RESUMO
Nonalcoholic fatty liver disease (NAFLD) is the leading cause of chronic liver disease worldwide. Liver biopsy remains the gold standard for diagnosis and staging of disease. There is a clinical need for noninvasive diagnostic tools for risk stratification, follow-up, and monitoring treatment response that are currently lacking, as well as preclinical models that recapitulate the etiology of the human condition. We have characterized the progression of NAFLD in eNOS-/- mice fed a high fat diet (HFD) using noninvasive Dixon-based magnetic resonance imaging and single voxel STEAM spectroscopy-based protocols to measure liver fat fraction at 3 T. After 8 weeks of diet intervention, eNOS-/- mice exhibited significant accumulation of intra-abdominal and liver fat compared with control mice. Liver fat fraction measured by 1 H-MRS in vivo showed a good correlation with the NAFLD activity score measured by histology. Treatment of HFD-fed NOS3-/- mice with metformin showed significantly reduced liver fat fraction and altered hepatic lipidomic profile compared with untreated mice. Our results show the potential of in vivo liver MRI and 1 H-MRS to noninvasively diagnose and stage the progression of NAFLD and to monitor treatment response in an eNOS-/- murine model that represents the classic NAFLD phenotype associated with metabolic syndrome.
Assuntos
Metformina , Hepatopatia Gordurosa não Alcoólica , Humanos , Animais , Camundongos , Hepatopatia Gordurosa não Alcoólica/diagnóstico por imagem , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Hepatopatia Gordurosa não Alcoólica/metabolismo , Ácidos Graxos/metabolismo , Metformina/farmacologia , Metformina/uso terapêutico , Modelos Animais de Doenças , Fígado/metabolismo , Espectroscopia de Ressonância Magnética/métodos , Camundongos Endogâmicos C57BLRESUMO
PURPOSE: To develop a novel simultaneous co-registered T1 , T2 , T2∗ , T1ρ , and fat fraction abdominal MR fingerprinting (MRF) approach for fully comprehensive liver-tissue characterization in a single breath-hold scan. METHODS: A gradient-echo liver MRF sequence with low fixed flip angle, multi-echo radial readout, and varying magnetization preparation pulses for multiparametric encoding is performed at 1.5 T. The T2∗ and fat fraction are estimated from a graph/cut water/fat separation method using a six-peak fat model. Water/fat singular images obtained are then matched to an MRF dictionary, estimating water-specific T1 , T2 , and T1ρ . The proposed approach was tested in phantoms and 10 healthy subjects and compared against conventional sequences. RESULTS: For the phantom studies, linear fits show excellent coefficients of determination (r2 > 0.9) for every parametric map. For in vivo studies, the average values measured within regions of interest drawn on liver, spleen, muscle, and fat are statistically different from the reference scans (p < 0.05) for T1 , T2 , and T1â´ but not for T2∗ and fat fraction, whereas correlation between MRF and reference scans is excellent for each parameter (r2 > 0.92 for every parameter). CONCLUSION: The proposed multi-echo inversion-recovery, T2 , and T1â´ prepared liver MRF sequence presented in this work allows for quantitative T1 , T2 , T2∗ , T1â´ , and fat fraction liver-tissue characterization in a single breath-hold scan of 18 seconds. The approach showed good agreement and correlation with respect to reference clinical maps.
Assuntos
Processamento de Imagem Assistida por Computador , Imageamento por Ressonância Magnética , Suspensão da Respiração , Humanos , Processamento de Imagem Assistida por Computador/métodos , Fígado/diagnóstico por imagem , Imageamento por Ressonância Magnética/métodos , Imagens de FantasmasRESUMO
PURPOSE: To develop a simultaneous T1 , T2 , and T1ρ cardiac magnetic resonance fingerprinting (MRF) approach to enable comprehensive contrast agent-free myocardial tissue characterization in a single breath-hold scan. METHODS: A 2D gradient-echo electrocardiogram-triggered cardiac MRF sequence with low flip angles, varying magnetization preparation, and spiral trajectory was acquired at 1.5 T to encode T1 , T2 , and T1â´ simultaneously. The MRF images were reconstructed using low-rank inversion, regularized with a multicontrast patch-based higher-order reconstruction. Parametric maps were generated and matched in the singular value domain to extended phase graph-based dictionaries. The proposed approach was tested in phantoms and 10 healthy subjects and compared against conventional methods in terms of coefficients of determination and best fits for the phantom study, and in terms of Bland-Altman agreement, average values and coefficient of variation of T1 , T2 , and T1â´ for the healthy subjects study. RESULTS: The T1 , T2 , and T1â´ MRF values showed excellent correlation with conventional spin-echo and clinical mapping methods in phantom studies (r2 > 0.97). Measured MRF values in myocardial tissue (mean ± SD) were 1133 ± 33 ms, 38.8 ± 3.5 ms, and 52.0 ± 4.0 ms for T1 , T2 and T1â´ , respectively, against 1053 ± 47 ms, 50.4 ± 3.9 ms, and 55.9 ± 3.3 ms for T1 modified Look-Locker inversion imaging, T2 gradient and spin echo, and T1â´ turbo field echo, respectively. CONCLUSION: A cardiac MRF approach for simultaneous quantification of myocardial T1 , T2 , and T1ρ in a single breath-hold MR scan of about 16 seconds has been proposed. The approach has been investigated in phantoms and healthy subjects showing good agreement with reference spin echo measurements and conventional clinical maps.
