RESUMO
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.
RESUMO
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.
Assuntos
Anti-Inflamatórios não Esteroides/farmacologia , Antioxidantes/farmacologia , Elastina/genética , Fibroblastos/efeitos dos fármacos , Taninos Hidrolisáveis/farmacologia , Metaloproteinase 9 da Matriz/genética , Poluição por Fumaça de Tabaco/análise , Animais , Misturas Complexas/antagonistas & inibidores , Misturas Complexas/farmacologia , Elastina/agonistas , Elastina/antagonistas & inibidores , Elastina/biossíntese , Fibroblastos/metabolismo , Fibroblastos/patologia , Regulação da Expressão Gênica , Inflamação , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Pulmão/patologia , Metaloproteinase 9 da Matriz/metabolismo , Cultura Primária de Células , Proteína-Lisina 6-Oxidase/antagonistas & inibidores , Proteína-Lisina 6-Oxidase/genética , Proteína-Lisina 6-Oxidase/metabolismo , Ratos , Espécies Reativas de Oxigênio/antagonistas & inibidores , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
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.
Assuntos
Aorta Abdominal/efeitos dos fármacos , Aneurisma da Aorta Abdominal/tratamento farmacológico , Portadores de Fármacos , Imunoconjugados/administração & dosagem , Inibidores de Metaloproteinases de Matriz/administração & dosagem , Metaloproteinases da Matriz/metabolismo , Nanopartículas , Fenilalanina/análogos & derivados , Poliésteres/química , Tiofenos/administração & dosagem , Animais , Aorta Abdominal/enzimologia , Aorta Abdominal/imunologia , Aorta Abdominal/patologia , Aneurisma da Aorta Abdominal/induzido quimicamente , Aneurisma da Aorta Abdominal/enzimologia , Aneurisma da Aorta Abdominal/imunologia , Aneurisma da Aorta Abdominal/patologia , Cloreto de Cálcio , Química Farmacêutica , Modelos Animais de Doenças , Progressão da Doença , Elastina/imunologia , Elastina/metabolismo , Imunoconjugados/química , Imunoconjugados/metabolismo , Macrófagos/efeitos dos fármacos , Macrófagos/enzimologia , Masculino , Inibidores de Metaloproteinases de Matriz/química , Inibidores de Metaloproteinases de Matriz/metabolismo , Camundongos , Fenilalanina/administração & dosagem , Fenilalanina/química , Fenilalanina/metabolismo , Proteólise , Células RAW 264.7 , Ratos Sprague-Dawley , Tiofenos/química , Tiofenos/metabolismo , Fatores de Tempo , Calcificação Vascular/enzimologia , Calcificação Vascular/patologia , Calcificação Vascular/prevenção & controleRESUMO
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.
Assuntos
Aorta/citologia , Proteína Morfogenética Óssea 2/metabolismo , Durapatita/química , Miócitos de Músculo Liso/efeitos dos fármacos , Proteína Smad5/metabolismo , Animais , Técnicas de Cultura de Células , Diferenciação Celular/efeitos dos fármacos , Regulação da Expressão Gênica , Naftalenos/química , Osteoblastos/citologia , Osteocalcina/metabolismo , Osteogênese , Reação em Cadeia da Polimerase , Pós , Pirazóis/química , Pirimidinas/química , RNA Mensageiro/metabolismo , Ratos , Receptores de Detecção de Cálcio/metabolismo , Transdução de Sinais , StentsRESUMO
Vascular calcification can be categorized into two different types. Intimal calcification related to atherosclerosis and elastin-specific medial arterial calcification (MAC). Osteoblast-like differentiation of vascular smooth muscle cells (VSMCs) has been shown in both types; however, how this relates to initiation of vascular calcification is unclear. We hypothesize that the initial deposition of hydroxyapatite-like mineral in MAC occurs on degraded elastin first and that causes osteogenic transformation of VSMCs. To test this, rat aortic smooth muscle cells (RASMCs) were cultured on hydroxyapatite crystals and calcified aortic elastin. Using RT-PCR and specific protein assays, we demonstrate that RASMCs lose their smooth muscle lineage markers like alpha smooth muscle actin (SMA) and myosin heavy chain (MHC) and undergo chondrogenic/osteogenic transformation. This is indicated by an increase in the expression of typical chondrogenic proteins such as aggrecan, collagen type II alpha 1(Col2a1) and bone proteins such as runt-related transcription factor 2 (RUNX2), alkaline phosphatase (ALP) and osteocalcin (OCN). Furthermore, when calcified conditions are removed, cells return to their original phenotype. Our data supports the hypothesis that elastin degradation and calcification precedes VSMCs' osteoblast-like differentiation.
