El financiamiento fue proporcionado por la lucha por la vista. Los tejidos fueron obtenidos a partir del Banco de Ojos Iowa Leones.

1. La disección del segmento anterior. <ol><li> La córnea se elimina en primer lugar hacer una incisión en el limbo en la cámara anterior con una hoja de 15 ° (Figura 1). A continuación, una hoja de curva escleral córnea-tijera se introduce en la cámara anterior. Cortes circunferenciales se hacen en la córnea, justo por delante del limbo. </li><li> 0,12 Colibri pinzas y tijeras Westcott se utilizan para cortar la circunferencia anterior del iris en el cuerpo ciliar. </li><li> El núcleo del cristalino se extrae con pinzas de 0,12 Colibri o una cuchilla de 15 ° (supersharp) y la corteza y la cápsula con las pinzas. </li></ol> 2. La aspiración central del vítreo. <ol><li> Una aguja de calibre 23 en una jeringa de 5 cc se inserta en la mitad de vítreo (Figura 1). </li><li> Aproximadamente 1 ml o más de núcleo vítreo es aspirada suavemente. </li><li> La muestra se coloca en un tubo de microcentrífuga y luego en nitrógeno líquido. </li></ol> 3. La disección hialoidea anterior. <ol><li> La hialoides anterior se ve como un anillo semi-transparente, mediante el ajuste de la luz incidente de una luz de gallina microscopio cuello. Aspiración previa de la base del vítreo se separa la hialoides anterior de otras estructuras para facilitar su identificación. </li><li> La hoja de tejido elástico se hala cuidadosamente lejos del cuerpo ciliar con Colibri o fórceps vestirse y cortarse con tijeras Vannas. Esta maniobra se repite. </li><li> La muestra se coloca en un tubo de microcentrífuga y luego en nitrógeno líquido. </li></ol> 4. Disección base del vítreo. <ol><li> Utilizando unas pinzas 0,12 para estabilizar el ocular, tijeras Westcott se utilizan para &quot;flor&quot; a los ojos, haciendo cuatro equidistantes para aliviar el estrés cortes del cuerpo ciliar hacia la cabeza del nervio óptico. </li><li> La pars plicata del cuerpo ciliar (Figura 2) se elimina de cada uno de los cuadrantes con tijeras de Westcott. </li><li> La base del vítreo se amarra a ambos lados de la ora serrata con unas pinzas sobre la pars plana y la retina (Figura 2). </li><li> La tracción continua sobre la base del vítreo se aplica con las pinzas. Corte con tijeras de Westcott secuencial extractos de un pañuelo de papel semi-transparente, con un &quot;collar de perlas&quot;, aspecto que se extirpa. </li><li> La muestra se coloca en un tubo de microcentrífuga y luego en nitrógeno líquido. </li></ol> 5. Eliminación de la corteza vítrea. <ol><li> La pars plana se extirpa de cada uno de los cuadrantes con tijeras de Westcott mediante la reducción de la hoja de aproximadamente 3 mm por detrás de la ora serrata (Figura 2). </li><li> Entre los prospectos de tejidos, la corteza del vítreo se visualiza como una película semi transparente. </li><li> La película elástica está bien agarrado con pinzas de 0,12 o en poder de la adhesión a una esponja Weck-Cel quirúrgica. La corteza del vítreo se tira a la retina y corta con tijeras Westcott (Figura 1). </li><li> La muestra se coloca en un tubo de microcentrífuga y luego en nitrógeno líquido. Todas las muestras se almacenan a -80 ° C hasta el momento utilizados para la experimentación. </li></ol> 6. Resultados representante Las muestras de tejido puede ser procesado por una variedad de métodos para experimentos específicos. En nuestro caso, las muestras fueron enviadas para análisis de proteínas por SDS-PAGE (Figura 3). <img src="/files/ftp_upload/2455/2455fig1.jpg" alt="Figura 1" /> Figura 1. Corte transversal del ojo humano que representa subestructuras diferentes partes del cuerpo vítreo. El humor vítreo más anterior es una fina capa de colágeno llamada la hialoides anterior. El núcleo vítreo comprende toda la región central del cuerpo vítreo. Esta parte del vítreo es más acuosa, en contraste con la base del vítreo, que es viscoso, lo suficiente como para ser captado por los fórceps y se une firmemente con el cuerpo ciliar y la retina subyacente. Que abarca el núcleo vítreo es un shell de colágeno muy fino llamado la corteza del vítreo. <img src="/files/ftp_upload/2455/2455fig2.jpg" alt="Figura 2" /> Figura 2. Anatomía base del vítreo. La base del vítreo es una subestructura semi-transparente del cuerpo vítreo, situado a lo largo de la ora serrata (flechas blancas), que es la línea divisoria que separa el cuerpo ciliar y la retina. El borde anterior de la base del vítreo se extiende sobre la pars plana (línea blanca) del cuerpo ciliar. El borde posterior de la base del vítreo se extiende de 2 a 3 mm por detrás de la ora serrata (tablero blanco). Para extirpar la base del vítreo, fórceps se utilizan para agarrar el tejido y tire de ella fuera del cuerpo ciliar y la retina subyacente. Una vez elevada, tijeras Westcott se utilizan para cortar a lo largo de la base. <img src="/files/ftp_upload/2455/2455fig3.jpg" alt="Figura 3" /> Figura 3. Una dimensión SDS-PAGE de los elementos del cuerpo vítreo. Concentración de proteínas totales de la hialoides anterior, base del vítreo, vitnúcleo vítreo, y la corteza vítrea fueron 11.24, 20.1, 16.61, 14.24 mg / ml, respectivamente. Electroforesis en gel se realizó en 200 kV durante 45 minutos, se tiñeron con Flamingo (Bio-Rad), y se visualizan mediante un sistema de VersaDoc imágenes (Bio-Rad). Los perfiles de los diferentes tejidos muestran varias bandas similares, lo que indica de conservación o la contaminación cruzada de las proteínas, así como bandas únicas (asterisco), lo que indica proteínas diferencialmente localizados.