Assuntos
Meios de Contraste , Imageamento por Ressonância Magnética , Coração/diagnóstico por imagem , Humanos , Processamento de Imagem Assistida por Computador , Espectroscopia de Ressonância Magnética , Imagens de FantasmasRESUMO
PURPOSE: Develop a novel 2D cardiac MR fingerprinting (MRF) approach to enable simultaneous T1, T2, T2*, and fat fraction (FF) myocardial tissue characterization in a single breath-hold scan. METHODS: Simultaneous, co-registered, multi-parametric mapping of T1, T2, and FF has been recently achieved with cardiac MRF. Here, we further incorporate T2* quantification within this approach, enabling simultaneous T1, T2, T2*, and FF myocardial tissue characterization in a single breath-hold scan. T2* quantification is achieved with an eight-echo readout that requires a long cardiac acquisition window. A novel low-rank motion-corrected (LRMC) reconstruction is exploited to correct for cardiac motion within the long acquisition window. The proposed T1/T2/T2*/FF cardiac MRF was evaluated in phantom and in 10 healthy subjects in comparison to conventional mapping techniques. RESULTS: The proposed approach achieved high quality parametric mapping of T1, T2, T2*, and FF with corresponding normalized RMS error (RMSE) T1 = 5.9%, T2 = 9.6% (T2 values <100 ms), T2* = 3.3% (T2* values <100 ms), and FF = 0.8% observed in phantom scans. In vivo, the proposed approach produced higher left-ventricular myocardial T1 values than MOLLI (1148 vs 1056 ms), lower T2 values than T2-GraSE (42.8 vs 50.6 ms), lower T2* values than eight-echo gradient echo (GRE) (35.0 vs 39.4 ms), and higher FF values than six-echo GRE (0.8 vs 0.3 %) reference techniques. The proposed approach achieved considerable reduction in motion artifacts compared to cardiac MRF without motion correction, improved spatial uniformity, and statistically higher apparent precision relative to conventional mapping for all parameters. CONCLUSION: The proposed cardiac MRF approach enables simultaneous, co-registered mapping of T1, T2, T2*, and FF in a single breath-hold for comprehensive myocardial tissue characterization, achieving higher apparent precision than conventional methods.
Assuntos
Coração , Imageamento por Ressonância Magnética , Suspensão da Respiração , Coração/diagnóstico por imagem , Humanos , Miocárdio , Imagens de Fantasmas , Reprodutibilidade dos TestesRESUMO
INTRODUCTION AND OBJECTIVES: Non-alcoholic fatty liver disease (NAFLD) encompasses a spectrum of liver abnormalities including steatosis, steatohepatitis, fibrosis, and cirrhosis. Liver biopsy remains the gold standard method to determine the disease stage in NAFLD but is an invasive and risky procedure. Studies have previously reported that changes in intrahepatic fatty acids (FA) composition are related to the progression of NAFLD, mainly in its early stages. The aim of this study was to characterize the liver FA composition in mice fed a Choline-deficient L-amino-defined (CDAA) diet at different stages of NAFLD using magnetic resonance spectroscopy (MRS). METHODS: We used in-vivo MRS to perform a longitudinal characterization of hepatic FA changes in NAFLD mice for 10 weeks. We validated our findings with ex-vivo MRS, gas chromatography-mass spectrometry and histology. RESULTS: In-vivo and ex-vivo results showed that livers from CDAA-fed mice exhibit a significant increase in liver FA content as well as a change in FA composition compared with control mice. After 4 weeks of CDAA diet, a decrease in polyunsaturated and an increase in monounsaturated FA were observed. These changes were associated with the appearance of early stages of steatohepatitis, confirmed by histology (NAFLD Activity Score (NAS) = 4.5). After 10 weeks of CDAA-diet, the liver FA composition remained stable while the NAS increased further to 6 showing a combination of early and late stages of steatohepatitis. CONCLUSION: Our results suggest that monitoring lipid composition in addition to total water/fat with MRS may yield additional insights that can be translated for non-invasive stratification of high-risk NAFLD patients.
Assuntos
Ácidos Graxos/metabolismo , Espectroscopia de Ressonância Magnética , Hepatopatia Gordurosa não Alcoólica/diagnóstico , Hepatopatia Gordurosa não Alcoólica/metabolismo , Animais , Modelos Animais de Doenças , Progressão da Doença , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Hepatopatia Gordurosa não Alcoólica/etiologia , Valor Preditivo dos Testes , Reprodutibilidade dos TestesRESUMO
PURPOSE: Cardiac magnetic resonance fingerprinting (cMRF) has been recently introduced to simultaneously provide T1 , T2 , and M0 maps. Here, we develop a 3-point Dixon-cMRF approach to enable simultaneous water specific T1 , T2 , and M0 mapping of the heart and fat fraction (FF) estimation in a single breath-hold scan. METHODS: Dixon-cMRF is achieved by combining cMRF with several innovations that were previously introduced for other applications, including a 3-echo GRE acquisition with golden angle radial readout and a high-dimensional low-rank tensor constrained reconstruction to recover the highly undersampled time series images for each echo. Water-fat separation of the Dixon-cMRF time series is performed to allow for water- and fat-specific T1 , T2 , and M0 estimation, whereas FF estimation is extracted from the M0 maps. Dixon-cMRF was evaluated in a standardized T1 -T2 phantom, in a water-fat phantom, and in healthy subjects in comparison to current clinical standards: MOLLI, SASHA, T2 -GRASE, and 6-point Dixon proton density FF (PDFF) mapping. RESULTS: Dixon-cMRF water T1 and T2 maps showed good agreement with reference T1 and T2 mapping techniques (R2 > 0.99 and maximum normalized RMSE ~5%) in a standardized phantom. Good agreement was also observed between Dixon-cMRF FF and reference PDFF (R2 > 0.99) and between Dixon-cMRF water T1 and T2 and water selective T1 and T2 maps (R2 > 0.99) in a water-fat phantom. In vivo Dixon-cMRF water T1 values were in good agreement with MOLLI and water T2 values were slightly underestimated when compared to T2 -GRASE. Average myocardium septal T1 values were 1129 ± 38 ms, 1026 ± 28 ms, and 1045 ± 32 ms for SASHA, MOLLI, and the proposed water Dixon-cMRF. Average T2 values were 51.7 ± 2.2 ms and 42.8 ± 2.6 ms for T2 -GRASE and water Dixon-cMRF, respectively. Dixon-cMRF FF maps showed good agreement with in vivo PDFF measurements (R2 > 0.98) and average FF in the septum was measured at 1.3%. CONCLUSION: The proposed Dixon-cMRF allows to simultaneously quantify myocardial water T1 , water T2 , and FF in a single breath-hold scan, enabling multi-parametric T1 , T2 , and fat characterization. Moreover, reduced T1 and T2 quantification bias caused by water-fat partial volume was demonstrated in phantom experiments.