Assuntos
Durapatita/metabolismo , Elastina/metabolismo , Túnica Íntima/metabolismo , Túnica Média/metabolismo , Calcificação Vascular/metabolismo , Actinas , Agrecanas/biossíntese , Fosfatase Alcalina/biossíntese , Animais , Aorta/metabolismo , Aorta/patologia , Técnicas de Cultura de Células , Diferenciação Celular , Células Cultivadas , Condrogênese/fisiologia , Colágeno Tipo II/biossíntese , Subunidade alfa 1 de Fator de Ligação ao Core/biossíntese , Cadeias Pesadas de Miosina , Osteoblastos/metabolismo , Osteocalcina/biossíntese , Osteogênese/fisiologia , RatosRESUMO
Abdominal aortic aneurysms (AAA) are progressive dilatations of infra-renal aorta causing structural weakening rendering the aorta prone to rupture. AAA can be potentially stabilized by inhibiting inflammatory enzymes such as matrix metalloproteinases (MMP); however, active regression of AAA is not possible without new elastic fiber regeneration. Here we report the elastogenic benefit of direct delivery of polyphenols such as pentagalloyl glucose (PGG), epigallocatechin gallate (EGCG), and catechin, to smooth muscle cells obtained either from healthy or from aneurysmal rat aorta. Addition of 10 µg/ml PGG and ECGC induce elastin synthesis, organization, and crosslinking while catechin does not. Our results indicate that polyphenols bind to monomeric tropoelastin and enhance coacervation, aid in crosslinking of elastin by increasing lysyl oxidase (LOX) synthesis, and by blocking MMP-2 activity. Thus, polyphenol treatments leads to increased mature elastin fibers synthesis without increasing the production of intracellular tropoelastin.
Assuntos
Tecido Elástico/metabolismo , Miócitos de Músculo Liso/efeitos dos fármacos , Polifenóis/farmacologia , Tropoelastina/metabolismo , Animais , Aorta Abdominal/citologia , Aorta Abdominal/metabolismo , Aorta Abdominal/fisiologia , Aneurisma da Aorta Abdominal/metabolismo , Aneurisma da Aorta Abdominal/patologia , Aneurisma da Aorta Abdominal/fisiopatologia , Catequina/análogos & derivados , Catequina/metabolismo , Catequina/farmacologia , Células Cultivadas , Tecido Elástico/ultraestrutura , Elastina/metabolismo , Elastina/ultraestrutura , Fibrilinas , Taninos Hidrolisáveis/metabolismo , Taninos Hidrolisáveis/farmacologia , Cinética , Masculino , Metaloproteinase 2 da Matriz/metabolismo , Proteínas dos Microfilamentos/metabolismo , Microscopia Eletrônica de Transmissão , Microscopia de Fluorescência , Músculo Liso Vascular/citologia , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/ultraestrutura , Polifenóis/metabolismo , Proteína-Lisina 6-Oxidase/metabolismo , Ratos , Ratos Sprague-Dawley , RegeneraçãoRESUMO
Significant challenges remain in targeting drugs to diseased vasculature; most important being rapid blood flow with high shear, limited availability of stable targets, and heterogeneity and recycling of cellular markers. We developed nanoparticles (NPs) to target degraded elastic lamina, a consistent pathological feature in vascular diseases. In-vitro organ and cell culture experiments demonstrated that these NPs were not taken up by cells, but instead retained within the extracellular space; NP binding was proportional to the extent of elastic lamina damage. With three well-established rodent models of vascular diseases such as aortic aneurysm (calcium chloride mediated aortic injury in rats), atherosclerosis (fat-fed apoE-/- mice), and vascular calcification (warfarin + vitamin K injections in rats), we show precise NPs spatial targeting to degraded vascular elastic lamina while sparing healthy vasculature when NPs were delivered systemically. Nanoparticle targeting degraded elastic lamina is attractive to deliver therapeutic or imaging agents to the diseased vasculature. FROM THE CLINICAL EDITOR: This novel work focuses on nanoparticle targeting of degraded elastic lamina in a variety of diseases, including atherosclerosis, vascular calcification, and aneurysm formation, and demonstrates the feasibility to deliver therapeutic or imaging agents to the diseased vasculature.
