periadventitially to the infrarenal aorta in mice and AAA was allowed to develop for 14 days. Nanoparticles loaded with PGG (EL-PGG-NPs) were then delivered via IV route at 14-day and 21-day (10 mg/kg of body weight). A control group of mice received no therapy. The targeting of NPs to the AAA site was confirmed with fluorescent dye marked NPs and gold NPs. Animals were sacrificed at 28-d. We found that targeted PGG therapy reversed the AAA by decreasing matrix metalloproteinases MMP-9 and MMP-2, and the infiltration of macrophages in the medial layer. The increase in diameter of the aorta was reversed to healthy controls. Moreover, PGG treatment restored degraded elastic lamina and increased the circumferential strain of aneurysmal aorta to the healthy levels. CONCLUSION: Our results support that site-specific delivery of PGG with targeted nanoparticles can be used to treat already developed AAA. Such therapy can reverse inflammatory markers and restore arterial homeostasis.

Gold nanoparticles that target degraded elastin improve imaging and rupture prediction in an AngII mediated mouse model of abdominal aortic aneurysm.

Background: Abdominal aortic aneurysms (AAA) are characterized by a progressive disruption and weakening of the extracellular matrix (ECM) leading to dilation of the aorta which can be fatal if not treated. Current diagnostic imaging modalities provides little insight on the varying degree of ECM degeneration that precedes rupture in AAAs. Targeted delivery of contrast agents such as gold nanoparticles (GNPs) that bind to degraded matrix could prove useful when combined with computed tomography (CT) to provide a non-invasive surrogate marker of AAA rupture potential. Methods: AAAs were induced by chronic infusion of angiotensin II (AngII) into low density-lipoprotein receptor-deficient (LDLr -/-) mice in combination with a high-fat diet. Abdominal ultrasound was used to monitor disease progression and to assess the circumferential strain throughout the cardiac cycle. At six weeks, GNPs conjugated with an elastin antibody (EL-GNP) were injected retro-orbitally. Mice were euthanized 24 hours after EL-GNP injection, and aortas were explanted and scanned ex-vivo with a micro-CT system. Histological assessment and 3D models of the aneurysms with micro-CT were used to determine the EL-GNPs distribution. Isolated vessel burst pressure testing was performed on each aneurysmal aorta to quantify rupture strength and to assess rupture location. Results: Aneurysms were found along the suprarenal aorta in AngII infused mice. Darkfield microscopy indicated EL-GNPs accumulation around the site of degraded elastin while avoiding the healthy and intact elastin fibers. Using nonlinear regression, the micro-CT signal intensity of EL-GNPs along the suprarenal aortas correlated strongly with burst pressures (R2=0.9415) but not the dilation as assessed by ultrasound measurements. Conclusions: Using an established mouse model of AAA, we successfully demonstrated in vivo targeting of EL-GNPs to damaged aortic elastin and correlated micro-CT-based signal intensities with burst pressures. Thus, we show that this novel targeting technique can be used as a diagnostic tool to predict the degree of elastin damage and therefore rupture potential in AAAs better than the extent of dilation.

Site-specific chelation therapy with EDTA-loaded albumin nanoparticles reverses arterial calcification in a rat model of chronic kidney disease.

Medial arterial calcification (MAC) is a common outcome in diabetes and chronic kidney disease (CKD). It occurs as linear mineral deposits along the degraded elastin lamellae and is responsible for increased aortic stiffness and subsequent cardiovascular events. Current treatments for calcification, particularly in CKD, are predominantly focused on regulating the mineral disturbance and other risk factors. Ethylene diamine tetraacetic acid (EDTA), a chelating agent, can resorb mineral deposits, but the systemic delivery of EDTA may cause side effects such as hypocalcemia and bone resorption. We have developed elastin antibody conjugated albumin nanoparticles that target only degraded elastin in vasculature while sparing healthy tissues. In this study, we tested a targeted nanoparticle-based EDTA chelation therapy to reverse CKD-associated MAC. Renal failure was induced in Sprague-Dawley rats by a high adenine diet supplemented by high P and Ca for 28 days that led to MAC. Intravenous delivery of DiR dye-loaded nanoparticles confirmed targeting to vascular degraded elastin and calcification sites within 24 hours. Next, EDTA-loaded albumin nanoparticles conjugated with an anti-elastin antibody were intravenously injected twice a week for two weeks. The targeted nanoparticles delivered EDTA at the site of vascular calcification and reversed mineral deposits without any untoward effects. Systemic EDTA injections or blank nanoparticles were ineffective in reversing MAC. Reversal of calcification seems to be stable as it did not return after the treatment was stopped for an additional four weeks. Targeted EDTA chelation therapy successfully reversed calcification in this adenine rat model of CKD. We consider that targeted NP therapy will provide an attractive option to reverse calcification and has a high potential for clinical translation.

