This work was supported by the National Institutes of Health (HL114669). CM is a recipient of a postdoctoral research fellowship grant awarded by the Myocarditis Foundation, NJ.

ÉTICA DECLARAÇÃO: Todos os procedimentos com animais foram conduzidos de acordo com as diretrizes para o Cuidado e Uso de Animais de Laboratório e aprovado pela Universidade de Nebraska-Lincoln, Lincoln, NE. 1. Indução da miocardite auto-imune experimental <ol><li> Preparar a solução de péptidos por dissolução myhc-α 334-352 em solução salina 1x tamponada com fosfato até uma concentração final de 2 ml mg/1.5. </li><li> Prepare a toxina pertussis (PT) por adição de 1 ml de solução estéril de 1x PBS a um frasco contendo 50 ug de PT liofilizadas para se obter uma concentração de estoque de 50 ng / ul. Tomar 20 ul de estoque para um tubo esterilizado de 1,5 mL e adicionar 980 ul de 1x PBS estéril para obter uma concentração de trabalho de 1 ng / ul. </li><li> Prepare o adjuvante de Freund completo (CFA) pela adição de 40 mg de Mycobacterium tuberculosis H37Rv (MTB) extrair 10 ml de CFA para obter uma concentração finalde 5 mg / ml. </li><li> Preparar a emulsão de péptido-CFA. NOTA: Para a indução de EAM, a emulsão de péptido-CFA foi administrado em 150 ul contendo 100 ug de myhc-α 334-352 por animal. Por exemplo, para imunizar ratinhos dez utilizar 1,5 ml da emulsão de péptido-CFA contendo 1 mg de myhc-α 334-352. <ol><li> Para preparar 1,5 ml de emulsão, alíquota de 750 ul de myhc-α 334-352 solução de péptido em um tubo de 1,5 ml, e CFA suplementado com MTB num outro tubo de 1,5 ml. Utilizando uma seringa de 3 ml luer-lok, extrair a solução de péptido em primeiro lugar, seguido pelo extracto de CFA / MTB. </li><li> Fixe a seringa a uma torneira de 3 vias e ligue a outra saída da torneira a um vazio de 3 ml seringa. Ajuste a desobstrução da torneira para que a mistura de péptidos-CFA flui de uma seringa para o outro com razoavelmente boa resistência. </li><li> Misture, empurrando o conteúdo de uma seringa para o outro várias vezes para ~ 1 min e, em seguida, coloque todo o conjunto no gelo para a 3 min. Repita esteProcesso de um mínimo de 10x. </li><li> Determinar a consistência da emulsão, colocando cuidadosamente uma pequena gota em água parada em um copo de 100 ml de vidro. Não se espera que a gotícula de dispersar em água. Se isso acontecer, continue misturando até que a consistência desejada seja atingida. </li><li> Transferir o conteúdo do emulsão a partir de seringas de 3 ml em 1 ml seringas luer-lok de substituir uma das duas seringas de 3 ml ligados à torneira com a seringa de 1 ml, e anexar uma agulha 27 ½ G para a seringa de 1 ml. </li></ol></li><li> Injectar 150 ul da emulsão de péptido-CFA em doses divididas por via subcutânea nas duas regiões inguinal de fêmeas de seis a oito semanas de idade, ratinhos A / J (~ 75 mL cada). </li><li> Administrar 100 ul de suspensão de PT (100 ng) por via intraperitoneal a cada um dos animais no dia 0 e no dia 2 após a imunização. </li><li> Repetir o procedimento de imunização, no dia 7, com a administração de 150 uL de péptido-CFA emulsão em doses divididas por via subcutânea em ambos os lados de tele esterno (~ 75 mL cada). Preparar esta emulsão fresco tal como acima. Em seguida, no dia 21, submeter os animais a imagem MRM, consulte a etapa 3. </li></ol> 2. Handling Animais <ol><li> Coloque cada rato em uma câmara de indução anestésica contendo 2% de mistura de isoflurano ao ar com uma almofada de aquecimento colocada por baixo para manter o calor, e transferir o animal para um suporte de animais especialmente projetado (Figura 1). </li><li> Imobilizar o animal em decúbito ventral sobre o titular do animal para seu focinho se encaixa no cone do nariz para manter a anestesia (Figura 1). Proteja a cabeça do rato com uma mordida-bar fixados aos dentes da frente do mouse. </li><li> Ligue o aquecedor de sopro de ar com a sua mangueira de saída inserido no furo vertical do scanner para manter a temperatura do corpo do animal durante o experimento. </li><li> Manter a anestesia com 0,5 a 2% de isoflurano, com uma taxa de fluxo de 2 ml / min durante toda a sessão de imagem. Confirmaranestesia utilizando o método de aperto do dedo do pé, sem movimento esperado. </li><li> Configure um sensor pneumático travesseiro, rato cauda / tornozelo sensor de pulso de fibra óptica, oximetria e sonda de temperatura retal para monitorar a respiração, a freqüência cardíaca ea temperatura corporal, respectivamente (Figura 1). Nota: gating cardíaca é realizada através de oximetria de pulso, que permite o monitoramento não invasivo da saturação arterial de oxigênio no sangue. O sensor de oximetria de pulso deve ser anexado ao tornozelo esquerdo, eo pé devem ser fixados com um laço da linha e gravou para manter o tornozelo no sensor. Imagiologia MRM é alcançado pela passagem da respiração e os sinais cardíacos, sem o uso de quaisquer agentes de contraste. </li></ol> 3. Aquisição de Imagem <ol><li> Depois de preparar o animal (Figura 1), colocar o rato no centro do scanner MRM com o coração está posicionado no centro do campo de visão (FOV), onde a homogeneidade do campo magnético é