Assuntos
Diagnóstico por Imagem/métodos , Sistemas de Liberação de Medicamentos/métodos , Ácido Láctico/química , Nanopartículas/química , Polímeros , Animais , Aneurisma Aórtico/diagnóstico , Aterosclerose/diagnóstico , Masculino , Camundongos , Polímeros/química , Ratos , Ratos Sprague-Dawley , Calcificação Vascular/diagnósticoRESUMO
Background: Pentagalloyl glucose (PGG) is a polyphenol with vasoprotective properties. Targeted delivery of PGG reversed aortic aneurysm growth in several rodent models associated with decreased number of macrophages and transforming growth factor-ß (TGF-ß) expression. Thus, we sought to determine cellular mechanisms by which PGG reduces macrophage-induced aortic pathogenicity and its relationship to TGF-ß. Methods: Using THP-1 cells, primary human aortic cells, and explanted rat aortas, we assessed the anti-inflammatory effect of PGG. Expression of pro/anti-inflammatory macrophage markers was analyzed. Adhesion of monocytes as well as oxidative stress status, viability, and TGF-ß expression after primary aortic cell exposure to macrophage-conditioned medium with and without PGG were assessed. The release of TGF-ß was also examined in elastase-treated cultured rat aortas. Results: PGG pre-treatment of human aortic cell monolayers reduced the adhesion of THP-1 monocytes. PGG enhanced the expression of anti-inflammatory markers in THP-1-derived macrophages, and increased mitochondrial reactive oxygen species as well as mitochondrial polarization. Conditioned medium from THP-1-derived macrophages induced reactive oxygen species, cell death, and TGF-ß release from human aortic cells, which was suppressed by PGG. In explanted rat aortas, PGG reduced elastase mediated TGF-ß release. Conclusions: Combining anti-inflammatory, cytotoxic, and oxidative effects, PGG has high cardiovascular therapeutic potential. We confirmed previous in vivo observations whereby PGG suppressed TGF-ß response associated with disease resolution.
Assuntos
Anti-Inflamatórios , Aorta , Taninos Hidrolisáveis , Macrófagos , Fator de Crescimento Transformador beta , Taninos Hidrolisáveis/farmacologia , Humanos , Animais , Fator de Crescimento Transformador beta/metabolismo , Células THP-1 , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Aorta/efeitos dos fármacos , Aorta/metabolismo , Aorta/patologia , Anti-Inflamatórios/farmacologia , Ratos , Espécies Reativas de Oxigênio/metabolismo , Masculino , Adesão Celular/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacosRESUMO
Elastin-specific medial vascular calcification, termed "Monckeberg's sclerosis," has been recognized as a major risk factor for various cardiovascular events. We hypothesize that chelating agents, such as disodium ethylenediaminetetraacetic acid (EDTA), diethylenetriaminepentaacetic acid (DTPA), and sodium thiosulfate (STS) might reverse elastin calcification by directly removing calcium from calcified tissues into soluble calcium complexes. We assessed the chelating ability of EDTA, DTPA, and STS on removal of calcium from hydroxyapatite (HA) powder, calcified porcine aortic elastin, and calcified human aorta in vitro. We show that both EDTA and DTPA could effectively remove calcium from HA and calcified tissues, while STS was not effective. The tissue architecture was not altered during chelation. In the animal model of aortic elastin-specific calcification, we further show that local periadventitial delivery of EDTA loaded in to poly(lactic-co-glycolic acid) nanoparticles regressed elastin-specific calcification in the aorta. Collectively, the data indicate that elastin-specific medial vascular calcification could be reversed by chelating agents.
Assuntos
Doenças da Aorta/tratamento farmacológico , Quelantes/uso terapêutico , Calcificação Vascular/tratamento farmacológico , Idoso , Animais , Doenças da Aorta/patologia , Cálcio/metabolismo , Quelantes/farmacologia , Avaliação Pré-Clínica de Medicamentos , Ácido Edético/farmacologia , Ácido Edético/uso terapêutico , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Ácido Pentético/farmacologia , Ácido Pentético/uso terapêutico , Ratos , Ratos Sprague-Dawley , Suínos , Resultado do Tratamento , Calcificação Vascular/patologiaRESUMO
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.