Pentagalloyl glucose increases elastin deposition, decreases reactive oxygen species and matrix metalloproteinase activity in pulmonary fibroblasts under inflammatory conditions.

Emphysema is characterized by degradation of lung alveoli that leads to poor airflow in lungs. Irreversible elastic fiber degradation by matrix metalloproteinases (MMPs) and reactive oxygen species (ROS) activity leads to loss of elasticity and drives the progression of this disease. We investigated if a polyphenol, pentagalloyl glucose (PGG) can increase elastin production in pulmonary fibroblasts. We also studied the effect of PGG treatment in reducing MMP activity and ROS levels in cells. We exposed rat pulmonary fibroblasts to two different types of inflammatory environments i.e., tumor necrosis factor-α (TNF-α) and cigarette smoke extract (CSE) to mimic the disease. Parameters like lysyl oxidase (LOX) and elastin gene expression, MMP-9 activity in the medium, lysyl oxidase (LOX) activity and ROS levels were studied to assess the effect of PGG on pulmonary fibroblasts. CSE inhibited lysyl oxidase (LOX) enzyme activity that resulted in a decreased elastin formation. Similarly, TNF-α treated cells showed less elastin in the cell layers. Both these agents caused increase in MMP activity and ROS levels in cells. However, when supplemented with PGG treatment along with these two inflammatory agents, we saw a significant increase in elastin deposition, reduction in both MMP activity and ROS levels. Thus PGG, which has anti-inflammatory, anti-oxidant properties coupled with its ability to aid in elastic fiber formation, can be a multifunctional drug to potentially arrest the progression of emphysema.

Systemic Delivery of Nanoparticles Loaded with Pentagalloyl Glucose Protects Elastic Lamina and Prevents Abdominal Aortic Aneurysm in Rats.

Degeneration of elastin plays a vital role in the pathology and progression of abdominal aortic aneurysm (AAA). Our previous study showed that pentagalloyl glucose (PGG), a core derivative of tannic acid, hinders the development of AAAs in a clinically relevant animal model when applied locally. In this study, we tested whether targeted nanoparticles (NPs) can deliver PGG to the site of an aneurysm and prevent aneurysmal growth by protecting elastin. PGG-loaded albumin NPs with a surface-conjugated elastin-specific antibody were prepared. Aneurysms were induced by calcium chloride-mediated injury to the abdominal aorta in rats. NPs were injected into the tail vein after 10 days of CaCl2 injury. Rats were euthanized after 38 days. PGG delivery led to reduction in macrophage recruitment, matrix metalloproteinase (MMP) activity, elastin degradation, calcification, and development of aortic aneurysm. Such NP delivery offers the potential for the development of effective and safe therapies for AAA.

Prevention of abdominal aortic aneurysm progression by targeted inhibition of matrix metalloproteinase activity with batimastat-loaded nanoparticles.