máximo. NOTA: Uma grande calibre(89 mm) 9.4 T vertical magneto equipado com gradientes de triplo eixo de 100 g / cm e 4cm de rádio-freqüência (RF) bobina de imagem é para ser usado para obter imagens de alta resolução tridimensional (3D). Nota: Certifique-se de suprimentos não-magnéticos ao usar um scanner de ressonância magnética. </li><li> Execute a interface de imagem e selecione &quot;New Study&quot; do menu &quot;Opções de Estudo&quot;. Digite &quot;mtune&quot; na barra de comandos e executá-lo para puxar para cima o &quot;GUI Tune&quot;. Em seguida, selecione &quot;Start Probe Tune&quot; e clique no botão &quot;Escala automática&quot;. A GUI Tune será alterado para mostrar o RFsignal. Use os botões sintonizar / fósforo remotos no final da bobina para ajustar a bobina de RF com a frequência de ressonância do protão (400 MHz). Na guia &quot;Iniciar&quot; ir à página &quot;Prescan&quot; para executar a frequência e capacidade de calibração, clicando nos botões correspondentes. Aperte o botão XYZ (rápida) no &quot;guia&quot; de &quot;Shim Estudo &#39;guia para puxar para cima a página de calços.Vá para a página de correção, selecione todas as iterações, e aperte o botão de correção para realizar auto calços. </li><li> Selecione uma seqüência olheiro na guia &quot;Study&quot; da interface de imagem para localizar o coração mouse. Na guia &quot;Acquire&quot; mudar o FOV a 35 mm 2 e manter as configurações padrão da máquina. Clique em Iniciar para executar a seqüência; ajustar a posição do suporte do animal, se o coração não está no centro do campo de visão, sintonizar a bobina RF e obter as imagens escuteiro novamente. </li><li> Clique no botão seqüência &quot;GEMS&quot; na guia &quot;Estudo&quot;, em seguida, digite os parâmetros de aquisição na aba &quot;Acquire&quot; correspondentes para obter dois planos ortogonais ao longo do eixo curto e eixo longo do coração 20,21. NOTA: os parâmetros de aquisição típicos para uma varredura gradiente-eco são: espessura do corte, a 1 mm; tempo de repetição (TR), a 200 ms; tempo de eco (TE), 2,67 ms (mínimo); ângulo de inclinação de 25 °; inplane tamanho da matriz, de 128 x 128; FOV, 22 milímetros2; número de aquisições, 4; e um tempo de aquisição de cerca de 1 min e 30 seg. </li><li> Na aba &quot;Avançado&quot;, selecione a seqüência &quot;CINE&quot; para recolher os eixos curto cine imagens fechado-oximetria de pulso de RM, a fim de medir os parâmetros anatômicos e funcionais do VE. Ajuste a posição eo ângulo das fatias de imagem baseada na visão de longo eixo do coração por pairando do mouse e arrastando. Digite os seguintes parâmetros de imagem na guia &quot;Acquire&quot; para obter o gradiente echo seqüência cine: TR, a 500 ms; TE, 5 milissegundos; FOV de 22 mm 2; matriz de aquisição, de 256 x 256; espessura de corte, a 1 mm; número de aquisições, 8; número de quadros, 6; e um tempo de aquisição de ~ 17 min. Clique em Iniciar para iniciar a aquisição. </li><li> Converter as imagens adquiridas para o formato DICOM usando a guia &quot;I / O&quot; do software de imagens e transferir os arquivos correspondentes ao centro de dados para processamento. </li><li> No final do procedimento de imagiologia, todosow os ratos para se recuperar da anestesia dentro do filtro-top gaiolas. Não deixe os ratos autônoma até que recuperar a consciência suficiente para manter decúbito esternal e mantê-los para estudos posteriores, conforme necessário. </li></ol> 4. Análise de Dados de Ressonância Magnética Cardíaca Microscopia de imagem <ol><li> Use um software de Segmento de analisar os parâmetros anatômicos e funcionais do VE 19. Ao carregar as imagens no formato DICOM cine para o software usando o &quot;Arquivo de Imagem Aberto (s)&quot; submenu do menu &quot;Arquivo&quot;. Na GUI selecione &#39;MRGE &quot;, como a técnica de imagem,&#39; Cine &#39;, como o tipo de imagem e&#39; de eixo curto médio-ventricular&quot;, como o plano de exibição de imagem. </li><li> Especificar os prazos a serem utilizados para o final de diástole e final da sístole, através da &quot;Definir Prazo atual para Acabar com Diástole&quot; e &quot;Set Prazo atual para Acabar sístole&quot; submenus do menu &quot;Editar&quot;, respectivamente. </li><li>Primeiro, clique no botão de comando &quot;LV&quot; e depois o &quot;PAI&quot; botões de comando no painel inferior direito &quot;ENDO&quot; e desenhar manualmente o endocárdio do ventrículo esquerdo e epicárdio respectivamente. Retire os músculos papilares por apertar o botão de comando correspondente, a fim de aumentar a precisão dos cálculos. </li><li> Leia os parâmetros quantificados LV como volume diastólica, volume sistólico, do volume sistólico e fração de ejeção no painel superior direito. Clique no botão de comando &quot;Diversos&quot; e, em seguida, o botão de comando &quot;Medida Caliper&quot; para medir os parâmetros do LV, como espessura da parede e diâmetro ventricular 22. </li><li> Clique no botão &quot;Salvar Imagem Ambos Pilhas e Segmentação&quot; submenu do menu &quot;Arquivo&quot; para salvar as imagens, incluindo as segmentações, no formato &#39;. Mat&#39; para reprocessar as imagens, se necessário. </li></ol>