Assuntos
Bioprótese , Cardiopatias Congênitas , Próteses Valvulares Cardíacas , Trombose , Criança , Humanos , Recém-Nascido , Bovinos , Animais , Ratos , Ovinos , Veias Jugulares , Resultado do Tratamento , Ventrículos do Coração , Estudos Retrospectivos , Cardiopatias Congênitas/cirurgia , Inflamação , Glucose/farmacologiaRESUMO
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.
Assuntos
Tecido Elástico , Elastina , Elastina/genética , Tecido Elástico/metabolismo , Matriz Extracelular/metabolismo , Proteínas da Matriz Extracelular/metabolismo , Pulmão/metabolismoRESUMO
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.
Assuntos
Aneurisma da Aorta Torácica , Tecido Elástico , Camundongos , Animais , Tecido Elástico/metabolismo , Aorta/patologia , Elastase Pancreática , Preparações Farmacêuticas/metabolismo , Glucose/metabolismoRESUMO
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.
Assuntos
Cardiopatias , Hipertensão Pulmonar , Humanos , Animais , Ratos , Artéria Pulmonar , Fenômenos Biomecânicos , ElastinaRESUMO
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.
Assuntos
Anti-Inflamatórios/administração & dosagem , Aneurisma da Aorta Abdominal/tratamento farmacológico , Taninos Hidrolisáveis/administração & dosagem , Sistemas de Liberação de Fármacos por Nanopartículas/química , Animais , Aorta Abdominal/efeitos dos fármacos , Aorta Abdominal/imunologia , Aorta Abdominal/patologia , Aneurisma da Aorta Abdominal/induzido quimicamente , Aneurisma da Aorta Abdominal/imunologia , Aneurisma da Aorta Abdominal/patologia , Modelos Animais de Doenças , Humanos , Inflamação/induzido quimicamente , Inflamação/tratamento farmacológico , Inflamação/imunologia , Inflamação/patologia , Masculino , Camundongos , Elastase Pancreática/efeitos adversosRESUMO
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.
Assuntos
Fibroblastos/efeitos dos fármacos , Polifenóis/administração & dosagem , Envelhecimento da Pele/efeitos dos fármacos , Higiene da Pele/métodos , Células Cultivadas , Colágeno/metabolismo , Tecido Elástico/química , Elastina/metabolismo , Matriz Extracelular/efeitos dos fármacos , Matriz Extracelular/metabolismo , Fibroblastos/química , Fibroblastos/metabolismo , Humanos , Oxirredução/efeitos dos fármacos , Oxirredução/efeitos da radiação , Cultura Primária de Células , Pele/citologia , Pele/efeitos dos fármacos , Pele/metabolismo , Pele/efeitos da radiação , Envelhecimento da Pele/efeitos da radiação , Raios Ultravioleta/efeitos adversosRESUMO
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.
Assuntos
Angiotensina II/farmacologia , Aneurisma da Aorta Abdominal/tratamento farmacológico , Sistemas de Liberação de Medicamentos/métodos , Taninos Hidrolisáveis/uso terapêutico , Nanopartículas/uso terapêutico , Animais , Anticorpos/imunologia , Aneurisma da Aorta Abdominal/induzido quimicamente , Elastina/imunologia , Taninos Hidrolisáveis/administração & dosagem , Injeções Intravenosas , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Nanopartículas/administração & dosagem , Soroalbumina BovinaRESUMO
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.
RESUMO
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.
Assuntos
Aorta Abdominal/efeitos dos fármacos , Aneurisma da Aorta Abdominal/tratamento farmacológico , Portadores de Fármacos/química , Taninos Hidrolisáveis/administração & dosagem , Imunoconjugados/administração & dosagem , Animais , Anticorpos/administração & dosagem , Anticorpos/imunologia , Aorta Abdominal/diagnóstico por imagem , Aorta Abdominal/patologia , Aneurisma da Aorta Abdominal/induzido quimicamente , Aneurisma da Aorta Abdominal/diagnóstico por imagem , Modelos Animais de Doenças , Elastina/antagonistas & inibidores , Elastina/imunologia , Matriz Extracelular/efeitos dos fármacos , Matriz Extracelular/patologia , Ouro , Humanos , Imunoconjugados/imunologia , Injeções Intravenosas , Masculino , Nanopartículas Metálicas/química , Camundongos , Elastase Pancreática/administração & dosagem , Elastase Pancreática/toxicidade , Soroalbumina Bovina/química , UltrassonografiaRESUMO
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.