Agradecemos a los Dres. Raimond Winslow, Elliot McVeigh, y Helm Patrick de la Johns Hopkins University de proporcionar conjuntos de datos ex vivo la investigación online.This fue apoyada por los Institutos Nacionales de Salud de subvención R01-HL082729 y National Science Foundation de subvención CBET-0.933.029.

1. Orientaciones de fibra Estimación <ol><li> Adquirir imágenes de MRI estructurales y DTMRI de un corazón humano adulto normal en diástole, con una resolución de 1 mm 3. Usando ImageJ, extraer el miocardio ventricular de la imagen estructural atlas por ajuste, para cada rebanada de eje corto, cerrado splines a través de un conjunto de puntos de señal colocados a lo largo de los límites epicárdicos y endocárdicos en la rebanada (Figura 2A y la figura 2B). Realizar la colocación de puntos de señal manualmente para cada rebanada 10 ª en la imagen. Obtener los puntos de señal para las rebanadas restantes al interpolar linealmente los puntos manualmente identificados, usando MATLAB.Reconstruct las orientaciones de las fibras del corazón atlas mediante el cálculo de los vectores propios primarias de los DTS en la imagen DTMRI (Figura 2C). </li><li> Adquirir una imagen de la geometría del corazón del paciente en diástole utilizando in vivo cardíaco CT o MRI. Reconstruct la geometría del corazón del paciente de la imagen de manera similar a la forma en que se construyó el atlas (Figura 3A y la Figura 3B). La imagen de paciente debe volver a tomar muestras antes de la reconstrucción de tal manera que la resolución en el plano es de 1 mm 2. De manera similar, el número de cortes para que puntos de referencia son recogidos manualmente, y el intervalo de interpolación fuera de plano debe ajustarse de modo que la imagen segmentada paciente corazón tiene un grosor de corte de 1 mm. </li><li> Deformar la imagen ventricular atlas para que coincida con la imagen geometría paciente en dos pasos. En el primer paso, realizar una transformación afín sobre la base de un conjunto de puntos de puntos de referencia de trece: el ventrículo izquierdo (LV) ápice, el ventrículo dos a la derecha (RV) puntos de inserción en la base, los dos puntos de inserción de RV a mitad de camino entre la base y el ápice, y cuatro conjuntos de dos puntos que dividen uniformemente RV LV epicardio y los contornos en la base, ya mitad de camino entre la base y el ápice (Figura 4A y <strong&gt; Figura 4B). En el segundo paso, deformar la transformada afín-atlas ventrículos aún más para que coincida con la geometría de la paciente, utilizando la correlación de gran difeomorfa deformación métrico (LDDMM) (Figura 4C). </li><li> Morph la imagen DTMRI del atlas por el cambio de posición de voxels de imagen y reorientando los DTS de acuerdo con la matriz de transformación de la correspondencia afín y el campo de deformación de la transformación LDDMM. Realizar la re-orientación de los DTs con la preservación de las direcciones principales (PPD) método. </li><li> Obtener la estimación de las orientaciones de las fibras de pacientes de la transformada imagen atlas DTMRI calculando el vector propio principal de la DT (Figura 5). </li></ol> 2. Medición del error de estimación <ol><li> Adquirir ex vivo estructural y MR imágenes DTMR de seis normales y tres corazones insuficientes caninos, a una resolución de 312,5 x 312,5 x 800 m 3. Aquí, el corazón faiseñuelo se debe generar en los caninos a través de la ablación por radiofrecuencia de la rama izquierda seguido de 3 semanas de tachypacing a 210 min -1. </li><li> Segmento de los ventrículos del corazón canino de manera similar al corazón humano, atlas, tal como se describe en el § 1.1. Denotar ventrículos segmentados de corazones caninos normales como corazones 1 a 6, y los de no segmentado corazones caninos como corazones 7 a 9 (Figura 6). </li><li> Obtener cinco diferentes estimaciones de orientaciones de las fibras ventriculares del corazón 1 mediante el uso de cada uno de los corazones de 2 a 6 como un