<ol>
	<li>Bishop, P. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Structural+macromolecules+and+supramolecular+organisation+of+the+vitreous+gel.">Structural macromolecules and supramolecular organisation of the vitreous gel.</a> Prog Retin Eye Res. 19, 323-344 (2000).</li><li>Sebag, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Molecular+biology+of+pharmacologic+vitreolysis.">Molecular biology of pharmacologic vitreolysis.</a> Trans Am Ophthalmol Soc. 103, 473-494 (2005).</li><li>Yoshimura, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comprehensive+analysis+of+inflammatory+immune+mediators+in+vitreoretinal+diseases.">Comprehensive analysis of inflammatory immune mediators in vitreoretinal diseases.</a> PLoS One. 4, e8158-e8158 (2009).</li><li>Gao, B. B., Chen, X., Timothy, N., Aiello, L. P., Feener, E. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Characterization+of+the+vitreous+proteome+in+diabetes+without+diabetic+retinopathy+and+diabetes+with+proliferative+diabetic+retinopathy.">Characterization of the vitreous proteome in diabetes without diabetic retinopathy and diabetes with proliferative diabetic retinopathy.</a> J Proteome Res. 7, 2516-2525 (2008).</li><li>Izuta, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Extracellular+SOD+and+VEGF+are+increased+in+vitreous+bodies+from+proliferative+diabetic+retinopathy+patients.">Extracellular SOD and VEGF are increased in vitreous bodies from proliferative diabetic retinopathy patients.</a> Mol Vis. 15, 2663-2672 (2009).</li><li>Azimzadeh, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Formalin-fixed+paraffin-embedded+(FFPE)+proteome+analysis+using+gel-free+and+gel-based+proteomics.">Formalin-fixed paraffin-embedded (FFPE) proteome analysis using gel-free and gel-based proteomics.</a> J Proteome Res. 9, 4710-4720 (2010).</li></ol>