Assuntos
Processamento de Imagem Assistida por Computador , Água , Coração/diagnóstico por imagem , Humanos , Imageamento por Ressonância Magnética , Espectroscopia de Ressonância Magnética , Imagens de Fantasmas , Reprodutibilidade dos TestesRESUMO
OBJECTIVE: Nitric oxide synthase 3 (NOS3) prevents neointima hyperplasia by still unknown mechanisms. To demonstrate the significance of endothelial nitric oxide in the polarization of infiltrated macrophages through the expression of matrix metalloproteinase (MMP)-13 in neointima formation. APPROACH AND RESULTS: After aortic endothelial denudation, NOS3 null mice show elevated neointima formation, detecting increased mobilization of LSK (lineage-negative [Lin]-stem-cell antigen 1 [SCA1]+KIT+) progenitor cells, and high ratios of M1 (proinflammatory) to M2 (resolving) macrophages, accompanied by high expression of interleukin-5, interleukin-6, MCP-1 (monocyte chemoattractant protein), VEGF (vascular endothelial growth factor), GM-CSF (granulocyte-macrophage colony stimulating factor), interleukin-1ß, and interferon-γ. In conditional c-Myc knockout mice, in which M2 polarization is defective, denuded aortas showed extensive wall thickening as well. Conditioned medium from NOS3-deficient endothelium induced extensive repolarization of M2 macrophages to an M1 phenotype, and vascular smooth muscle cells proliferated and migrated faster in conditioned medium from M1 macrophages. Among the different proteins participating in cell migration, MMP-13 was preferentially expressed by M1 macrophages. M1-mediated vascular smooth muscle cell migration was inhibited when macrophages were isolated from MMP-13-deficient mice, whereas exogenous administration of MMP-13 to vascular smooth muscle cell fully restored migration. Excess vessel wall thickening in mice lacking NOS3 was partially reversed by simultaneous deletion of MMP-13, indicating that NOS3 prevents neointimal hyperplasia by preventing MMP-13 activity. An excess of M1-polarized macrophages that coexpress MMP-13 was also detected in human carotid samples from endarterectomized patients. CONCLUSIONS: These findings indicate that at least M1 macrophage-mediated expression of MMP-13 in NOS3 null mice induces neointima formation after vascular injury, suggesting that MMP-13 may represent a new promising target in vascular disease.
Assuntos
Doenças da Aorta/metabolismo , Macrófagos/metabolismo , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Neointima , Óxido Nítrico/metabolismo , Idoso , Idoso de 80 Anos ou mais , Animais , Aorta/metabolismo , Aorta/patologia , Doenças da Aorta/enzimologia , Doenças da Aorta/genética , Doenças da Aorta/patologia , Biomarcadores/metabolismo , Estudos de Casos e Controles , Diferenciação Celular , Linhagem da Célula , Movimento Celular , Proliferação de Células , Células Cultivadas , Modelos Animais de Doenças , Feminino , Humanos , Hiperplasia , Mediadores da Inflamação/metabolismo , Macrófagos/enzimologia , Macrófagos/patologia , Masculino , Metaloproteinase 13 da Matriz/deficiência , Metaloproteinase 13 da Matriz/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pessoa de Meia-Idade , Músculo Liso Vascular/enzimologia , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/enzimologia , Miócitos de Músculo Liso/patologia , Óxido Nítrico Sintase Tipo III/deficiência , Óxido Nítrico Sintase Tipo III/genética , Fenótipo , Proteínas Proto-Oncogênicas c-myc/deficiência , Proteínas Proto-Oncogênicas c-myc/genética , Fatores de TempoRESUMO
Cardiovascular diseases are the first leading cause of death and morbidity in developed countries. The use of animal models have contributed to increase our knowledge, providing new approaches focused to improve the diagnostic and the treatment of these pathologies. Several models have been developed to address cardiovascular complications, including atherothrombotic and cardiac diseases, and the same pathology have been successfully recreated in different species, including small and big animal models of disease. However, genetic and environmental factors play a significant role in cardiovascular pathophysiology, making difficult to match a particular disease, with a single experimental model. Therefore, no exclusive method perfectly recreates the human complication, and depending on the model, additional considerations of cost, infrastructure, and the requirement for specialized personnel, should also have in mind. Considering all these facts, and depending on the budgets available, models should be selected that best reproduce the disease being investigated. Here we will describe models of atherothrombotic diseases, including expanding and occlusive animal models, as well as models of heart failure. Given the wide range of models available, today it is possible to devise the best strategy, which may help us to find more efficient and reliable solutions against human cardiovascular diseases.
Assuntos
Aneurisma Aórtico , Aterosclerose , Modelos Animais de Doenças , Insuficiência Cardíaca , Trombose , Animais , Aneurisma Aórtico/etiologia , Aneurisma Aórtico/genética , Aneurisma Aórtico/fisiopatologia , Apolipoproteínas E/genética , Aterosclerose/etiologia , Aterosclerose/genética , Aterosclerose/fisiopatologia , Cães , Insuficiência Cardíaca/etiologia , Insuficiência Cardíaca/genética , Insuficiência Cardíaca/fisiopatologia , Humanos , Camundongos , Coelhos , Ratos , Receptores de LDL/genética , Suínos , Trombose/etiologia , Trombose/genética , Trombose/fisiopatologiaRESUMO
Cardiovascular diseases are the leading causes of death worldwide. A permeable/leaky and dysfunctional endothelium is considered the earliest marker of vascular damage and thought to drive atherosclerosis. A method to identify these changes in vivo would be desirable in the clinic. Magnetic resonance imaging (MRI)-based tools and other technologies have enabled a profound understanding of the role of the endothelium in cardiovascular diseases and risk in vivo. There is, however, a need for reproducible and simple approaches for extracting quantifiable data reflective of endothelial damage from a single imaging study. A non-invasive, easy-to-implement, and quantitative MRI workflow was developed to acquire and analyze images that allow the quantification of two imaging biomarkers of arterial endothelial damage (leakiness/permeability and dysfunction). Here, the protocol describes the application of this method in the brachiocephalic artery of atherosclerotic ApoE-/- mice using a clinical MRI scanner. First, late gadolinium enhancement (LGE) and Modified Look-Locker Inversion Recovery (MOLLI) T1 mapping protocols to quantify endothelial leakage using an albumin-binding probe are described. Second, anatomic, and quantitative blood flow sequences to measure endothelial dysfunction, in response to acetylcholine are described. Importantly, the method outlined here allows the acquisition of high-spatial-resolution 3D images with large volumetric coverage enabling accurate segmentation of vessel wall structures to improve inter- and intra-observer variability and to increase reliability and reproducibility. Additionally, it provides quantitative data without the need for high-temporal resolution for complex kinetic modeling, making it model-independent and even allowing for imaging of highly mobile vessels (coronary arteries). Therefore, the approach simplifies and expedites data analysis. Finally, this method can be implemented on different scanners, can be extended to image different arterial beds, and is clinically applicable for use in humans. This method could be used to diagnose and treat patients with atherosclerosis by adopting a precision-medicine approach.