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No conflicts of interests were declared.

Este estudo descreve o procedimento MRM e sua utilidade como uma ferramenta não-invasiva para determinar anormalidades cardíacas em ratos afetados com miocardite auto-imune. Desde as características histológicas de EAM assemelham miocardite pós-infecciosa dos seres humanos, modelos de mouse são comumente empregados para delinear os mecanismos imunológicos de lesão miocárdica 23-25. No entanto, os animais afetados com miocardite aparecem clinicamente normal, eo diagnóstico é feito com base na histol...

A insuficiência cardíaca é a principal causa de mortes, e miocardite é uma das causas predominantes de insuficiência cardíaca em jovens adolescentes 1. A maioria dos pacientes afetados com miocardite permanecem assintomáticos e que a doença é resolvida espontaneamente 2. No entanto, 10-20% das pessoas afetadas podem desenvolver doença crônica, levando a cardiomiopatia dilatada (DCM) 3. Vários modelos animais têm sido desenvolvidos para estudar a patogénese imune miocardite. A doença pode ser induzida em miocardite suscetíveis A / J e ratinhos Balb / c através da imunização de animais com miosina cardíaca cadeia pesada (MyHC)-α ou os seus fragmentos de péptidos imunodominantes ou por infecção com agentes patogénicos como vírus coxsackie B3 4-9. Este estudo envolve myhc-α miocardite induzida-334-352 em ratinhos A / J. Apesar de mostrar infiltrações do miocárdio, os animais afetados por miocardite aparecem clinicamente normal; diagnóstico é baseado na avaliação histológica dos corações para a inflamação 7 aª ecocardiografia 10. Microscopia de ressonância magnética (MRM) é um método vulgarmente utilizado para obtenção de imagens de alta resolução cardiovascular com planos tridimensionais, permitindo que a avaliação de detalhes funcionais ao nível dos vasos sanguíneos pequenos (até 10 um de diâmetro), mas este nível de resolução de energia é não alcançável com a rotina de ressonância magnética (MRI) procedimento de digitalização, no qual, a resolução é geralmente obtida até 1 milímetro 11-14. MRM oferece uma vantagem, pois permite a aquisição de imagens de alta resolução e também para obter os parâmetros de desempenho nos momentos iniciais do processo da doença 14. Clinicamente, MRM imagem tem sido amplamente aplicado para estudar os parâmetros funcionais do doente coração, pulmão ou cérebro 15-17. Neste estudo, o uso de uma técnica de MRM como um meio não-invasivo para determinar anomalias cardíacas em ratinhos A / J afectados com miocardite auto-imune é mostrado. Especificamente, tele MRM imagem permite a quantificação dos parâmetros funcionais, tais como ventrículo esquerdo (VE) volume diastólico final e fração de ejeção (EF) com uma precisão razoável 18. As definições dos respectivos parâmetros são: o volume do ventrículo esquerdo que, o volume de sangue no ventrículo esquerdo no final do ciclo diastólica e fração de ejeção, o volume sistólico / volume diastólico final. A análise dos dados é realizada utilizando o software Segmento disponível gratuitamente desenvolvido para DICOM-compliant imagens cardiovasculares processamento adquiridos por scanners de ressonância magnética 19. Os dados revelaram um aumento na espessura da parede do VE nos animais myocarditic, correspondendo a uma diminuição do volume diastólico final do VE, do volume sistólico e fração de ejeção, em comparação com esses parâmetros funcionais em ratos saudáveis.