El cuerpo vítreo es un gel semi-transparente, cuya composición molecular es poco conocida, especialmente en el ámbito de sus subestructuras: la base del vítreo, el núcleo, la corteza y hialoidea anterior. El núcleo vítreo contiene colágeno II, V, IX y XI, junto con el sulfato de condroitina proteoglicanos, proteoglicanos heparán sulfato, y ácido hialurónico. 1,2 biomarcadores de proteínas en el núcleo vítreo se han asociado con enfermedades como la retinopatía diabética. 3.5 ¿Cómo estas proteínas se expresan diferencialmente en cada una de las subestructuras, y en muchos casos las identidades de proteínas específicas, no se conocen. Estos detalles pueden dar una idea sobre el origen de las proteínas asociadas con enfermedades específicas vítreo-retiniana y ayudar a las terapias blanco en el futuro. Aunque el mensaje óptimo intervalo mortem para la disección de los tejidos no se ha determinado, la degradación de proteínas puede afectar a los experimentos posteriores. Por ejemplo, la inmunohistoquímica se ve afectado en los ojos postmortem de 12 horas y algunas actividades enzimáticas específicas se puede reducir en unas pocas horas (observaciones no publicadas). Todos los tejidos de este estudio fueron recogidos entre 2 y 8 horas de la muerte, sin cambios significativos en la expresión de la proteína o la idoneidad para analsyis proteómica. El método de congelación de nitrógeno líquido de conservación se elige sobre la fijación con el fin de evitar que pequeños cambios en la estructura de las proteínas causada por fijador de reticulación, que se ha demostrado en otros tejidos por LC-MS/MS. Seis estudios proteómicos dependerá de la capacidad de precisión analizar los diferentes compartimentos del vítreo, como se demuestra en este experimento de vídeo. Hemos validado la técnica de disección con una dimensión SDS-PAGE. A medida que nuestros resultados sugieren, que se expresan diferencialmente en las proteínas de las subestructuras de diversas partes del cuerpo vítreo. La identificación de estas proteínas proporcionan una comprensión más detallada de la compartimentalización del vítreo.

<table border="1"><tbody><tr><td> Nombre </td><td> Empresa </td><td> Número de catálogo </td></tr><tr><td> 0,12 fórceps </td><td> Storz Ophthalmics </td><td> E1502 </td></tr><tr><td> 5 cc jeringa </td><td> Becton-Dickinson </td><td> 309603 </td></tr><tr><td> Recta Pinzas Vestirse con dientes </td><td> Storz Ophthalmics </td><td> E1400 </td></tr><tr><td> 23 calibre de la aguja </td><td> Becton-Dickinson </td><td> 305145 </td></tr><tr><td> Colibri fórceps </td><td> Storz Ophthalmics </td><td> 2 / 132 </td></tr><tr><td> Castroviejo córnea tijeras en ángulo </td><td> Storz Ophthalmics </td><td> E3223 </td></tr><tr><td> Vannas tijeras curvas de la capsulotomía </td><td> Storz Ophthalmics </td><td> E3387 </td></tr><tr><td> Weck-Cel quirúrgica lanzas </td><td> Medtronic </td><td> 0008680 </td></tr><tr><td> Westcott Tijeras tenotomía curvas </td><td> Storz Ophthalmics </td><td> E3320 </td></tr></tbody></table>

dissection of human vitreous body elements for proteomic analysis

El humor vítreo es un ópticamente transparente, la matriz extracelular de colágeno que llena el interior del ojo y se superpone a la retina. 1,2 interacciones anormales entre las subestructuras de vítreo y la retina la base de varias enfermedades vítreo-retiniana, incluyendo desgarro retiniano y desprendimiento, pliegue macular, agujero macular, relacionada con la edad degeneración macular, la tracción vitreomacular, vitreorretinopatía proliferativa, retinopatía diabética proliferativa, y vitreoretinopathies heredado. 1,2 La composición molecular de las subestructuras del vítreo no se conoce. Dado que el cuerpo vítreo es transparente con el acceso quirúrgico limitado, ha sido difícil de estudiar sus subestructuras en el nivel molecular. Hemos desarrollado un método para separar y conservación de estos tejidos para análisis proteómico y bioquímicos. La técnica de disección en este video experimental muestra la forma de aislar a base del vítreo, hialoidea anterior, vítreo núcleo y la corteza vítreo de ojos de cadáver. Una dimensión SDS-PAGE análisis de cada componente vítreo mostró que nuestra técnica de disección resultó en cuatro perfiles de proteínas único correspondiente a cada sub-estructura del cuerpo vítreo humano. Identificación de proteínas diferencialmente en compartimientos revelará las moléculas candidatas subyacente de diversas enfermedades vítreo-retiniana.