atlas (Figura 7). </li><li> Estimar orientaciones de las fibras para cada uno de los ventrículos del corazón fallan al usar 1 como el atlas (Figura 8). </li><li> Foreach punto de datos de cada conjunto de orientaciones de fibra estimados, calcular el error de estimación como | θ c-θ a |, donde θ θ c y A son los ángulos de inclinación de un estimadod adquirido orientaciones de las fibras en ese punto, respectivamente. </li><li> Para cada punto de datos en cada conjunto de orientaciones de fibra estimados, calcular el ángulo agudo betweenestimated y adquirió direcciones de las fibras en tres dimensiones (3D) por medio de thevector producto escalar. </li></ol> 3. La medición de los efectos del error de estimación en las simulaciones <ol><li> De corazón 1, la construcción de seis modelos, uno con las orientaciones de fibra DTMRI-adquiridas de corazón 1 (denominado modelo 1), y cinco con las cinco orientaciones de fibra estimados conjuntos de datos (modelos de 2 a 6). Para cada uno de el corazón tres defecto geometrías, construir dos modelos ventriculares, uno con los DTMRI adquiridas en orientaciones de fibra y el otro con las orientaciones de fibra estimados. Aquí, la resolución espacial de los modelos, calculado en términos de la longitud de borde media de las mallas, debe ser de aproximadamente 600 micras. Denotemos los modelos de insuficiencia cardiaca con fibras DTMRI adquiridas como modelos de 7 a 9, y los que tienen estifibras ed como modelos de 10 a 12.In los modelos, representación monodominio utilizar para describir el tejido cardíaco, con las ecuaciones que rigen: </li></ol><img src="/files/ftp_upload/50125/50125eq1.jpg" alt="Ecuación 1" fo:content-width="2.18in" fo:src="/files/ftp_upload/50125/50125eq1highres.jpg" /> donde σ b es el tensor de conductividad a granel que se calcula a partir de los tensores de conductividad bidomain como se describe por Potse et al 30; V m es el potencial transmembrana; m C es la capacitancia de membrana específico, y de iones I es la densidad de la corriente transmembrana, que a su vez depende de V m y un conjunto de variables de estado μ describir la dinámica de los flujos iónicos a través de la m membrane.For C, utilizar un valor de 1 μ F / cm 2. Para i σ en los modelos normales del corazón canino, utilice longitudinalnales y transversales valores de conductividad de 0,34 S / m y 0,06 S / m, respectivamente. Representar lon l en los modelos Greenstein Winslow-iónicos del miocito ventricular canino. Disminuir las conductividades eléctricas en modelos caninos cardíacos ventriculares falla en un 30% (Figura 9). <ol start="2"><li> Usando el paquete de software CARP (CardioSolv, LLC), el ritmo sinusal simular con todos los modelos. Inducir taquicardia ventricular reentrante (VT) en los seis modelos fallan al usar un protocolo de estimulación S1-S2. Elige la sincronización entre S1 y S2 para obtener actividad TV sostenida durante 2 segundos después de la entrega S2. Si VT no se induce para cualquier sincronización S1-S2, disminuir las conductividades de hasta un 70% hasta VT fue inducida (Figura 10). </li><li> Para cada simulación, calcular pseudo-ECGs tomando la diferencia de los potenciales extracelulares entre dos puntos en un baño de isotrópica que rodea el corazón. Coloque los dos puntos cerca de la base del corazón separadospor 18 cm, de tal manera que la línea que conecta ellos es perpendicular al plano de base-vértice del septo como se ilustra en la Figura 10. Para cada simulación con orientaciones de fibra estimados, calcular la métrica como MAD </li></ol><img src="/files/ftp_upload/50125/50125eq2.jpg" alt="Ecuación 2" fo:content-width="2.77in" fo:src="/files/ftp_upload/50125/50125eq2highres.jpg" /> donde X es la forma de onda de ECG de la simulación con orientaciones de fibra estimados, Y es la forma de onda de ECG de thecorresponding simulación con orientaciones de fibra adquiridos, X es el valor medio de X, Y es el valor medio de Y, y n es la longitud de X e Y.