Assuntos
Aterosclerose , Gadolínio , Animais , Aterosclerose/diagnóstico por imagem , Meios de Contraste , Endotélio Vascular/diagnóstico por imagem , Humanos , Imageamento por Ressonância Magnética/métodos , Camundongos , Permeabilidade , Reprodutibilidade dos TestesRESUMO
Dysfunctional elastin turnover plays a major role in the progression of atherosclerotic plaques. Failure of tropoelastin cross-linking into mature elastin leads to the accumulation of tropoelastin within the growing plaque, increasing its instability. Here we present Gd4-TESMA, an MRI contrast agent specifically designed for molecular imaging of tropoelastin within plaques. Gd4-TESMA is a tetrameric probe composed of a tropoelastin-binding peptide (the VVGS-peptide) conjugated with four Gd(III)-DOTA-monoamide chelates. It shows a relaxivity per molecule of 34.0 ± 0.8 mM-1 s-1 (20 MHz, 298 K, pH 7.2), a good binding affinity to tropoelastin (KD = 41 ± 12 µM), and a serum half-life longer than 2 h. Gd4-TESMA accumulates specifically in atherosclerotic plaques in the ApoE-/- murine model of plaque progression, with 2 h persistence of contrast enhancement. As compared to the monomeric counterpart (Gd-TESMA), the tetrameric Gd4-TESMA probe shows a clear advantage regarding both sensitivity and imaging time window, allowing for a better characterization of atherosclerotic plaques.
Assuntos
Aterosclerose/metabolismo , Meios de Contraste/química , Elastina/metabolismo , Gadolínio/química , Imageamento por Ressonância Magnética , Tropoelastina/análise , Animais , Meios de Contraste/síntese química , Modelos Animais de Doenças , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Estrutura Molecular , Ressonância de Plasmônio de SuperfícieRESUMO
Nitric Oxide (NO) is involved in the development and progression of abdominal aortic aneurysms (AAA). We found that inhibition of inducible NO synthase (iNOS) protects mice in an elastase-induced AAA model, significantly inhibiting the production of matrix metalloproteinase-13 (MMP-13). The extracellular MMP inducer (EMMPRIN; CD147) was increased in human AAA biopsies and in wild-type murine AAA but not in AAA from iNOS null mice. In cells overexpressing ectopic EMMPRIN, MMP-13 secretion was stimulated, whereas silencing of EMMPRIN by RNA interference led to significant inhibition of MMP-13 expression. In addition, elastase infusion of MMP-13 null mouse aortas induced a significant increase of EMMPRIN but reduced aortic dilatation when compared with wild-type mice, suggesting that NO-mediated AAA may be mediated through EMMPRIN induction of MMP-13. These findings were further verified in elastase-infused iNOS null mice, in which daily administration of NO caused a significant aortic dilatation and the expression of EMMPRIN and MMP-13. By contrast, in iNOS wild-type mice, pharmacological inhibition of iNOS by administration of 1400 W induced a reduction of aortic diameter and inhibition of MMP-13 and EMMPRIN expression when compared with control mice. Our results suggest that NO may regulate the development of AAA in part by inducing the expression of EMMPRIN and modulating the activity of MMP-13 in murine and human aneurysms.
Assuntos
Aneurisma da Aorta Abdominal/enzimologia , Aneurisma da Aorta Abdominal/patologia , Basigina/metabolismo , Metaloproteinase 13 da Matriz/biossíntese , Óxido Nítrico/metabolismo , Animais , Progressão da Doença , Regulação para Baixo , Endotélio Vascular/enzimologia , Endotélio Vascular/patologia , Indução Enzimática , Espaço Extracelular/enzimologia , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Óxido Nítrico Sintase Tipo II/deficiência , Óxido Nítrico Sintase Tipo II/metabolismo , Tirosina/análogos & derivados , Tirosina/metabolismoRESUMO
AIMS: Dysfunctional matrix turnover is present at sites of abdominal aortic aneurysm (AAA) and leads to the accumulation of monomeric tropoelastin rather than cross-linked elastin. We used a gadolinium-based tropoelastin-specific magnetic resonance contrast agent (Gd-TESMA) to test whether quantifying regional tropoelastin turnover correlates with aortic expansion in a murine model. The binding of Gd-TESMA to excised human AAA was also assessed. METHODS AND RESULTS: We utilized the angiotensin II (Ang II)-infused apolipoprotein E gene knockout (ApoE-/-) murine model of aortic dilation and performed in vivo imaging of tropoelastin by administering Gd-TESMA followed by late gadolinium enhancement (LGE) magnetic resonance imaging (MRI) and T1 mapping at 3 T, with subsequent ex vivo validation. In a cross-sectional study (n = 66; control = 11, infused = 55) we found that Gd-TESMA enhanced MRI was elevated and confined to dilated aortic segments (control: LGE=0.13 ± 0.04 mm2, control R1= 1.1 ± 0.05 s-1 vs. dilated LGE =1.0 ± 0.4 mm2, dilated R1 =2.4 ± 0.9 s-1) and was greater in segments with medium (8.0 ± 3.8 mm3) and large (10.4 ± 4.1 mm3) compared to small (3.6 ± 2.1 mm3) vessel volume. Furthermore, a proof-of-principle longitudinal study (n = 19) using Gd-TESMA enhanced MRI demonstrated a greater proportion of tropoelastin: elastin expression in dilating compared to non-dilating aortas, which correlated with the rate of aortic expansion. Treatment with pravastatin and aspirin (n = 10) did not reduce tropoelastin turnover (0.87 ± 0.3 mm2 vs. 1.0 ± 0.44 mm2) or aortic dilation (4.86 ± 2.44 mm3 vs. 4.0 ± 3.6 mm3). Importantly, Gd-TESMA-enhanced MRI identified accumulation of tropoelastin in excised human aneurysmal tissue (n = 4), which was confirmed histologically. CONCLUSION: Tropoelastin MRI identifies dysfunctional matrix remodelling that is specifically expressed in regions of aortic aneurysm or dissection and correlates with the development and rate of aortic expansion. Thus, it may provide an additive imaging marker to the serial assessment of luminal diameter for surveillance of patients at risk of or with established aortopathy.