Watch this Scientific Journal Video about Dissection of Human Vitreous Body Elements for Proteomic Analysis at JoVE.com

Dissection of Human Vitreous Body Elements for Proteomic Analysis

Este video muestra una técnica eficaz para la diferenciación y la disección de las diferentes estructuras semi-transparente del cuerpo humano en el vítreo posterior ojos mortem.

La disección de los elementos humanos Cuerpo Vítreo para el análisis proteómico

The vitreous is an optically clear, collagenous extracellular matrix that fills the inside of the eye and overlies the retina. 1,2 Abnormal interactions ...

dissection-of-human-vitreous-body-elements-for-proteomic-analysis

Research

JoVE Journal

Medicine

31.7K vistas.  University of Iowa. Este video muestra una técnica eficaz para la diferenciación y la disección de las diferentes estructuras semi-transparente del cuerpo humano en el vítreo posterior ojos mortem.

Video: Dissection of Human Vitreous Body Elements for Proteomic Analysis

Whole Vitreous Humor Dissection for Vitreodynamic Analysis

The authors propose an effective technique to isolate whole, intact vitreous core and cortex from post mortem enucleated porcine eyes. While previous studies have shown the results of such dissections, the detailed steps have not been described, precluding researchers outside the field from replicating their methods. Other studies harvest vitreous either through aspiration, which does not maintain the vitreous structure anatomy, or through partial dissection, which only isolates the vitreous core. The proposed method isolates the whole vitreous body, with the vitreous core and cortex intact, while maintaining vitreous anatomy and structural integrity. In this method, a full thickness scleral flap in an enucleated porcine eye is first created and through this, the choroid tissue can be separated from the sclera. The scleral flap is then expanded and the choroid is completely separated from the sclera. Finally the choroid-retina tissue is peeled off the vitreous to leave an isolated intact vitreous body. The proposed vitreous dissection technique can be used to study physical properties of the vitreous humor. In particular, this method has significance for experimental studies involving drug delivery, vitreo-retinal oxygen transport, and intraocular convection.

The goal of this protocol is to show an effective technique to isolate whole, intact vitreous core and cortex from post mortem enucleated porcine eyes.

Todo el humor vítreo Disección de Análisis Vitreodynamic

The authors propose an effective technique to isolate whole, intact vitreous core and cortex from post mortem enucleated porcine eyes. While previous ...

Investigación

Medicina

Laser-capture Microdissection of Human Prostatic Epithelium for RNA Analysis

The prostate gland contains a heterogeneous milieu of stromal, epithelial, neuroendocrine and immune cell types. Healthy prostate is comprised of fibromuscular stroma surrounding discrete epithelial-lined secretory lumens and a very small population of immune and neuroendocrine cells. In contrast, areas of prostate cancer have increased dysplastic luminal epithelium with greatly reduced or absent stromal population. Given the profound difference between stromal and epithelial cell types, it is imperative to separate the cell types for any type of downstream molecular analysis. Despite this knowledge, the bulk of gene expression studies compare benign prostate to cancer without micro-dissection, leading to stromal bias in the benign samples. Laser-capture micro-dissection (LCM) is an effective method to physically separate different cell types from a specimen section. The goal of this protocol is to show that RNA can be successfully isolated from LCM-collected human prostatic epithelium and used for downstream gene expression studies such as RT-qPCR and RNAseq.

The goal of this protocol is to use laser-capture micro-dissection as an effective method to isolate pure populations of cell types from heterogeneous prostate tissues for downstream RNA analysis.

Laser-captura microdisección de Human Prostática epitelio de Análisis de ARN

The prostate gland contains a heterogeneous milieu of stromal, epithelial, neuroendocrine and immune cell types.  Healthy prostate is comprised of ...

Dissection of the Mouse Pancreas for Histological Analysis and Metabolic Profiling

We have been investigating the pancreas specific transcription factor, 1a cre-recombinase; lox-stop-lox- Kristen rat sarcoma, glycine to aspartic acid at the 12 codon (Ptf1acre/+;LSL-KrasG12D/+) mouse strain as a model of human pancreatic cancer. The goal of our current studies is to identify novel metabolic biomarkers of pancreatic cancer progression. We have performed metabolic profiling of urine, feces, blood, and pancreas tissue extracts, as well as histological analyses of the pancreas to stage the cancer progression. The mouse pancreas is not a well-defined solid organ like in humans, but rather is a diffusely distributed soft tissue that is not easily identified by individuals unfamiliar with mouse internal anatomy or by individuals that have little or no experience performing mouse organ dissections. The purpose of this article is to provide a detailed step-wise visual demonstration to guide novices in the removal of the mouse pancreas by dissection. This article should be especially valuable to students and investigators new to research that requires harvesting of the mouse pancreas by dissection for metabolic profiling or histological analyses.