<table border="0" cellpadding="0" cellspacing="0" width="1185">
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			<td>List Biologicals Laboratories, Campbell, CA</td>
			<td>181</td>
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			<td>Corning, Manassas, VA</td>
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			<td>BD, Franklin Lakes, NJ</td>
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			<td>Energy Savers Unlimited, Inc. Carson, CA</td>
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			<td>Summit Anesthesia Solutions, Ann Harbor, MI</td>
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			<td>Agilent Technologies, Santa Clara, CA</td>
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noninvasive assessment of cardiac abnormalities in experimental autoimmune myocarditis by magnetic resonance microscopy imaging in the mouse

Miocardite é uma inflamação do miocárdio, mas apenas ~ 10% dessas manifestações clínicas mostram afetados da doença. Para estudar os eventos imunológicos das lesões do miocárdio, vários modelos de mouse de miocardite têm sido amplamente utilizados. Este estudo envolveu experimental miocardite auto-imune (EAM) induzida com miosina cardíaca cadeia pesada (MyHC)-α 334-352 em camundongos A / J; os animais afetados desenvolvem miocardite linfocítica mas sem sinais clínicos aparentes. Neste modelo, a utilidade de microscopia de ressonância magnética (MRM) como uma modalidade não invasivo para determinar as alterações estruturais e funcionais do coração em animais imunizados com myhc-α 334-352 é mostrada. EAM e ratos saudáveis ​​foram fotografadas usando um 9.4 T (400 MHz) 89 milímetros scanner de núcleo furo vertical, equipado com uma sonda de imagem de rádio-freqüência 4 centímetros centopéia e 100 g / cm gradientes eixos triplos. Imagens cardíacos foram adquiridos a partir de animais anestesiados utilizando uma seqüência de pulsos cine-based gradiente-eco, ea animals foram monitorados pela respiração e oximetria de pulso. A análise revelou um aumento na espessura da parede ventricular em murganhos EAM, com uma diminuição correspondente no diâmetro interior dos ventrículos, quando comparados com os ratos saudáveis. Os dados sugerem que as alterações morfológicas e funcionais nos corações inflamadas pode ser monitorizada de forma não invasiva por MRM em animais vivos. Em conclusão, MRM oferece uma vantagem de avaliar a progressão e a regressão de lesões do miocárdio em doenças causadas por agentes infecciosos, como resposta às terapias.

Neste relatório, a utilidade da técnica MRM como uma modalidade não-invasivo para determinar as mudanças estruturais e funcionais no coração dos animais afetados com EAM é mostrado. Miocardite foi induzida em ratinhos A / J por imunização dos animais com myhc-α 334-352 em CFA 7, e os animais foram submetidos a experiências MRM no dia 21 pós-imunização. A imagiologia foi realizada MRM em animais vivos, sob anestesia de isoflurano em 9,4 T (400 MHz para protões) utilizando um íman 89 milímetros...

Watch this Scientific Journal Video about Noninvasive Assessment of Cardiac Abnormalities in Experimental Autoimmune Myocarditis by Magnetic Resonance Microscopy Imaging in the Mouse at JoVE.com

Noninvasive Assessment of Cardiac Abnormalities in Experimental Autoimmune Myocarditis by Magnetic Resonance Microscopy Imaging in the Mouse

This study demonstrates the successful establishment of magnetic resonance microscopy imaging as a non-invasive tool to assess the cardiac abnormalities in mice affected with autoimmune myocarditis. The data indicate that the technique can be used to monitor the disease-progression in live animals.