Assuntos
Aorta Abdominal/diagnóstico por imagem , Aneurisma da Aorta Abdominal/diagnóstico por imagem , Dissecção Aórtica/diagnóstico por imagem , Meios de Contraste/administração & dosagem , Matriz Extracelular/metabolismo , Imageamento por Ressonância Magnética , Tropoelastina/metabolismo , Remodelação Vascular , Dissecção Aórtica/induzido quimicamente , Dissecção Aórtica/metabolismo , Dissecção Aórtica/patologia , Angiotensina II , Animais , Aorta Abdominal/metabolismo , Aorta Abdominal/patologia , Aneurisma da Aorta Abdominal/induzido quimicamente , Aneurisma da Aorta Abdominal/metabolismo , Aneurisma da Aorta Abdominal/patologia , Biomarcadores/metabolismo , Meios de Contraste/metabolismo , Dilatação Patológica , Modelos Animais de Doenças , Progressão da Doença , Matriz Extracelular/patologia , Humanos , Camundongos Knockout para ApoE , Valor Preditivo dos Testes , Estudo de Prova de Conceito , Fatores de Tempo , Regulação para CimaRESUMO
Overview: Cardiovascular disease remains a leading cause of death worldwide, with vulnerable plaque rupture the underlying cause of many heart attacks and strokes. Much research is focused on identifying an imaging biomarker to differentiate stable and vulnerable plaque. Magnetic Resonance Imaging (MRI) is a non-ionising and non-invasive imaging modality with excellent soft tissue contrast. However, MRI has relatively low sensitivity (micromolar) for contrast agent detection compared to nuclear imaging techniques. There is also an increasing emphasis on developing MRI probes that are not based on gadolinium chelates because of increasing concerns over associated systemic toxicity and deposits1. To address the sensitivity and safety concerns of gadolinium this project focused on the development of a high relaxivity probe based on superparamagnetic iron oxide nanoparticles for the imaging of atherosclerotic plaque with MRI. With development, this may facilitate differentiating stable and vulnerable plaque in vivo.Aim: To develop a range of MRI contrast agents based on superparamagnetic iron oxide nanoparticles (SPIONs), and test them in a murine model of advanced atherosclerosis. Methods: Nanoparticles of four core sizes were synthesised by thermal decomposition and coated with poly(maleicanhydride-alt-1-octadecene) (PMAO), poly(ethyleneimine) (PEI) or alendronate, then characterised for core size, hydrodynamic size, surface potential and relaxivity. On the basis of these results, one candidate was selected for further studies. In vivo studies using 10 nm PMAO-coated SPIONs were performed in ApoE-/- mice fed a western diet and instrumented with a perivascular cuff on the left carotid artery. Control ApoE-/- mice were fed a normal chow diet and were not instrumented. Mice were scanned on a 3T MR scanner (Philips Achieva) with the novel SPION contrast agent, and an elastin-targeted gadolinium agent that was shown previously to enable visualisation of plaque burden. Histological analysis was undertaken to confirm imaging findings through staining for macrophages, CX3CL1, elastin, tropoelastin, and iron. Results: The lead SPION agent consisted of a 10 nm iron oxide core with poly(maleicanhydride-alt-1-octadecene), (-36.21 mV, r2 18.806 mmol-1/s-1). The irregular faceting of the iron oxide core resulted in high relaxivity and the PMAO provided a foundation for further functionalisation on surface -COOH groups. The properties of the contrast agent, including the negative surface charge and hydrodynamic size, were designed to maximise circulation time and evade rapid clearance through the renal system or phagocytosis. In vitro testing showed that the SPION agent was non-toxic. In vivo results show that the novel contrast agent accumulates in similar vascular regions to a gadolinium-based contrast agent (Gd-ESMA) targeted to elastin, which accumulates in plaque. There was a significant difference in SPION signal between the instrumented and the contralateral non-instrumented vessels in diseased mice (p = 0.0411, student's t-test), and between the instrumented diseased vessel and control vessels (p = 0.0043, 0.0022, student's t-test). There was no significant difference between the uptake of either contrast agent between stable and vulnerable plaques (p = 0.3225, student's t-test). Histological verification was used to identify plaques, and Berlin Blue staining confirmed the presence of nanoparticle deposits within vulnerable plaques and co-localisation with macrophages. Conclusion: This work presents a new MRI contrast agent for atherosclerosis which uses an under-explored surface ligand, demonstrating promising properties for in vivo behaviour, is still in circulation 24 hours post-injection with limited liver uptake, and shows good accumulation in a murine plaque model.
Assuntos
Meios de Contraste , Imageamento por Ressonância Magnética/métodos , Nanopartículas de Magnetita/química , Imagem Molecular/métodos , Placa Aterosclerótica , Animais , Artérias Carótidas/diagnóstico por imagem , Artérias Carótidas/patologia , Meios de Contraste/química , Meios de Contraste/farmacocinética , Dieta Hiperlipídica , Feminino , Camundongos , Camundongos Knockout , Placa Aterosclerótica/diagnóstico por imagem , Placa Aterosclerótica/metabolismo , Placa Aterosclerótica/patologiaRESUMO
BACKGROUND AND AIMS: Acute ischemia is associated with myocardial endothelial damage and microvessel formation, resulting in leakage of plasma albumin into the myocardial extravascular space. In this study, we tested whether an albumin-binding intravascular contrast agent (gadofosveset) allows for improved quantification of myocardial permeability compared to the conventional extracellular contrast agent Gd-DTPA using late gadolinium enhancement (LGE) and T1 mapping in vivo. METHODS: MI was induced in C57BL/6 mice (nâ¯=â¯6) and cardiac magnetic resonance imaging (CMR) was performed at 3, 10 and 21 days post-MI using Gd-DTPA and 24â¯h later using gadofosveset. Functional, LGE and T1 mapping protocols were performed 45 min post-injection of the contrast agent. RESULTS: LGE images showed that both contrast agents provided similar measurements of infarct area at all time points following MI. Importantly, the myocardial R1 measurements after administration of gadofosveset were higher in the acute phase-day 3 (R1 [s-1]â¯=â¯6.29⯱â¯0.29) compared to the maturation phase-days 10 and 21 (R1 [s-1]â¯=â¯4.76⯱â¯0.30 and 4.48⯱â¯0.14), suggesting that the uptake of this agent could be used to stage myocardial remodeling. No differences in myocardial R1 were observed after administration of Gd-DTPA at different time points post-MI (R1 [s-1]â¯=â¯3d: 3.77⯱â¯0.37; 10d: 2.74⯱â¯0.06; 21d: 3.35⯱â¯0.26). The MRI results were validated by ex vivo histology that showed albumin leakage in the myocardium in the acute phase and microvessel formation at later stages. CONCLUSIONS: We demonstrate the merits of an albumin-binding contrast agent for monitoring changes in myocardial permeability between acute ischemia and chronic post-MI myocardial remodeling.
