Los autores desean agradecer a Hartwig Wieboldt por su excelente asistencia técnica, y al Centro de Microscopía de Imágenes UKE (Umif) del Centro Médico Universitario Hamburgo-Eppendorf por proporcionar microscopios y apoyo. Esta investigación fue financiada por el DZHK (Centro Alemán de Investigación Cardiovascular) [FKZ 81Z4710141].

Los autores no tienen nada que revelar.

La comprensión de los procesos celulares y moleculares en neuronas y células gliales del sistema nervioso simpático que preceden a la aparición del AV es de alto interés, ya que la parada cardíaca súbita sigue siendo la causa más común de muerte en todo el mundo5. Por lo tanto, en el manuscrito actual, proporcionamos un repertorio básico de métodos para identificar el SG murino , un elemento murino dentro de esta red , y realizar análisis posteriores a nivel de ARN, proteína y celular...

El sistema nervioso autónomo cardíaco es un equilibrio estrechamente regulado de componentes simpáticos, parasimpáticos y sensoriales que permite al corazón adaptarse a los cambios ambientales con la respuesta fisiológica adecuada1,2. Las alteraciones en este equilibrio, por ejemplo, un aumento de la actividad simpática, se han establecido como un factor clave para la aparición, así como el mantenimiento de las arritmias ventriculares (AV)3,4. Por lo tanto, la modulación autonómica, lograda a través de la reducción farmacológica de la actividad simpática con betabloqueantes, ha sido una piedra angular en el tratamiento de pacientes con AV durante décadas5,6. Pero a pesar de las intervenciones farmacológicas y basadas en catéteres, un número relevante de pacientes todavía sufre de VArecurrente 7.
La entrada simpática al corazón está mediada principalmente a través de cuerpos celulares neuronales en los ganglios estrellados (SG), estructuras bilaterales en forma de estrella de la cadena simpática, que transmiten información a través de numerosos nervios intratorácicos desde el tronco encefálico hasta elcorazón 8,9,10. La brotación nerviosa del SG después de la lesión se asocia con el AV y la muerte súbita cardíaca11,12,destacando el SG como objetivo de la modulación autonómica13,14. Una reducción de la entrada simpática al corazón se puede lograr temporalmente a través de la inyección percutánea de anestésicos locales o permanentemente mediante la extirpación parcial de la SG a través de la toracoscopia asistida por video15,16. La denervación simpática cardíaca presenta una opción para los pacientes con AV refractaria a la terapia con resultados prometedores14,16,17. Hemos aprendido de la SG explantada de estos pacientes que la remodelación neuronal y neuroquímica, la neuroinflamación y la activación glial son sellos de remodelación simpática que podrían contribuir o agravar la disfunción autonómica18,19. Aún así, los procesos celulares y moleculares subyacentes en estas neuronas siguen siendo oscuros hasta la fecha, por ejemplo, el papel de la transdiferenciación neuronal en un fenotipo colinérgico20,21. Los estudios experimentales presentan nuevos enfoques para tratar el AV, por ejemplo, la reducción de la actividad nerviosa simpática a través de la optogenética22,pero la caracterización en profundidad de la SG todavía falta en muchas patologías cardíacas que van de la mano con el AV. Los modelos de ratón que imitan estas patologías permiten estudiar la remodelación neuronal que potencialmente precede a la aparición de arritmias12,23. Éstos se pueden completar por otros análisis morfológicos y funcionales para la caracterización autonómica del corazón y del sistema nervioso. En el presente protocolo, proporcionamos un repertorio básico de métodos que permiten diseccionar y caracterizar el SG murino para mejorar la comprensión del AV.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>96-well plate</td>
			<td>TPP</td>
			<td>92097</td>
			<td>RNAscope</td>
		</tr>
		<tr>
			<td>Adhesion Slides SuperFrost plus&#160; 25 x 75 x 1 mm</td>
			<td>R. Langenbrinck</td>
			<td>03-0060</td>
			<td>Microscopy</td>
		</tr>
		<tr>
			<td>Albumin bovine Fraction V receptor grade lyophil.</td>
			<td>Serva</td>
			<td>11924.03</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>bisBenzimide H33342 trihydrochloride (Hoechst)</td>
			<td>Sigma-Aldrich, St. Louis, MO, USA</td>
			<td>B2261</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>Chicken anti neurofilament</td>
			<td>EMD Millipore</td>
			<td>AB5539</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>Dimethyl sulfoxide (DMSO)</td>
			<td>Merck, KGA, Darmstadt, Germany</td>
			<td>D8418</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>Donkey anti chicken IgY Alexa 647&#160;</td>
			<td>Merck, KGA, Darmstadt, Germany</td>
			<td>AP194SA6</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>Donkey anti goat IgG Alexa 568&#160;</td>
			<td>Thermo Fisher Scientific</td>
			<td>A11057</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>Donkey anti rabbit IgG Alexa 488&#160;</td>
			<td>Thermo Fisher Scientific</td>
			<td>A21206</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>Drying block 37-100 mm</td>