Avaliação não invasiva da Cardiac Anormalidades em Experimental Auto-imune Miocardite por Ressonância Magnética Microscopia de imagem no Rato

Myocarditis is an inflammation of the myocardium, but only ~10% of those affected show clinical manifestations of the disease. To study the immune events ...

noninvasive-assessment-cardiac-abnormalities-experimental-autoimmune

Research

JoVE Journal

Medicine

9.8K visualizações.  University of Nebraska-Lincoln. This study demonstrates the successful establishment of magnetic resonance microscopy imaging as a non-invasive tool to assess the cardiac abnormalities in mice affected with autoimmune myocarditis. The data indicate that the technique can be used to monitor the disease-progression in live animals.

Vídeo: Noninvasive Assessment of Cardiac Abnormalities in Experimental Autoimmune Myocarditis by Magnetic Resonance Microscopy Imaging in the Mouse

Magnetic Resonance Derived Myocardial Strain Assessment Using Feature Tracking

Purpose: An accurate and practical method to measure parameters like strain in myocardial tissue is of great clinical value, since it has been shown, that strain is a more sensitive and earlier marker for contractile dysfunction than the frequently used parameter EF. Current technologies for CMR are time consuming and difficult to implement in clinical practice. Feature tracking is a technology that can lead to more automization and robustness of quantitative analysis of medical images with less time consumption than comparable methods.
Methods: An automatic or manual input in a single phase serves as an initialization from which the system starts to track the displacement of individual patterns representing anatomical structures over time. The specialty of this method is that the images do not need to be manipulated in any way beforehand like e.g. tagging of CMR images.
Results: The method is very well suited for tracking muscular tissue and with this allowing quantitative elaboration of myocardium and also blood flow.
Conclusions: This new method offers a robust and time saving procedure to quantify myocardial tissue and blood with displacement, velocity and deformation parameters on regular sequences of CMR imaging. It therefore can be implemented in clinical practice.

An accurate and practical method to measure parameters like strain in myocardial tissue is of great clinical value, since it has been shown, that strain is a more sensitive and earlier marker for contractile dysfunction than the frequently used parameter EF.

Avaliação Strain Ressonância Magnética Derivado do Miocárdio Usando o controle de recursos

Purpose: An accurate and practical method to measure parameters like strain in myocardial tissue is of great clinical value, since it has been shown, that ...

Medicina

Multi-modal Imaging of Angiogenesis in a Nude Rat Model of Breast Cancer Bone Metastasis Using Magnetic Resonance Imaging, Volumetric Computed Tomography and Ultrasound

Angiogenesis is an essential feature of cancer growth and metastasis formation. In bone metastasis, angiogenic factors are pivotal for tumor cell proliferation in the bone marrow cavity as well as for interaction of tumor and bone cells resulting in local bone destruction. Our aim was to develop a model of experimental bone metastasis that allows in vivo assessment of angiogenesis in skeletal lesions using non-invasive imaging techniques.
For this purpose, we injected 105 MDA-MB-231 human breast cancer cells into the superficial epigastric artery, which precludes the growth of metastases in body areas other than the respective hind leg1. Following 25-30 days after tumor cell inoculation, site-specific bone metastases develop, restricted to the distal femur, proximal tibia and proximal fibula1. Morphological and functional aspects of angiogenesis can be investigated longitudinally in bone metastases using magnetic resonance imaging (MRI), volumetric computed tomography (VCT) and ultrasound (US).
MRI displays morphologic information on the soft tissue part of bone metastases that is initially confined to the bone marrow cavity and subsequently exceeds cortical bone while progressing. Using dynamic contrast-enhanced MRI (DCE-MRI) functional data including regional blood volume, perfusion and vessel permeability can be obtained and quantified2-4. Bone destruction is captured in high resolution using morphological VCT imaging. Complementary to MRI findings, osteolytic lesions can be located adjacent to sites of intramedullary tumor growth. After contrast agent application, VCT angiography reveals the macrovessel architecture in bone metastases in high resolution, and DCE-VCT enables insight in the microcirculation of these lesions5,6. US is applicable to assess morphological and functional features from skeletal lesions due to local osteolysis of cortical bone. Using B-mode and Doppler techniques, structure and perfusion of the soft tissue metastases can be evaluated, respectively. DCE-US allows for real-time imaging of vascularization in bone metastases after injection of microbubbles7.
In conclusion, in a model of site-specific breast cancer bone metastases multi-modal imaging techniques including MRI, VCT and US offer complementary information on morphology and functional parameters of angiogenesis in these skeletal lesions.

In the pathogenesis of bone metastasis, angiogenesis is a crucial process and therefore represents a target for imaging and therapy. Here, we present a rat model of site-specific breast cancer bone metastasis and describe strategies to non-invasively image angiogenesis in vivo using magnetic resonance imaging, volumetric computed tomography and ultrasound.