Assuntos
Permeabilidade Capilar , Meios de Contraste/administração & dosagem , Vasos Coronários/metabolismo , Gadolínio DTPA/administração & dosagem , Gadolínio/administração & dosagem , Imagem Cinética por Ressonância Magnética/métodos , Infarto do Miocárdio/diagnóstico por imagem , Compostos Organometálicos/administração & dosagem , Animais , Meios de Contraste/farmacocinética , Vasos Coronários/fisiopatologia , Modelos Animais de Doenças , Feminino , Gadolínio/farmacocinética , Gadolínio DTPA/farmacocinética , Camundongos Endogâmicos C57BL , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/patologia , Infarto do Miocárdio/fisiopatologia , Miocárdio/patologia , Compostos Organometálicos/farmacocinética , Valor Preditivo dos Testes , Fatores de Tempo , Função Ventricular Esquerda , Remodelação VentricularRESUMO
Background: Elastolysis and ineffective elastogenesis favor the accumulation of tropoelastin, rather than cross-linked elastin, in atherosclerotic plaques. We developed gadolinium-labeled tropoelastin-specific magnetic resonance contrast agents (Gd-TESMAs) for tropoelastin imaging in animal models. Methods and Results: Two peptides, VVGSPSAQDEASPLS and YPDHVQYTHY were selected to target tropoelastin. In vitro binding, relaxivity, and biodistribution experiments enabled characterization of the probes and selecting the best candidate for in vivo MRI. MRI was performed in atherosclerotic apolipoprotein E-deficient (ApoE-/-) mice and New Zealand white rabbits with stable and rupture-prone plaques using Gd-TESMA. Additionally, human carotid endarterectomy specimens were imaged ex vivo. The VVGSPSAQDEASPLS-based probe discriminated between tropoelastin and cross-linked elastin (64±7% vs 1±2%, P=0.001), had high in vitro relaxivity in solution (r1-free=11.7±0.6mM-1s-1, r1-bound to tropoelastin = 44±1mM-1s-1) and favorable pharmacokinetics. In vivo mice vascular enhancement (4wks=0.13±0.007mm2, 8wks=0.22±0.01mm2, 12wks=0.33±0.01mm2, P<0.001) and R1 relaxation rate (4wks=0.90±0.01 s-1, 8wks=1.40±0.03 s-1, 12wks=1.87±0.04s-1, P<0.001) increased with atherosclerosis progression after Gd-TESMA injection. Conversely, statin-treated (0.13±0.01mm2, R1 =1.37±0.03s-1) and control (0.10±0.005mm2, R1 =0.87±0.05s-1) mice showed less enhancement. Rupture-prone rabbit plaques had higher R1 relaxation rate compared with stale plaques (R1=2.26±0.1s-1vs R1=1.43±0.02s-1, P=0.001), after administration of Gd-TESMA that allowed detection of rupture-prone plaques with high sensitivity (84.4%) and specificity (92.3%). Increased vascular R1 relaxation rate was observed in carotid endarterectomy plaques after soaking (R1pre= 1.1±0.26 s-1 vs R1post= 3.0±0.1s-1, P=0.01). Ex vivo analyses confirmed the MRI findings and showed uptake of the contrast agent to be specific for tropoelastin. Conclusions: MRI of tropoelastin provides a novel biomarker for atherosclerotic plaque progression and instability.
Assuntos
Doenças da Aorta/diagnóstico por imagem , Aterosclerose/diagnóstico por imagem , Doenças das Artérias Carótidas/diagnóstico por imagem , Meios de Contraste/administração & dosagem , Compostos Heterocíclicos/administração & dosagem , Imageamento por Ressonância Magnética , Imagem Molecular/métodos , Oligopeptídeos/administração & dosagem , Compostos Organometálicos/administração & dosagem , Placa Aterosclerótica , Tropoelastina/metabolismo , Animais , Doenças da Aorta/metabolismo , Doenças da Aorta/patologia , Aterosclerose/genética , Aterosclerose/metabolismo , Aterosclerose/patologia , Biomarcadores/metabolismo , Doenças das Artérias Carótidas/metabolismo , Doenças das Artérias Carótidas/patologia , Meios de Contraste/farmacocinética , Modelos Animais de Doenças , Progressão da Doença , Compostos Heterocíclicos/farmacocinética , Humanos , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout para ApoE , Oligopeptídeos/farmacocinética , Compostos Organometálicos/farmacocinética , Valor Preditivo dos Testes , Coelhos , Ruptura EspontâneaRESUMO
Background: Optimal healing of the myocardium following myocardial infarction (MI) requires a suitable degree of inflammation and its timely resolution, together with a well-orchestrated deposition and degradation of extracellular matrix (ECM) proteins. Methods and Results: MI and SHAM-operated animals were imaged at 3,7,14 and 21 days with 3T magnetic resonance imaging (MRI) using a 19F/1H surface coil. Mice were injected with 19F-perfluorocarbon (PFC) nanoparticles to study inflammatory cell recruitment, and with a gadolinium-based elastin-binding contrast agent (Gd-ESMA) to evaluate elastin content. 19F MRI signal co-localized with infarction areas, as confirmed by late-gadolinium enhancement, and was highest 7days post-MI, correlating with macrophage content (MAC-3 immunohistochemistry) (ρ=0.89,P<0.0001). 19F quantification with in vivo (MRI) and ex vivo nuclear magnetic resonance (NMR) spectroscopy correlated linearly (ρ=0.58,P=0.020). T1 mapping after Gd-ESMA injection showed increased relaxation rate (R1) in the infarcted regions and was significantly higher at 21days compared with 7days post-MI (R1[s-1]:21days=2.8 [IQR,2.69-3.30] vs 7days=2.3 [IQR,2.12-2.5], P<0.05), which agreed with an increased tropoelastin content (ρ=0.89, P<0.0001). The predictive value of each contrast agent for beneficial remodeling was evaluated in a longitudinal proof-of-principle study. Neither R1 nor 19F at day 7 were significant predictors for beneficial remodeling (P=0.68;P=0.062). However, the combination of both measurements (R1<2.34Hz and 0.55≤19F≤1.85) resulted in an odds ratio of 30.0 (CI95%:1.41-638.15;P=0.029) for favorable post-MI remodeling. Conclusions: Multinuclear 1H/19F MRI allows the simultaneous assessment of inflammation and elastin remodeling in a murine MI model. The interplay of these biological processes affects cardiac outcome and may have potential for improved diagnosis and personalized treatment.