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No hay conflictos de interés declarado.

Esta investigación demuestra cuantitativamente que, en ausencia de DTMRI, las orientaciones de fibra de miocardio ventrículos normales y no puede ser estimado a partir de imágenes in vivo de sus geometrías para uso en las simulaciones de la electrofisiología cardiaca. La metodología propuesta se demuestra con datos de la TC en vivo, pero es igualmente aplicable a las imágenes de RM in vivo de la geometría ventricular, frente a la falta de capacidad para adquirir directamente del pacien...

<html> El enfoque computacional se está convirtiendo central para el avance de la comprensión de la función del corazón en la salud y la enfermedad. State-of-the-art de todo corazón-modelos de electrofisiología y electromecánica se están utilizando para estudiar una amplia gama de fenómenos, tales como la propagación ventricular normal, la arritmia, la desfibrilación, el acoplamiento electromecánico, y la resincronización cardiaca 1. Sin embargo, para el enfoque computacional para ser directamente aplicables en el entorno clínico, es imperativo que los modelos de ser específica para el paciente, es decir, los modelos debe estar basado en la arquitectura específica y propiedades electrofisiológicas o electromecánicas de corazón enfermo del paciente. Simulación con estos modelos ayudarán a los médicos a llegar a decisiones altamente personalizadas para las intervenciones electrofisiológicos, así como la profilaxis, así dramáticamente mejorar la atención de la salud cardiaca 2-4. contenido &quot;&gt; Creación de modelos realistas cardíacos requiere la adquisición de la geometría y la estructura de fibra de un paciente cardíaco. orientaciones de fibra determinar direcciones de propagación eléctrica y distribuciones de tensión en el corazón, y por lo tanto la adquisición de ellos es esencial para el modelado cardíaco 5, 6. Con los recientes avances en imagen médica, es ahora posible obtener la geometría de un corazón paciente, remodelación includingstructural tales como el infarto, in vivo con alta resolución usando imágenes por resonancia magnética (MRI) y tomografía computarizada tecnologías (CT). Sin embargo, no hay método práctico para la adquisición de estructura de la fibra de un corazón paciente in vivo. tensor de difusión (DT) RM 7, 8, la única técnica para adquirir orientaciones de las fibras del corazón intacto, no está ampliamente disponible en vivo debido a ciertas limitaciones 9. Una breve descripción de los esfuerzos anteriores para traducir DTMRI para el entorno clínico se pueden encontrar elsewhere 2. Aunque metodologías como regla basada en la asignación de las orientaciones de fibra de ofrecer alternativas a DTMRI, estos métodos tienen ciertas limitaciones graves 2, 10. Así, las dificultades en la adquisición de estructura de la fibra cardiaca in vivo en la actualidad impiden la aplicación de simulaciones cardíacos electrofisiológicos y electromecánicos en la clínica. El objetivo de esta investigación fue abordar directamente esta necesidad. La hipótesis de que las orientaciones de fibra ventricular de un corazón puede predecirse con precisión debido a la geometría del corazón y de un atlas, donde el atlas es un corazón cuya geometría y orientaciones de fibra están disponibles. En consecuencia, se utilizó el estado de la técnica para desarrollar una metodología para la estimación de las orientaciones de las fibras cardíacas en vivo, y probaron la hipótesis de normalidad y de no ventrículos caninos 2. La idea central de nuestra metodología de estimación de fibra es explotar similarities en orientaciones de las fibras, con respecto a la geometría, entre corazones diferentes con el fin de aproximarse a la estructura de la fibra de un corazón (objetivo) para que sólo la información de geometría se encuentra disponible. En el corazón de nuestra metodología de estimación es el registro de la geometría atlas con la geometría de destino mediante mapas difeomorfa gran deformación métrica (LDDMM) 11, y la transformación de las orientaciones de fibra atlas con la preservación de los componentes principales (PPD) 2, 12. El diffeomorphicproperty de garantías LDDMM que el atlas no &quot;plegable&quot; sí durante la deformación, preservando así las estructuras anatómicas integrityof. Figura 1 ilustra la canalización de procesamiento de nuestra metodología. La sección del texto protocolo § 1 describe los diversos componentes de la tubería mediante la demostración de cómo la estimación se puede realizar para un paciente ejemplo. Los números dentro de algunos de los bloques en la Figura 1 se refieren a la correspondientesubsecciones en la sección § 1 del texto del protocolo. Se evaluó el desempeño de la metodología propuesta por la cuantificación del error de estimación, y medir el efecto de esta onsimulations de error de la electrofisiología cardiaca, por computacionalmente simulando locales mapas de activación eléctricas, así como pseudo-electrocardiogramas (ECGs pseudo-). Debido a la falta de disponibilidad de los corazones humanos, la evaluación del desempeño se realizó con corazones caninos disponibles a partir de estudios previos 13-15. El error de estimación se calcula por medio de ángulos de inclinación 16, la tradición followingthe de histología, donde las mediciones angulares son secciones performedon tejido que son cortadas en paralelo a la epicardialsurface. Desde el anglebetween la dirección de la fibra y el plano tangente epicárdica es generallysmall 17, 18, ​​la pérdida de información en la descripción de un fiberdirection enteramente usando su ángulo de inclinación es insignificante. Para la computsimulaciones acionales, basados ​​en imágenes modelos se construyeron como se informó anteriormente 19, 20, y el tejido cardíaco en los modelos fue representado establecidos sobre la base de técnicas matemáticas y los datos experimentales 21-25. El ritmo sinusal se simuló por la activación de replicación procedente de la red de Purkinje 26, y la taquicardia ventricular, por una estimulación S1-S2 protocolo 27. Los pseudo-ECG se calcularon 28 y compararon mediante la desviación media absoluta (MAD) métrica 29.