Multi-modal de imagem de angiogênese em um modelo de rato nu de Câncer de mama utilizando a ressonância magnética, tomografia computadorizada volumétrica e ultra-som

Angiogenesis is an essential feature of cancer growth and metastasis formation. In bone metastasis, angiogenic factors are pivotal for tumor cell ...

Diffusion Tensor Magnetic Resonance Imaging in the Analysis of Neurodegenerative Diseases

Diffusion tensor imaging (DTI) techniques provide information on the microstructural processes of the cerebral white matter (WM) in vivo. The present applications are designed to investigate differences of WM involvement patterns in different brain diseases, especially neurodegenerative disorders, by use of different DTI analyses in comparison with matched controls. 	
DTI data analysis is performed in a variate fashion, i.e. voxelwise comparison of regional diffusion direction-based metrics such as fractional anisotropy (FA), together with fiber tracking (FT) accompanied by tractwise fractional anisotropy statistics (TFAS) at the group level in order to identify differences in FA along WM structures, aiming at the definition of regional patterns of WM alterations at the group level. Transformation into a stereotaxic standard space is a prerequisite for group studies and requires thorough data processing to preserve directional inter-dependencies. The present applications show optimized technical approaches for this preservation of quantitative and directional information during spatial normalization in data analyses at the group level. On this basis, FT techniques can be applied to group averaged data in order to quantify metrics information as defined by FT. Additionally, application of DTI methods, i.e. differences in FA-maps after stereotaxic alignment, in a longitudinal analysis at an individual subject basis reveal information about the progression of neurological disorders. Further quality improvement of DTI based results can be obtained during preprocessing by application of a controlled elimination of gradient directions with high noise levels.	
In summary, DTI is used to define a distinct WM pathoanatomy of different brain diseases by the combination of whole brain-based and tract-based DTI analysis.

Diffusion tensor imaging (DTI) basically serves as an MRI-based tool to identify in vivo the microstructure of the brain and pathological processes due to neurological disorders within the cerebral white matter. DTI-based analyses allow for application to brain diseases both at the group level and in single subject data.

Diffusion Tensor Imagem por Ressonância Magnética na Análise de Doenças Neurodegenerativas

Diffusion tensor imaging (DTI) techniques provide information on the microstructural processes of the cerebral white matter (WM) in vivo. The present ...

Implanting Glass Spinal Cord Windows in Adult Mice with Experimental Autoimmune Encephalomyelitis

Experimental autoimmune encephalomyelitis (EAE) in adult rodents is the standard experimental model for studying autonomic demyelinating diseases such as multiple sclerosis. Here we present a low-cost and reproducible glass window implantation protocol that is suitable for intravital microscopy and studying the dynamics of spinal cord cytoarchitecture with subcellular resolution in live adult mice with EAE. Briefly, we surgically expose the vertebrae T12-L2 and construct a chamber around the exposed vertebrae using a combination of cyanoacrylate and dental cement. A laminectomy is performed from T13 to L1, and a thin layer of transparent silicone elastomer is applied to the dorsal surface of the exposed spinal cord. A modified glass cover slip is implanted over the exposed spinal cord taking care that the glass does not directly contact the spinal cord. To reduce the infiltration of inflammatory cells between the window and spinal cord, anti-inflammatory treatment is administered every 2 days (as recommended by ethics committee) for the first 10 days after implantation. EAE is induced only 2-3 weeks after the cessation of anti-inflammatory treatment.
Using this approach we successfully induced EAE in 87% of animals with implanted windows and, using Thy1-CFP-23 mice (blue axons in dorsal spinal cord), quantified axonal loss throughout EAE progression. Taken together, this protocol may be useful for studying the recruitment of various cell populations as well as their interaction dynamics, with subcellular resolution and for extended periods of time. This intravital imaging modality represents a valuable tool for developing therapeutic strategies to treat autoimmune demyelinating diseases such as EAE.

We describe a method for implanting and maintaining glass windows over the exposed spinal cords of adult mice for studying experimental autoimmune encephalomyelitis. These windows provide chronic optical access to the spinal cord for monitoring cell dynamics at the subcellular level in live animals using two-photon microscopy.

Implantação de vidro Medular do Windows em camundongos adultos com Experimental Auto-imune Encephalomyelitis

Experimental autoimmune encephalomyelitis (EAE) in adult rodents is the standard experimental model for studying autonomic demyelinating diseases such as ...