RATIONALE: Matrix metalloproteinases (MMPs)-mediated extracellular matrix destruction is the major cause of development and progression of abdominal aortic aneurysms. Systemic treatments of MMP inhibitors have shown effectiveness in animal models, but it did not translate to clinical success either because of low doses used or systemic side effects of MMP inhibitors. We propose a targeted nanoparticle (NP)-based delivery of MMP inhibitor at low doses to the abdominal aortic aneurysms site. Such therapy will be an attractive option for preventing expansion of aneurysms in patients without systemic side effects. OBJECTIVE: Our previous study showed that poly(d,l-lactide) NPs conjugated with an antielastin antibody could be targeted to the site of an aneurysm in a rat model of abdominal aortic aneurysms. In the study reported here, we tested whether such targeted NPs could deliver the MMP inhibitor batimastat (BB-94) to the site of an aneurysm and prevent aneurysmal growth. METHODS AND RESULTS: Poly(d,l-lactide) NPs were loaded with BB-94 and conjugated with an elastin antibody. Intravenous injections of elastin antibody-conjugated BB-94-loaded NPs targeted the site of aneurysms and delivered BB-94 in a calcium chloride injury-induced abdominal aortic aneurysms in rats. Such targeted delivery inhibited MMP activity, elastin degradation, calcification, and aneurysmal development in the aorta (269% expansion in control versus 40% elastin antibody-conjugated BB-94-loaded NPs) at a low dose of BB-94. The systemic administration of BB-94 alone at the same dose was ineffective in producing MMP inhibition. CONCLUSIONS: Targeted delivery of MMP inhibitors using NPs may be an attractive strategy to inhibit aneurysmal progression.

A novel crosslinking method for improved tear resistance and biocompatibility of tissue based biomaterials.

Over 300,000 heart valve replacements are performed annually to replace stenotic and regurgitant heart valves. Bioprosthetic heart valves (BHVs), derived from glutaraldehyde crosslinked (GLUT) porcine aortic valve leaflets or bovine pericardium are often used. However, valve failure can occur within 12-15 years due to calcification and/or progressive degeneration. In this study, we have developed a novel fabrication method that utilizes carbodiimide, neomycin trisulfate, and pentagalloyl glucose crosslinking chemistry (TRI) to better stabilize the extracellular matrix of porcine aortic valve leaflets. We demonstrate that TRI treated leaflets show similar biomechanics to GLUT crosslinked leaflets. TRI treated leaflets had better resistance to enzymatic degradation in vitro and demonstrated better tearing toughness after challenged with enzymatic degradation. When implanted subcutaneously in rats for up to 90 days, GLUT control leaflets calcified heavily while TRI treated leaflets resisted calcification, retained more ECM components, and showed better biocompatibility.

Rat aortic smooth muscle cells cultured on hydroxyapatite differentiate into osteoblast-like cells via BMP-2-SMAD-5 pathway.

Vascular calcification is an important pathological condition associated with increased risk of cardiovascular mortality. Hydroxyapatite (HA) found in such deposits is the same polymorph of calcium (Ca) found in bone, indicating calcification may involve mechanisms akin to bone formation. Vascular smooth muscle cells (Vsmcs) have been shown to undergo phenotypic change to osteoblast-like cells. However, the mechanisms underlying this phenotypic change are unclear, and whether the stimulus to become osteogenic is a result of loss of mineralization inhibitors or early mineral deposits is not known. Our aim in this study is to identify mechanisms and signal transduction pathways that cause differentiation of Vsmcs into osteoblast-like cells in the presence of HA. We first characterized vascular origin of Vsmcs by studying the expression of smooth muscle cell markers: myosin heavy chain and smooth muscle actin along with SM22α at both mRNA and protein levels. Vsmcs grown on HA exhibited progressive change in cellular morphology at 3-, 7-, and 14-day time points. Culturing of Vsmcs on HA disc resulted in decrease in media Ca levels and increased expression of Ca-sensing receptor (CaSR) on Vsmcs resulting in upregulation of intracellular CaSR signaling leading to increased BMP-2 secretion. BMP-2 pathway mediated differentiation of Vsmcs to osteoblast-like cells shown by expression of osteogenic markers like runt-related transcription factor 2, osteocalcin, and alkaline phosphatase at mRNA and protein levels. Blocking CaSR by NPS-2143 reduced BMP-2 secretion and blocking the BMP-2 pathway by LDN-193189, a BMP inhibitor, modulated expression of osteogenic markers confirming their role in osteogenesis of Vsmcs.