<table border="1"><tbody><tr><td> LDDMM </td><td> Johns Hopkins University </td><td></td><td> <a href="http://cis.jhu.edu/software/lddmm-volume/index.php" target="_blank">http://cis.jhu.edu/software/lddmm-volume/index.php</a> </td></tr><tr><td> MATLAB </td><td> Mathworks, Inc. </td><td> R2011b </td><td> <a href="http://www.mathworks.com/products/matlab/" target="_blank">http://www.mathworks.com/products/matlab/</a> </td></tr><tr><td> ImageJ </td><td> Los Institutos Nacionales de la Salud </td><td></td><td> <a href="http://rsbweb.nih.gov/ij/" target="_blank">http://rsbweb.nih.gov/ij/</a> </td></tr><tr><td> Tarántula </td><td> CAE Software Solutions </td><td></td><td> <a href="http://www.meshing.at/Spiderhome/Tarantula.html" target="_blank">http://www.meshing.at/Spiderhome/Tarantula.html</a> </td></tr><tr><td> CARP </td><td> CardioSolv</td><td></td><td> <a href="http://cardiosolv.com/" target="_blank">http://cardiosolv.com/</a> </td></tr><tr><td> Imágenes caninas </td><td> Johns Hopkins University </td><td></td><td> <a href="http://www.ccbm.jhu.edu/research/DTMRIDS.php" target="_blank">http://www.ccbm.jhu.edu/research/DTMRIDS.php</a> </td></tr></tbody></table>

patient-specific modeling of the heart: estimation of ventricular fiber orientations

Específicos del paciente simulaciones de corazón (dis) función dirigida a la personalización de la terapia cardiaca se ve obstaculizada por la ausencia de la tecnología de formación de imágenes in vivo para clínicamente adquirir orientaciones de fibra de miocardio. El objetivo de este proyecto fue desarrollar una metodología para estimar las orientaciones de fibras cardíacas de imágenes in vivo de las geometrías del corazón del paciente. Una representación precisa de la geometría ventricular y orientaciones de las fibras fue reconstruido, respectivamente, de alta resolución ex vivo resonancia estructural magnética (RM) y tensor de difusión (DT) imágenes de MR de un corazón humano normal, referido como el atlas. Geometría ventricular de un corazón paciente se extrajo, a través de la segmentación semiautomática, a partir de un estudio in vivo tomografía computarizada (TC) de imagen. Usando algoritmos de transformación de la imagen, la geometría ventricular atlas se deforma para coincidir con la del paciente. Por último, el campo de deformación se aplicó a la orientat atlas fibraiones para obtener una estimación de orientaciones de fibra de pacientes. La precisión de las estimaciones de fibra se evaluó utilizando seis normales y tres corazones insuficientes caninos. La diferencia absoluta media entre los ángulos de inclinación de las orientaciones de fibra adquiridos y estimado fue de 15,4 °. Simulaciones computacionales de mapas de activación ventricular y pseudo-ECG en ritmo sinusal y taquicardia ventricular se indica que no hay diferencias significativas entre las orientaciones de fibra estimados y adquiridos a clínicamente observables ideas nuevas level.The obtenidos del proyecto allanará el camino para el desarrollo de específicos del paciente modelos del corazón que puede ayudar a los médicos en el diagnóstico personalizado y decisiones sobre las intervenciones electrofisiológicos.

La figura 11, muestra AC visualizaciones estilizadas de estimados, así como orientaciones de fibra DTMRI derivados en corazones normales y fallando. Examen cualitativo muestra que las orientaciones de fibra estimados alinean bien con DTMRI derivados de los. Panel D ilustra, superpuesta a la geometría del corazón 1, la distribución del error en ángulos de inclinación de los corazones normales &#39;, promediado en los cinco estimaciones. Panel E muestra la distribución media de error de no haber á...

Watch this Scientific Journal Video about Patient-specific Modeling of the Heart: Estimation of Ventricular Fiber Orientations at JoVE.com

Patient-specific Modeling of the Heart: Estimation of Ventricular Fiber Orientations

Una metodología para estimar las orientaciones de fibras ventriculares de imágenes in vivo de las geometrías del corazón del paciente para el modelado personalizado se describe. Validación de la metodología a cabo utilizando normal y no corazones caninos demostrar que que no hay diferencias significativas entre las orientaciones de las fibras estimados y adquirido a un nivel clínicamente observable.