Assuntos
Proteínas da Matriz Extracelular/metabolismo , Infarto do Miocárdio/complicações , Miocardite/metabolismo , Miocárdio/metabolismo , Animais , Biomarcadores/metabolismo , Modelos Animais de Doenças , Feminino , Imuno-Histoquímica , Imagem Cinética por Ressonância Magnética , Espectroscopia de Ressonância Magnética , Camundongos , Camundongos Endogâmicos C57BL , Infarto do Miocárdio/diagnóstico , Infarto do Miocárdio/metabolismo , Miocardite/diagnóstico , Miocardite/etiologia , Miocárdio/patologiaRESUMO
BACKGROUND: Compromised structural integrity of the endothelium and higher microvessel density increase vascular permeability. We investigated whether vascular permeability measured in vivo by magnetic resonance imaging using the albumin-binding contrast agent, gadofosveset, is a surrogate marker of rupture-prone atherosclerotic plaque in a rabbit model. METHODS AND RESULTS: New Zealand white rabbits (n=10) were rendered atherosclerotic by cholesterol-diet and endothelial denudation. Plaque rupture was triggered with Russell's viper venom and histamine. Animals were imaged pre-triggering, at 3 and 12 weeks, to quantify plaque area, vascular permeability, vasodilation, and stiffness and post-triggering to identify thrombus. Plaques identified on the pretrigger scans were classified as stable or rupture-prone based on the absence or presence of thrombus on the corresponding post-trigger magnetic resonance imaging, respectively. All rabbits had developed atherosclerosis, and 60% had ruptured plaques. Rupture-prone plaques had higher vessel wall relaxation rate (R1; 2.30±0.5 versus 1.86±0.3 s-1; P<0.001), measured 30 minutes after gadofosveset administration, and higher R1/plaque area ratio (0.70±0.06 versus 0.47±0.02, P= 0.01) compared with stable plaque at 12 weeks. Rupture-prone plaques had higher percent change in R1 between the 3 and 12 weeks compared with stable plaque (50.80±7.2% versus 14.22±2.2%; P<0.001). Immunohistochemistry revealed increased vessel wall albumin and microvessel density in diseased aortas and especially in ruptured plaque. Electron microscopy showed lack of structural integrity in both luminal and microvascular endothelium in diseased vessels. Functionally, the intrinsic vasodilation of the vessel wall decreased at 12 weeks compared with 3 weeks (18.60±1.0% versus 23.43±0.8%; P<0.001) and in rupture-prone compared with stable lesions (16.40±2.0% versus 21.63±1.2%; P<0.001). Arterial stiffness increased at 12 weeks compared with 3 weeks (5.00±0.1 versus 2.53±0.2 m/s; P<0.001) both in animals with stable and rupture-prone lesions. CONCLUSIONS: T1 mapping using an albumin-binding contrast agent (gadofosveset) could quantify the changes in vascular permeability associated with atherosclerosis progression and rupture-prone plaques.
Assuntos
Aorta/diagnóstico por imagem , Doenças da Aorta/diagnóstico por imagem , Aterosclerose/diagnóstico por imagem , Permeabilidade Capilar , Meios de Contraste/metabolismo , Gadolínio/metabolismo , Imageamento por Ressonância Magnética , Microvasos/diagnóstico por imagem , Compostos Organometálicos/metabolismo , Placa Aterosclerótica , Albumina Sérica/metabolismo , Animais , Aorta/metabolismo , Aorta/fisiopatologia , Doenças da Aorta/etiologia , Doenças da Aorta/metabolismo , Doenças da Aorta/fisiopatologia , Área Sob a Curva , Aterosclerose/etiologia , Aterosclerose/metabolismo , Aterosclerose/fisiopatologia , Colesterol na Dieta , Modelos Animais de Doenças , Progressão da Doença , Endotélio Vascular/diagnóstico por imagem , Endotélio Vascular/metabolismo , Endotélio Vascular/fisiopatologia , Histamina , Masculino , Microvasos/metabolismo , Microvasos/fisiopatologia , Valor Preditivo dos Testes , Ligação Proteica , Curva ROC , Coelhos , Ruptura Espontânea , Daboia , Fatores de Tempo , Rigidez Vascular , Vasodilatação , Venenos de VíborasRESUMO
BACKGROUND: Despite the beneficial effects of vascular interventions, these procedures may damage the endothelium leading to increased vascular permeability and remodeling. Re-endothelialization of the vessel wall, with functionally and structurally intact cells, is controlled by endothelial nitric oxide synthase (NOS3) and is crucial for attenuating adverse effects after injury. We investigated the applicability of the albumin-binding MR contrast agent, gadofosveset, to noninvasively monitor focal changes in vascular permeability and remodeling, after injury, in NOS3-knockout (NOS3(-/-)) and wild-type (WT) mice in vivo. METHODS AND RESULTS: WT and NOS3(-/-) mice were imaged at 7, 15, and 30 days after aortic denudation or sham-surgery. T1 mapping (R1=1/T1, s(-1)) and delayed-enhanced MRI were used as measurements of vascular permeability (R1) and remodeling (vessel wall enhancement, mm(2)) after gadofosveset injection, respectively. Denudation resulted in higher vascular permeability and vessel wall enhancement 7 days after injury in both strains compared with sham-operated animals. However, impaired re-endothelialization and increased neovascularization in NOS3(-/-) mice resulted in significantly higher R1 at 15 and 30 days post injury compared with WT mice that showed re-endothelialization and lack of neovascularization (R1 [s(-1)]=15 days: NOS3 (-/-)4.02 [interquartile range, IQR, 3.77-4.41] versus WT2.39 [IQR, 2.35-2.92]; 30 days: NOS3 (-/-)4.23 [IQR, 3.94-4.68] versus WT2.64 [IQR, 2.33-2.80]). Similarly, vessel wall enhancement was higher in NOS3(-/-) but recovered in WT mice (area [mm(2)]=15 days: NOS3 (-/-)5.20 [IQR, 4.68-6.80] versus WT2.13 [IQR, 0.97-3.31]; 30 days: NOS3 (-/-)7.35 [IQR, 5.66-8.61] versus WT1.60 [IQR, 1.40-3.18]). Ex vivo histological studies corroborated the MRI findings. CONCLUSIONS: We demonstrate that increased vascular permeability and remodeling, after injury, can be assessed noninvasively using an albumin-binding MR contrast agent and may be used as surrogate markers for evaluating the healing response of the vessel wall after injury.