Simultaneous PET/MRI Imaging During Mouse Cerebral Hypoxia-ischemia

Dynamic changes in tissue water diffusion and glucose metabolism occur during and after hypoxia in cerebral hypoxia-ischemia reflecting a bioenergetics disturbance in affected cells. Diffusion weighted magnetic resonance imaging (MRI) identifies regions that are damaged, potentially irreversibly, by hypoxia-ischemia. Alterations in glucose utilization in the affected tissue may be detectable by positron emission tomography (PET) imaging of 2-deoxy-2-(18F)fluoro-&#7429;-glucose ([18F]FDG) uptake. Due to the rapid and variable nature of injury in this animal model, acquisition of both modes of data must be performed simultaneously in order to meaningfully correlate PET and MRI data. In addition, inter-animal variability in the hypoxic-ischemic injury due to vascular differences limits the ability to analyze multi-modal data and observe changes to a group-wise approach if data is not acquired simultaneously in individual subjects. The method presented here allows one to acquire both diffusion-weighted MRI and [18F]FDG uptake data in the same animal before, during, and after the hypoxic challenge in order to interrogate immediate physiological changes.

The method presented here uses simultaneous positron emission tomography and magnetic resonance imaging. In the cerebral hypoxia-ischemia model, dynamic changes in diffusion and glucose metabolism occur during and after injury. The evolving and irreproducible damage in this model necessitates simultaneous acquisition if meaningful multi-modal imaging data are to be acquired.

Simultânea de imagem PET / MRI Durante Rato isquemia-hipoxia cerebral

Dynamic changes in tissue water diffusion and glucose metabolism occur during and after hypoxia in cerebral hypoxia-ischemia reflecting a bioenergetics ...

Assessment of Cardiac Morphological and Functional Changes in Mouse Model of Transverse Aortic Constriction by Echocardiographic Imaging

Transverse aortic constriction (TAC) in mice has been used as a valuable model to study mechanisms of cardiac hypertrophy and heart failure1. A reliable noninvasive method is essential to assess real-time cardiac morphological and functional changes in animal models of heart disease. Transthoracic echocardiography represents an important tool for noninvasive assessment of cardiac structure and function2. Here we used a high-resolution ultrasound imaging system to monitor myocardial remodeling and heart failure progression over time in a mouse model of TAC. B-mode, M-mode, and Doppler imaging were used to precisely assess cardiac hypertrophy, ventricular dilatation, and functional deterioration in mice following TAC. Color and pulse wave (PW) Doppler imaging was used to noninvasively measure pressure gradient across the aortic constriction created by TAC and to assess transmitral blood flow in mice. Thus transthoracic echocardiographic imaging provides comprehensive noninvasive measurements of cardiac dimensions and function in mouse models of heart disease.

The goal of this protocol is to noninvasively assess cardiac structural and functional changes in a mouse model of heart disease created by transverse aortic constriction, using B- and M-mode echocardiography and color/pulse wave Doppler imaging.

Avaliação da morfológicas e funcionais cardíacas Mudanças no Modelo rato de Transverse aórtica constrição por ecocardiográfica de imagem

Transverse aortic constriction (TAC) in mice has been used as a valuable model to study mechanisms of cardiac hypertrophy and heart failure1. A reliable ...

Quantitative Visualization of Leukocyte Infiltrate in a Murine Model of Fulminant Myocarditis by Light Sheet Microscopy

Light-sheet fluorescence microscopy (LSFM), in combination with chemical clearing protocols, has become the gold standard for analyzing fluorescently labelled structures in large biological specimens, and is down to cellular resolution. Meanwhile, the constant refinement of underlying protocols and the enhanced availability of specialized commercial systems enable us to investigate the microstructure of whole mouse organs and even allow for the characterization of cellular behavior in various live-cell imaging approaches. Here, we describe a protocol for the spatial whole-mount visualization and quantification of the CD45+ leukocyte population in inflamed mouse hearts. The method employs a transgenic mouse strain (CD11c.DTR)that has recently been shown to serve as a robust, inducible model for the study of the development of fulminant fatal myocarditis, characterized by lethal cardiac arrhythmias. This protocol includes myocarditis induction, intravital antibody-mediated cell staining, organ preparation, and LSFM with subsequent computer-assisted image post-processing. Although presented as a highly-adapted method for our particular scientific question, the protocol represents the blueprint of an easily adjustable system that can also target completely different fluorescent structures in other organs and even in other species.

Here, we describe a light-sheet microscopy approach to visualize the cardiac CD45+ leukocyte infiltrate in a murine model of aseptic fulminant myocarditis, which is induced by the intratracheal diphtheria toxin treatment of CD11c.DTR mice.

Visualização Quantitativa de Infiltrado de Leucócitos em um Modelo Murino de Miocardite Fulminante por Microscopia de Folha de Luz

Light-sheet fluorescence microscopy (LSFM), in combination with chemical clearing protocols, has become the gold standard for analyzing fluorescently ...