Modelado específico del paciente del corazón: Estimación de las orientaciones de las fibras ventriculares

Patient-specific  simulations of heart (dys)function aimed at personalizing cardiac therapy are hampered  by the absence of in vivo imaging technology for ...

patient-specific-modeling-heart-estimation-ventricular-fiber

Research

JoVE Journal

Bioengineering

13.6K vistas.  Johns Hopkins University. Una metodología para estimar las orientaciones de fibras ventriculares de imágenes in vivo de las geometrías del corazón del paciente para el modelado personalizado se describe. Validación de la metodología a cabo utilizando normal y no corazones caninos demostrar que que no hay diferencias significativas entre las orientaciones de las fibras estimados y adquirido a un nivel clínicamente observable.

Video: Patient-specific Modeling of the Heart: Estimation of Ventricular Fiber Orientations

High-resolution Fiber-optic Microendoscopy for in situ Cellular Imaging

Many biological and clinical studies require the longitudinal study and analysis of morphology and function with cellular level resolution. Traditionally, multiple experiments are run in parallel, with individual samples removed from the study at sequential time points for evaluation by light microscopy. Several intravital techniques have been developed, with confocal, multiphoton, and second harmonic microscopy all demonstrating their ability to be used for imaging in situ 1. With these systems, however, the required infrastructure is complex and expensive, involving scanning laser systems and complex light sources. Here we present a protocol for the design and assembly of a high-resolution microendoscope which can be built in a day using off-the-shelf components for under US$5,000. The platform offers flexibility in terms of image resolution, field-of-view, and operating wavelength, and we describe how these parameters can be easily modified to meet the specific needs of the end user.
We and others have explored the use of the high-resolution microendoscope (HRME) in in vitro cell culture 2-5, in excised 6 and living animal tissues 2,5, and in human tissues in vivo 2,7. Users have reported the use of several different fluorescent contrast agents, including proflavine 2-4, benzoporphyrin-derivative monoacid ring A (BPD-MA) 5, and fluoroscein 6,7, all of which have received full, or investigational approval from the FDA for use in human subjects. High-resolution microendoscopy, in the form described here, may appeal to a wide range of researchers working in the basic and clinical sciences. The technique offers an effective and economical approach which complements traditional benchtop microscopy, by enabling the user to perform high-resolution, longitudinal imaging in situ.

High-resolution Fiber-optic Microendoscopy for in situ Cellular Imaging

In many biological and clinical situations it is advantageous to study cellular processes as they evolve in their native microenvironment. Here we describe the assembly and use of a low-cost fiber-optic microscope which can provide real time imaging in cell culture, animal studies, and clinical patient studies.

De alta resolución de fibra óptica para Microendoscopy In situ Celular imágenes

Many biological and clinical studies require the longitudinal study and analysis of morphology and function with cellular level resolution.  ...

Investigación

Bioingeniería

Multiparametric Optical Mapping of the Langendorff-perfused Rabbit Heart

Optical imaging and fluorescent probes have significantly advanced research methodology in the field of cardiac electrophysiology in ways that could not have been accomplished by other approaches1. With the use of the calcium- and voltage-sensitive dyes, optical mapping allows measurement of transmembrane action potentials and calcium transients with high spatial resolution without the physical contact with the tissue. This makes measurements of the cardiac electrical activity possible under many conditions where the use of electrodes is inconvenient or impossible1. For example, optical recordings provide accurate morphological changes of membrane potential during and immediately after stimulation and defibrillation, while conventional electrode techniques suffer from stimulus-induced artifacts during and after stimuli due to electrode polarization1. 
The Langendorff-perfused rabbit heart is one of the most studied models of human heart physiology and pathophysiology. Many types of arrhythmias observed clinically could be recapitulated in the rabbit heart model. It was shown that wave patterns in the rabbit heart during ventricular arrhythmias, determined by effective size of the heart and the wavelength of reentry, are very similar to that in the human heart2. It was also shown that critical aspects of excitation-contraction (EC) coupling in rabbit myocardium, such as the relative contribution of sarcoplasmic reticulum (SR), is very similar to human EC coupling3. Here we present the basic procedures of optical mapping experiments in Langendorff-perfused rabbit hearts, including the Langendorff perfusion system setup, the optical mapping systems setup, the isolation and cannulation of the heart, perfusion and dye-staining of the heart, excitation-contraction uncoupling, and collection of optical signals. These methods could be also applied to the heart from species other than rabbit with adjustments to flow rates, optics, solutions, etc.
Two optical mapping systems are described. The panoramic mapping system is used to map the entire epicardium of the rabbit heart4-7. This system provides a global view of the evolution of reentrant circuits during arrhythmogenesis and defibrillation, and has been used to study the mechanisms of arrhythmias and antiarrhythmia therapy8,9. The dual mapping system is used to map the action potential (AP) and calcium transient (CaT) simultaneously from the same field of view10-13. This approach has enhanced our understanding of the important role of calcium in the electrical alternans and the induction of arrhythmia14-16.