This video article provides a detailed demonstration of the procedures required to successfully remove the pancreas from a mouse by dissection for histological analysis and metabolic profiling.

Disección del páncreas de ratón para análisis histológico y perfiles metabólicos

We have been investigating the pancreas specific transcription factor, 1a cre-recombinase; lox-stop-lox- Kristen rat sarcoma, glycine to aspartic acid at ...

Dissection and Explant Culture of Murine Allantois for the In Vitro Analysis of Allantoic Attachment

The placenta is essential for the growth and development of mammalian embryos. For this reason, numerous genetic alterations and likely also environmental insults that disturb placenta development or function can cause early pregnancy loss in mice and humans. Nevertheless, simple in vitro assays to screen for potential effects on placenta formation are lacking. Here, we focus on modeling the first and critical step in placenta formation, which consists of the attachment of the allantois to the chorion. We describe a method to rapidly assess the attachment of allantoic explants on immobilized &#945;4&#946;1 integrin, which serves as a chorio-mimetic substrate.This in vitro approach enables a qualitative evaluation of the attachment and spreading behavior of multiple allantois explants at different consecutive time points. The protocol may be used to investigate the effect of targeted mouse mutations, drugs, or various environmental factors that have been linked to pregnancy complications or fetal loss on allantois attachment ex vivo.

Dissection and Explant Culture of Murine Allantois for the In Vitro Analysis of Allantoic Attachment

We describe an in vitro assay to model chorioallantoic attachment, the first step in placenta formation. The protocol demonstrates the dissection and explant culture of murine allantoides on immobilized &#945;4&#946;1 integrin. Allantois attachment is evaluated microscopically at pre-determined time points.

Disección y la cultura de explante de Allantois murino para el análisis In Vitro del accesorio alantoideo

The placenta is essential for the growth and development of mammalian embryos. For this reason, numerous genetic alterations and likely also environmental ...

Body Composition and Metabolic Caging Analysis in High Fat Fed Mice

Alterations to body composition (fat or lean mass), metabolic parameters such as whole-body oxygen consumption, energy expenditure, and substrate utilization, and behaviors such as food intake and physical activity can provide important information regarding the underlying mechanisms of disease. Given the importance of body composition and metabolism to the development of obesity and its subsequent sequelae, it is necessary to make accurate measures of these parameters in the pre-clinical research setting. Advances in technology over the past few decades have made it possible to derive these measures in rodent models in a non-invasive and longitudinal fashion. Consequently, these metabolic measures have proven useful when assessing the response of genetic manipulations (for example knockout or transgenic mice, viral knock-down or overexpression of genes), experimental drug/compound screening and dietary, behavioral or physical activity interventions. Herein, we describe the protocols used to measure body composition and metabolic parameters using an animal monitoring system in chow-fed and high fat diet-fed mice.

This protocol describes the use of a body composition analyzer and metabolic animal monitoring system to characterize body composition and metabolic parameters in mice. An obesity model induced by high-fat feeding is used as an example for the application of these techniques.

Composición corporal y análisis jaulas metabólico en ratones de reserva grasa alta

Alterations to body composition (fat or lean mass), metabolic parameters such as whole-body oxygen consumption, energy expenditure, and substrate ...