Cardiac Magnetic Resonance Imaging at 7 Tesla

CMR at an ultra-high field (magnetic field strength B0&#160;&#8805; 7 Tesla) benefits from the signal-to-noise ratio (SNR) advantage inherent at higher magnetic field strengths and potentially provides improved signal contrast and spatial resolution. While promising results have been achieved, ultra-high field CMR is challenging due to energy deposition constraints and physical phenomena such as transmission field non-uniformities and magnetic field inhomogeneities. In addition, the magneto-hydrodynamic effect renders the synchronization of the data acquisition with the cardiac motion difficult. The challenges are currently addressed by explorations into novel magnetic resonance technology. If all impediments can be overcome, ultra-high field CMR may generate new opportunities for functional CMR, myocardial tissue characterization, microstructure imaging or metabolic imaging. Recognizing this potential, we show that multi-channel radio frequency (RF) coil technology tailored for CMR at 7 Tesla together with higher order B0 shimming and a backup signal for cardiac triggering facilitates high fidelity functional CMR. With the proposed setup, cardiac chamber quantification can be accomplished in examination times similar to those achieved at lower field strengths. To share this experience and to support the dissemination of this expertise, this work describes our setup and protocol tailored for functional CMR at 7 Tesla.

The sensitivity gain inherent to ultrahigh field magnetic resonance holds promise for high spatial resolution imaging of the heart. Here, we describe a protocol customized for functional cardiovascular magnetic resonance (CMR) at 7 Tesla using an advanced multi-channel radio-frequency coil, magnetic field shimming and a triggering concept.

Ressonância magnética cardíaca na Tesla 7

CMR at an ultra-high field (magnetic field strength B0&#160;&#8805; 7 Tesla) benefits from the signal-to-noise ratio (SNR) advantage inherent at higher ...

Phase Contrast Magnetic Resonance Imaging in the Rat Common Carotid Artery

Phase contrast magnetic resonance imaging (PC-MRI) is a noninvasive approach that can quantify flow-related parameters such as blood flow. Previous studies have shown that abnormal blood flow may be associated with systemic vascular risk. Thus, PC-MRI can facilitate the translation of data obtained from animal models of cardiovascular diseases to pertinent clinical investigations. In this report, we describe the procedure for measuring blood flow in the common carotid artery (CCA) of rats using cine-gated PC-MRI and discuss relevant analysis methods. This procedure can be performed in a live, anesthetized animal and does not require euthanasia after the procedure. The proposed scanning parameters yield repeatable measurements for blood flow, indicating excellent reproducibility of the results. The PC-MRI procedure described in this article can be used for pharmacological testing, pathophysiological assessment, and cerebral hemodynamics evaluation.

The overall goal of this procedure is to measure the blood flow in the rat common carotid artery by using noninvasive phase contrast magnetic resonance imaging.

Imagens de ressonância magnética fase contraste na artéria carótida comum de rato

Phase contrast magnetic resonance imaging (PC-MRI) is a noninvasive approach that can quantify flow-related parameters such as blood flow. Previous ...

Quantification of Levator Ani Hiatus Enlargement by Magnetic Resonance Imaging in Males and Females with Pelvic Organ Prolapse

Here we present a protocol to examine the levator ani hiatus in males and females with pelvic organ prolapse, during the Valsalva maneuver and while evacuating acoustic gel, using a horizontally oriented 1.5 T magnetic resonance (MR) scanner. On midsagittal images, the vertical distance of pelvic organs is measured in millimeters relative to the hymen plane (female) and to the lower border of the symphysis pubis (male), preceded by - (above) or + ( below) signs. On axial images, the levator ani area is calculated in square centimeters with a free-hand tracing method from three key images, passing through the midsymphysis (level I), tangential to the lower border of the symphysis (level II), and at the maximal anterior rectal wall bulging (level III). Areas at rest and strained are compared to find evidence of a percentage of increase. The purpose is to provide objective evidence of the maximal extent of pelvic organs descent and hiatus enlargement without the interference of foreign objects or the examiner&#39;s proximity, so as to overcome the limitations of pelvic examination and transperineal sonography (i.e., subjectivity and sex-related limitations [female only]).

Here we present a protocol to standardize the measurement of the levator ani hiatus size&#160;by magnetic resonance imaging. The purpose is to extract biomechanical inferences from image analysis by comparing resting and straining values in patients of both sexes with pelvic prolapse, using consistent anatomical bony landmarks.

Quantificação do Levator Ani hiato alargamento por ressonância magnética em machos e fêmeas com prolapso de órgãos pélvicos

Here we present a protocol to examine the levator ani hiatus in males and females with pelvic organ prolapse, during the Valsalva maneuver and while ...