Assuntos
Permeabilidade Capilar , Meios de Contraste , Endotélio Vascular/lesões , Gadolínio , Angiografia por Ressonância Magnética , Compostos Organometálicos , Remodelação Vascular , Animais , Permeabilidade Capilar/fisiologia , Endotélio Vascular/metabolismo , Humanos , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos , Camundongos Knockout , Modelos Animais , Óxido Nítrico Sintase Tipo III , Remodelação Vascular/fisiologia , Vasodilatação/fisiologiaRESUMO
BACKGROUND: Well-defined inflammation, proliferation, and maturation phases orchestrate the remodeling of the injured myocardium after myocardial infarction (MI) by controlling the formation of new extracellular matrix. The extracellular matrix consists mainly of collagen but also fractions of elastin. It is thought that elastin is responsible for maintaining elastic properties of the myocardium, thus reducing the risk of premature rupture. An elastin/tropoelastin-specific contrast agent (Gd-ESMA) was used to image tropoelastin and mature elastin fibers for in vivo assessment of extracellular matrix remodeling post-MI. METHODS AND RESULTS: Gd-ESMA enhancement was studied in a mouse model of myocardial infarction using a 7 T MRI scanner and results were compared to those achieved after injection of a nonspecific control contrast agent, gadolinium-diethylenetriamine pentaacetic acid (Gd-DTPA). In the infarcted tissue, Gd-ESMA uptake (measured as R1 relaxation rate) steadily increased from day 3 to day 21 as a result of the synthesis of elastin/tropoelastin. R1 values were in good agreement with histological findings. A similar R1 behavior was observed in the remote myocardium. No mature cross-linked elastin was found at any time point. In contrast, Gd-DTPA uptake was only observed in the infarct with no changes in R1 values between 3 and 21 days post-MI. CONCLUSIONS: We demonstrate the feasibility of in vivo imaging of extracellular matrix remodeling post-MI using a tropoelastin/elastin binding MR contrast agent, Gd-ESMA. We found that tropoelastin is the main contributor to the increased MRI signal at late stages of MI where its augmentation in areas of infarction was in good agreement with the R1 increase.
Assuntos
Meios de Contraste/farmacocinética , Elastina/metabolismo , Imagem Cinética por Ressonância Magnética , Infarto do Miocárdio/diagnóstico , Miocárdio/metabolismo , Miocárdio/patologia , Remodelação Ventricular , Animais , Biomarcadores/metabolismo , Meios de Contraste/administração & dosagem , Modelos Animais de Doenças , Estudos de Viabilidade , Feminino , Gadolínio DTPA/administração & dosagem , Injeções Intravenosas , Camundongos Endogâmicos C57BL , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/patologia , Valor Preditivo dos Testes , Ligação Proteica , Tropoelastina/metabolismoRESUMO
BACKGROUND AND PURPOSE: There are conflicting data regarding whether netrin-1 retards or accelerates atherosclerosis progression, as it can lead either to monocyte repulsion from or retention within plaques depending on its cellular source. We investigated the effect of aspirin, which is widely used in cardiovascular prophylaxis, on the synthesis of different isoforms of netrin-1 by endothelial cells under pro-inflammatory conditions, and defined the net effect of aspirin-dependent systemic modulation of netrin-1 on atherosclerosis progression. EXPERIMENTAL APPROACH: Netrin-1 synthesis was studied in vitro using human endothelial cells stimulated with TNF-α, with or without aspirin treatment. In vivo experiments were conducted in ApoE(-/-) mice fed with a high-fat diet (HFD), receiving either aspirin or clopidogrel. KEY RESULTS: TNF-α-induced NF-κB activation up-regulated the nuclear isoform of netrin-1, while simultaneously reducing secreted netrin-1. Down-regulation of the secreted isoform compromised the chemorepellent action of the endothelium against monocyte chemotaxis. Aspirin counteracted TNF-α-mediated effects on netrin-1 synthesis by endothelial cells through COX-dependent inhibition of NF-κB and concomitant histone hyperacetylation. Administration of aspirin to ApoE(-/-) mice on HFD increased blood and arterial wall levels of netrin-1 independently of its effects on platelets, accompanied by reduced plaque size and content of monocytes/macrophages, compared with untreated or clopidogrel-treated mice. In vivo blockade of netrin-1 enhanced monocyte plaque infiltration in aspirin-treated ApoE(-/-) mice. CONCLUSIONS AND IMPLICATIONS: Aspirin counteracts down-regulation of secreted netrin-1 induced by pro-inflammatory stimuli in endothelial cells. The aspirin-dependent increase of netrin-1 in ApoE(-/-) mice exerts anti-atherogenic effects by preventing arterial accumulation of monocytes.