Quantitative Micro-CT Analysis of Aortopathy in a Mouse Model of &#946;-aminopropionitrile-induced Aortic Aneurysm and Dissection

Aortic aneurysm and dissection is associated with significant morbidity and mortality in the population and can be highly lethal. While animal models of aortic disease exist, in vivo imaging of the vasculature has been limited. In recent years, micro-computerized tomography (micro-CT) has emerged as a preferred modality for imaging both large and small vessels both in vivo and ex vivo. In conjunction with a method of vascular casting, we have successfully used micro-CT to characterize the frequency and distribution of aortic pathology in &#946;-aminopropionitrile-treated C57/Bl6 mice. Technical limitations of this method include variations in the quality of the perfusion introduced by poor animal preparation, the application of proper methodologies for vessel size quantification, and the non-survivability of this procedure. This article details a methodology for the intravascular perfusion of a lead-based radiopaque silicone rubber for the quantitative characterization of aortopathy in a mouse model of aneurysm and dissection. In addition to visualizing aortic pathology, this method may be used for examining other vascular beds in vivo or vascular beds removed post-mortem.

Quantitative Micro-CT Analysis of Aortopathy in a Mouse Model of &amp;#946;-aminopropionitrile-induced Aortic Aneurysm and Dissection

This article describes a detailed methodology of using a radiopaque lead-based silicone rubber to perfuse the murine vasculature for aortic diameter quantification in a mouse model of aortic aneurysm and dissection.

Análisis de Micro-CT cuantitativa de Aortopathy en un modelo murino de disección y aneurisma de la aorta inducida por β-oxidasa

Aortic aneurysm and dissection is associated with significant morbidity and mortality in the population and can be highly lethal. While animal models of ...

Whole Body and Regional Quantification of Active Human Brown Adipose Tissue Using 18F-FDG PET/CT

In endothermic animals, brown adipose tissue (BAT) is activated to produce heat for defending body temperature in response to cold. BAT&#39;s ability to expend energy has made it a potential target for novel therapies to ameliorate obesity and associated metabolic disorders in humans. Though this tissue has been well studied in small animals, BAT&#39;s thermogenic capacity in humans remains largely unknown due to the difficulties of measuring its volume, activity, and distribution. Identifying and quantifying active human BAT is commonly performed using 18F-Fluorodeoxyglucose (18F-FDG) positron emission tomography and computed tomography (PET/CT) scans following cold-exposure or pharmacological activation. Here we describe a detailed image-analysis approach to quantify total-body human BAT from 18F-FDG PET/CT scans using an open-source software. We demonstrate the drawing of user-specified regions of interest to identify metabolically active adipose tissue while avoiding common non-BAT tissues, to measure BAT volume and activity, and to further characterize its anatomical distribution. Although this rigorous approach is time-consuming, we believe it will ultimately provide a foundation to develop future automated BAT quantification algorithms.

Whole Body and Regional Quantification of Active Human Brown Adipose Tissue Using 18F-FDG PET/CT

Using free, open-source software, we have developed an analytical approach to quantify total and regional brown adipose tissue (BAT) volume and metabolic activity of BAT using 18F-FDG PET/CT.

Cuerpo entero y cuantificación Regional de activo humano marrón tejido adiposo utilizando 18F-FDG PET/CT

In endothermic animals, brown adipose tissue (BAT) is activated to produce heat for defending body temperature in response to cold. BAT&#39;s ability to ...

Segmentation and Linear Measurement for Body Composition Analysis using Slice-O-Matic and Horos

Body composition is associated with risk of disease progression and treatment complications in a variety of conditions. Therefore, quantification of skeletal muscle mass and adipose tissues on Computed Tomography (CT) and/or Magnetic Resonance Imaging (MRI) may inform surgery risk evaluation and disease prognosis. This article describes two quantification methods originally described by Mourtzakis et al. and Avrutin et al.: tissue segmentation and linear measurement of skeletal muscle. Patients' cross-sectional image at the midpoint of the third lumbar vertebra was obtained for both measurements. For segmentation, the images were imported into Slice-O-Matic and colored for skeletal muscle, intramuscular adipose tissue, visceral adipose tissue, and subcutaneous adipose tissue. Then, surface areas of each tissue type were calculated using the tag surface area function. For linear measurements, the height and width of bilateral psoas and paraspinal muscles at the level of the third lumbar vertebra are measured and the calculation using these four values yield the estimated skeletal muscle mass. Segmentation analysis provides quantitative, comprehensive information about the patients' body composition, which can then be correlated with disease progression. However, the process is more time-consuming and requires specialized training. Linear measurements are an efficient and clinic-friendly tool for quick preoperative evaluation. However, linear measurements do not provide information on adipose tissue composition. Nonetheless, these methods have wide applications in a variety of diseases to predict surgical outcomes, risk of disease progression and inform treatment options for patients.