This article describes the basic procedures for conducting optical mapping experiments in the Langendorff-perfused rabbit heart using the panoramic imaging system, and the dual (voltage and calcium) imaging modality.

Multiparamétrico óptico Cartografía del corazón de conejo Langendorff perfundidos

Optical imaging and fluorescent probes have significantly advanced research methodology in the field of cardiac electrophysiology in ways that could not ...

Targeting Neuronal Fiber Tracts for Deep Brain Stimulation Therapy Using Interactive, Patient-Specific Models

Deep brain stimulation (DBS), which involves insertion of an electrode to deliver stimulation to a localized brain region, is an established therapy for movement disorders and is being applied to a growing number of disorders. Computational modeling has been successfully used to predict the clinical effects of DBS; however, there is a need for novel modeling techniques to keep pace with the growing complexity of DBS devices. These models also need to generate predictions quickly and accurately. The goal of this project is to develop an image processing pipeline to incorporate structural magnetic resonance imaging (MRI) and diffusion weighted imaging (DWI) into an interactive, patient specific model to simulate the effects of DBS. A virtual DBS lead can be placed inside of the patient model, along with active contacts and stimulation settings, where changes in lead position or orientation generate a new finite element mesh and solution of the bioelectric field problem in near real-time, a timespan of approximately 10 seconds. This system also enables the simulation of multiple leads in close proximity to allow for current steering by varying anodes and cathodes on different leads. The techniques presented in this paper reduce the burden of generating and using computational models while providing meaningful feedback about the effects of electrode position, electrode design, and stimulation configurations to researchers or clinicians who may not be modeling experts.

The goal of this project is to develop an interactive, patient-specific modeling pipeline to simulate the effects of deep brain stimulation in near real-time and provide meaningful feedback as to how these devices influence neural activity in the brain.

Dirigidos a los tractos de fibras neuronales para la terapia de estimulación cerebral profunda mediante modelos interactivos, específico para cada paciente

Deep brain stimulation (DBS), which involves insertion of an electrode to deliver stimulation to a localized brain region, is an established therapy for ...

Light-sheet Fluorescence Microscopy for the Study of the Murine Heart

Light-sheet fluorescence microscopy has been widely used for rapid image acquisition with a high axial resolution from micrometer to millimeter scale. Traditional light-sheet techniques involve the use of a single illumination beam directed orthogonally at sample tissue. Images of large samples that are produced using a single illumination beam contain stripes or artifacts and suffer from a reduced resolution due to the scattering and absorption of light by the tissue. This study uses a dual-sided illumination beam and a simplified CLARITY optical clearing technique for the murine heart. These techniques allow for deeper imaging by removing lipids from the heart and produce a large field of imaging, greater than 10 x 10 x 10 mm3. As a result, this strategy enables us to quantify the ventricular dimensions, track the cardiac lineage, and localize the spatial distribution of cardiac-specific proteins and ion-channels from the post-natal to adult mouse hearts with sufficient contrast and resolution.

This study uses a dual-sided illumination light-sheet fluorescence microscopy (LSFM) technique combined with optical clearing to study the murine heart.

Hoja de luz fluorescencia microscopía para el estudio del corazón murino

Light-sheet fluorescence microscopy has been widely used for rapid image acquisition with a high axial resolution from micrometer to millimeter scale. ...

Biaxial Mechanical Characterizations of Atrioventricular Heart Valves

Extensive biaxial mechanical testing of the atrioventricular heart valve leaflets can be utilized to derive&#160;optimal parameters used in constitutive models, which provide a mathematical representation of the mechanical function of those structures. This presented biaxial mechanical testing protocol involves (i) tissue acquisition, (ii) the preparation of tissue specimens, (iii) biaxial mechanical testing, and (iv) postprocessing of the acquired data. First, tissue acquisition requires obtaining porcine or ovine hearts from a local Food and Drug Administration-approved abattoir for later dissection to retrieve the valve leaflets. Second, tissue preparation requires using tissue specimen cutters on the leaflet tissue to extract a clear zone for testing. Third, biaxial mechanical testing of the leaflet specimen requires the use of a commercial biaxial mechanical tester, which consists of force-controlled, displacement-controlled, and stress-relaxation testing protocols to characterize the leaflet tissue&#39;s mechanical properties. Finally, post-processing requires the use of data image correlation techniques and force and displacement readings to summarize the tissue&#39;s mechanical behaviors in response to external loading. In general, results from biaxial testing demonstrate that the leaflet tissues yield a nonlinear, anisotropic mechanical response. The presented biaxial testing procedure is advantageous to other methods since the method presented here allows for a more comprehensive characterization of the valve leaflet tissue under one unified testing scheme, as opposed to separate testing protocols on different tissue specimens. The proposed testing method has its limitations in that shear stress is potentially present in the tissue sample. However, any potential shear is presumed negligible.