Segmentation and linear measurements quantify skeletal muscle mass and adipose tissues using Computed Tomography and/or Magnetic Resonance Imaging images. Here, we outline the use of Slice-O-Matic software and Horos image viewer for rapid and accurate analysis of body composition. These methods can provide important information for prognosis and risk stratification.

Segmentación y medición lineal para el análisis de la composición corporal utilizando Slice-O-Matic y Horos

Body composition is associated with risk of disease progression and treatment complications in a variety of conditions. Therefore, quantification of ...

Location, Dissection, and Analysis of the Murine Stellate Ganglion

The autonomic nervous system is a substantial driver of cardiac electrophysiology. Especially&#160;the role of its sympathetic branch is an ongoing matter of investigation in the pathophysiology of ventricular arrhythmias (VA). Neurons in the stellate ganglia (SG)&#160;&#8211;&#160;bilateral star-shaped structures of the sympathetic chain &#8211;&#160;are an important component of the sympathetic infrastructure. The SG are a recognized target for&#160;treatment via cardiac sympathetic denervation in patients with therapy-refractory VA. While neuronal remodeling and glial activation in the SG have been described in patients with VA, the underlying cellular and molecular processes that potentially precede the onset of arrhythmia are only insufficiently understood and should be elucidated to improve autonomic modulation. Mouse models allow us to study sympathetic neuronal remodeling, but identification of the murine SG is challenging for the inexperienced investigator. Thus, in-depth cellular and molecular biological studies of the murine SG are lacking for many common cardiac diseases. Here, we describe a basic repertoire for dissecting and studying the SG in adult mice for analyses at RNA level (RNA isolation for gene expression analyses, in situ hybridization), protein level (immunofluorescent whole mount staining), and cellular level (basic morphology, cell size measurement). We present potential solutions to overcome challenges in the preparation technique, and how to improve staining via quenching of autofluorescence. This allows for the visualization of neurons as well as glial cells via established markers in order to determine cell composition and remodeling processes. The methods presented here allow characterizing the SG to gain further information on autonomic dysfunction in mice prone to VA and can be complemented by additional techniques investigating neuronal and glial components of the autonomic nervous system in the heart.

Pathophysiological changes in the cardiac autonomic nervous system, especially in its sympathetic branch, contribute to the onset and maintenance of ventricular arrhythmias. In the present protocol, we show how to characterize murine stellate ganglia to improve the understanding of the underlying molecular and cellular processes.

Localización, Disección, Y Análisis Del Ganglio Estrellado Murino

The autonomic nervous system is a substantial driver of cardiac electrophysiology. Especially&#160;the role of its sympathetic branch is an ongoing matter ...

A Non-Invasive Method for Generating the Cyclic Loading-Induced Intra-Articular Cartilage Lesion Model of the Rat Knee

The pathophysiology of primary osteoarthritis (OA) remains unclear. However, a specific subclassification of OA in relatively younger age groups is likely correlated with a history of articular cartilage damage and ligament avulsion. Surgical animal models of OA of the knee play an important role in understanding the onset and progression of post-traumatic OA and aid in the development of novel therapies for this disease. However, non-surgical models have been recently considered to avoid traumatic inflammation that could affect the evaluation of the intervention. 
In this study, an intra-articular cartilage lesion rat model induced by in vivo cyclic compressive loading was developed, which allowed researchers to (1) determine the optimal magnitude, speed, and duration of load that could cause focal cartilage damage; (2) assess post-traumatic spatiotemporal pathological changes in chondrocyte vitality; and (3) evaluate the histological expression of destructive or protective molecules that are involved in the adaptation and repair mechanisms against joint compressive loads. This report describes the experimental protocol for this novel cartilage lesion in a rat model.

Here, we present the cyclic loading-induced intra-articular cartilage lesion model of the rat knee, generated by 60 cyclic compressions over 20 N, resulting in damage to the femoral condylar cartilage in rats.

Un método no invasivo para generar el modelo de lesión de cartílago intraarticular inducida por carga cíclica de la rodilla de rata

The pathophysiology of primary osteoarthritis (OA) remains unclear. However, a specific subclassification of OA in relatively younger age groups is likely ...