This protocol involves characterizations of atrioventricular valve leaflets with force-controlled, displacement-controlled, and stress-relaxation biaxial mechanical testing procedures. Results acquired with this protocol can be used for constitutive model development to simulate the mechanical behavior of functioning valves under a finite element simulation framework.

Biaxiales caracterizaciones mecánicas de válvulas auriculoventricular del corazón

Extensive biaxial mechanical testing of the atrioventricular heart valve leaflets can be utilized to derive&#160;optimal parameters used in constitutive ...

Optical Imaging of Isolated Murine Ventricular Myocytes

The ability to isolate adult cardiac myocytes has permitted researchers to study a variety of cardiac pathologies at the single cell level. While advances in calcium sensitive dyes have permitted the robust optical recording of single cell calcium dynamics, recording of robust transmembrane optical voltage signals has remained difficult. Arguably, this is because of the low single to noise ratio, phototoxicity, and photobleaching of traditional potentiometric dyes. Therefore, single cell voltage measurements have long been confined to the patch clamp technique which while the gold standard, is technically demanding and low throughput. However, with the development of novel potentiometric dyes, large, fast optical responses to changes in voltage can be obtained with little to no phototoxicity and photobleaching. This protocol describes in detail how to isolate adult murine myocytes which can be used for cellular shortening, calcium, and optical voltage measurements. Specifically, the protocol describes how to use a ratiometric calcium dye, a single-excitation calcium dye, and a single excitation voltage dye. This approach can be used to assess the cardiotoxicity and arrhythmogenicity of various chemical agents. While phototoxicity is still an issue at the single cell level, methodology is discussed on how to reduce it.

We present the methodology for the isolation of murine myocytes and how to obtain voltage or calcium traces simultaneously with sarcomere shortening traces using fluorescence photometry with simultaneous digital cell geometry measurements.

Imágenes ópticas de miocitos ventriculares murinos aislados

The ability to isolate adult cardiac myocytes has permitted researchers to study a variety of cardiac pathologies at the single cell level. While advances ...

Evaluation of Left Ventricular Structure and Function using 3D Echocardiography

Three-dimensional (3D) quantification of the left ventricle (LV) provides significant added value in terms of diagnostic accuracy and precise risk stratification in various cardiac disorders. Recently, 3D echocardiography became available in routine cardiology practice; however, high-quality image acquisition and subsequent analysis have a steep learning curve. The present article aims to guide the reader through a detailed 3D protocol by presenting tips and tricks and also by highlighting the potential pitfalls to facilitate the widespread but technically sound use of this important technique concerning the LV. First and foremost, we show the acquisition of a high-quality 3D dataset with optimal spatial and temporal resolution. Then, we present the analytical steps toward a detailed quantification of the LV by using one of the most widely applied built-in software. We will quantify LV volumes, sphericity, mass and also systolic function by measuring ejection fraction and myocardial deformation (longitudinal and circumferential strain). We will discuss and provide clinical examples about the essential scenarios where the transition from a conventional echocardiographic approach to a 3D-based quantification is highly recommended.

In this article, we provide a step-by-step acquisition and analysis protocol for the volumetric assessment and speckle-tracking analysis of the left ventricle by 3D echocardiography, particularly focusing on practical aspects that maximize the feasibility of this technique.

Evaluación de la estructura y función del ventrículo izquierdo mediante ecocardiografía 3D

Three-dimensional (3D) quantification of the left ventricle (LV) provides significant added value in terms of diagnostic accuracy and precise risk ...

Patient-Specific Polyvinyl Alcohol Phantom Fabrication with Ultrasound and X-Ray Contrast for Brain Tumor Surgery Planning

Phantoms are essential tools for clinical training, surgical planning and the development of novel medical devices. However, it is challenging to create anatomically accurate head phantoms with realistic brain imaging properties because standard fabrication methods are not optimized to replicate any patient-specific anatomical detail and 3D printing materials are not optimized for imaging properties. In order to test and validate a novel navigation system for use during brain tumor surgery, an anatomically accurate phantom with realistic imaging and mechanical properties was required. Therefore, a phantom was developed using real patient data as input and 3D printing of molds to fabricate a patient-specific head phantom comprising the skull, brain and tumor with both ultrasound and X-ray contrast. The phantom also had mechanical properties that allowed the phantom tissue to be manipulated in a similar manner to how human brain tissue is handled during surgery. The phantom was successfully tested during a surgical simulation in a virtual operating room.
The phantom fabrication method uses commercially available materials and is easy to reproduce. The 3D printing files can be readily shared, and the technique can be adapted to encompass many different types of tumor.

This protocol describes the fabrication of a patient specific skull, brain and tumor phantom. It uses 3D printing to create molds, and polyvinyl alcohol (PVA-c) is used as the tissue mimicking material.

Fabricación fantasma de alcohol polivinílico específica del paciente con ultrasonido y contraste de rayos X para la planificación de la cirugía de tumor cerebral

Phantoms are essential tools for clinical training, surgical planning and the development of novel medical devices. However, it is challenging to create ...

Evaluating Regional Pulmonary Deposition using Patient-Specific 3D Printed Lung Models

Development of targeted therapies for pulmonary diseases is limited by the availability of preclinical testing methods with the ability to predict regional aerosol delivery. Leveraging 3D printing to generate patient-specific lung models, we outline the design of a high-throughput, in vitro experimental setup for quantifying lobular pulmonary deposition. This system is made with a combination of commercially available and 3D printed components and allows the flow rate through each lobe of the lung to be independently controlled. Delivery of fluorescent aerosols to each lobe is measured using fluorescence microscopy. This protocol has the potential to promote the growth of personalized medicine for respiratory diseases through its ability to model a wide range of patient demographics and disease states. Both the geometry of the 3D printed lung model and the air flow profile setting can be easily modulated to reflect clinical data for patients with varying age, race, and gender. Clinically relevant drug delivery devices, such as the endotracheal tube shown here, can be incorporated into the testing setup to more accurately predict a device&#8217;s capacity to target therapeutic delivery to a diseased region of the lung. The versatility of this experimental setup allows it to be customized to reflect a multitude of inhalation conditions, enhancing the rigor of preclinical therapeutic testing.

We present a high-throughput, in vitro method for quantifying regional pulmonary deposition at the lobe level using CT scan-derived, 3D printed lung models with tunable air flow profiles.

Evaluación de la deposición pulmonar regional utilizando modelos pulmonares impresos en 3D específicos del paciente

Development of targeted therapies for pulmonary diseases is limited by the availability of preclinical testing methods with the ability to predict ...

Quantification of Mouse Heart Left Ventricular Function, Myocardial Strain, and Hemodynamic Forces by Cardiovascular Magnetic Resonance Imaging

Mouse models have contributed significantly to understanding genetic and physiological factors involved in healthy cardiac function, how perturbations result in pathology, and how myocardial diseases may be treated. Cardiovascular magnetic resonance imaging (CMR) has become an indispensable tool for a comprehensive in vivo assessment of cardiac anatomy and function. This protocol shows detailed measurements of mouse heart left ventricular function, myocardial strain, and hemodynamic forces using 7-Tesla CMR. First, animal preparation and positioning in the scanner are demonstrated. Survey scans are performed for planning imaging slices in various short- and long-axis views. A series of prospective ECG-triggered short-axis (SA) movies (or CINE images) are acquired covering the heart from apex to base, capturing end-systolic and end-diastolic phases. Subsequently, single-slice, retrospectively gated CINE images are acquired in a midventricular SA view, and in 2-, 3-, and 4-chamber views, to be reconstructed into high-temporal resolution CINE images using custom-built and open-source software. CINE images are subsequently analyzed using dedicated CMR image analysis software.
Delineating endomyocardial and epicardial borders in SA end-systolic and end-diastolic CINE images allows for the calculation of end-systolic and end-diastolic volumes, ejection fraction, and cardiac output. The midventricular SA CINE images are delineated for all cardiac time frames to extract a detailed volume-time curve. Its time derivative allows for the calculation of the diastolic function as the ratio of the early filling and atrial contraction waves. Finally, left ventricular endocardial walls in the 2-, 3-, and 4-chamber views are delineated using feature-tracking, from which longitudinal myocardial strain parameters and left ventricular hemodynamic forces are calculated. In conclusion, this protocol provides detailed in vivo quantification of the mouse cardiac parameters, which can be used to study temporal alterations in cardiac function in various mouse models of heart disease.

This study describes a comprehensive cardiovascular magnetic resonance imaging (CMR) protocol to quantify the left ventricular functional parameters of the mouse heart. The protocol describes the acquisition, post-processing, and analysis of the CMR images as well as assessment of different cardiac functional parameters.

Cuantificación de la función ventricular izquierda del corazón de ratón, la distensión miocárdica y las fuerzas hemodinámicas mediante imágenes de resonancia magnética cardiovascular

Mouse models have contributed significantly to understanding genetic and physiological factors involved in healthy cardiac function, how perturbations ...