			<td>Whatman (Sigma Aldrich)</td>
			<td>WHA10310992&#160;</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>Eosin Y</td>
			<td>Sigma Aldrich</td>
			<td>E4009</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>Ethanol 99 % denatured with MEK, IPA and Bitrex (min. 99,8 %)</td>
			<td>Th.Geyer</td>
			<td>2212.5000</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>Eukitt mounting medium</td>
			<td>AppliChem</td>
			<td>253681.0008</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>Fluoromount-G</td>
			<td>Southern Biotech</td>
			<td>0100-01</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>Fluoromount-G + DAPI</td>
			<td>Southern Biotech</td>
			<td>0100-20</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>Goat anti choline acetyltransferase</td>
			<td>EMD Millipore</td>
			<td>AP144P</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>H2O2 30% (w/w)</td>
			<td>Merck, KGA, Darmstadt, Germany</td>
			<td>H1009</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>Heparin Sodium 25.000 UI / 5ml</td>
			<td>Rotexmedica</td>
			<td>PZN: 3862340</td>
			<td>Preparation SG</td>
		</tr>
		<tr>
			<td>High-capacity cDNA reverse transctiption kit</td>
			<td>Life technologies</td>
			<td>&#160;4368813</td>
			<td>RNA isolation</td>
		</tr>
		<tr>
			<td>Isoflurane (Forene)</td>
			<td>Abbott Laboratories</td>
			<td>2594.00.00</td>
			<td>Preparation SG</td>
		</tr>
		<tr>
			<td>Mayer&#39;s hemalum solution</td>
			<td>Merck</td>
			<td>1.09249.0500</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>Methanol</td>
			<td>Sigma-Aldrich</td>
			<td>34860</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>Microscope cover glasses 20x20 mm or smaller</td>
			<td>Marienfeld</td>
			<td>0101040</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>miRNeasy Mini Kit</td>
			<td>Qiagen</td>
			<td>217004</td>
			<td>RNA isolation</td>
		</tr>
		<tr>
			<td>NanoDrop 2000c</td>
			<td>Thermo Fisher Scientific</td>
			<td>ND-2000C</td>
			<td>RNA isolation</td>
		</tr>
		<tr>
			<td>Opal 570 Reagent Pack</td>
			<td>Akoya Bioscience</td>
			<td>FP1488001KT</td>
			<td>RNAscope</td>
		</tr>
		<tr>
			<td>Paraformaldehyde, 16% w/v aq. soln., methanol free&#160;</td>
			<td>Alfa Aesar</td>
			<td>43368</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>Pasteur pipettes, LDPE, unsterile, 3 ml, 154 mm</td>
			<td>Th.Geyer</td>
			<td>7691202</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>Phosphate-buffered saline tablets</td>
			<td>Gibco</td>
			<td>18912-014</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>Pinzette Dumont SS Forceps</td>
			<td>FineScienceTools</td>
			<td>11203-25</td>
			<td>Preparation SG</td>
		</tr>
		<tr>
			<td>QIAzol Lysis Reagent</td>
			<td>Qiagen&#160;</td>
			<td>79306</td>
			<td>RNA isolation</td>
		</tr>
		<tr>
			<td>Rabbit anti tyrosine hydroxylase</td>
			<td>EMD Millipore</td>
			<td>AB152</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>RNAlater</td>
			<td>Merck</td>
			<td>R0901-100ML</td>
			<td>RNA isolation (optional)</td>
		</tr>
		<tr>
			<td>RNAscope Multiplex Fluorescent Reagent Kit v2</td>
			<td>biotechne (ACD)</td>
			<td>323100</td>
			<td>RNAscope</td>
		</tr>
		<tr>
			<td>RNAscope Probe-Mm-S100b-C2</td>
			<td>biotechne (ACD)</td>
			<td>431738-C2</td>
			<td>RNAscope</td>
		</tr>
		<tr>
			<td>RNAscope Probe-Mm-Tubb3</td>
			<td>biotechne (ACD)</td>
			<td>423391</td>
			<td>RNAscope</td>
		</tr>
		<tr>
			<td>Stainless steel beads 7 mm&#160;</td>
			<td>Qiagen&#160;</td>
			<td>69990</td>
			<td>RNA isolation</td>
		</tr>
		<tr>
			<td>Sudan black B</td>
			<td>Roth</td>
			<td>0292.2</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>TaqMan Gene Expression Assay Cdkn1b (Mm00438168_m1)</td>
			<td>Thermo Fisher Scientific</td>
			<td>4331182</td>
			<td>Gene expression analysis</td>
		</tr>
		<tr>
			<td>TaqMan Gene Expression Assay Choline acetyltransferase (Mm01221880_m1)</td>
			<td>Thermo Fisher Scientific</td>
			<td>4331182</td>
			<td>Gene expression analysis</td>
		</tr>
		<tr>
			<td>TaqMan Gene Expression Assay MKi67 (Mm01278617_m1)</td>
			<td>Thermo Fisher Scientific</td>
			<td>4331182</td>
			<td>Gene expression analysis</td>
		</tr>
		<tr>
			<td>TaqMan Gene Expression Assay PTPCR (Mm01293577_m1)</td>
			<td>Thermo Fisher Scientific</td>
			<td>4331182</td>
			<td>Gene expression analysis</td>
		</tr>
		<tr>
			<td>TaqMan Gene Expression Assay S100b (Mm00485897_m1)</td>
			<td>Thermo Fisher Scientific</td>
			<td>4331182</td>
			<td>Gene expression analysis</td>
		</tr>
		<tr>
			<td>TaqMan Gene Expression Assay Tyrosin Hydroxylase (Mm00447557_m1)</td>
			<td>Thermo Fisher Scientific</td>
			<td>4331182</td>
			<td>Gene expression analysis</td>
		</tr>
		<tr>
			<td>TaqMan mastermix</td>
			<td>Applied biosystems</td>
			<td>4370074</td>
			<td>Gene Expression analysis&#160;</td>
		</tr>
		<tr>
			<td>Tissue Lyser II</td>
			<td>Qiagen</td>
			<td>85300</td>
			<td>RNA isolation</td>
		</tr>
		<tr>
			<td>Triton X-100 10% solution</td>
			<td>Sigma-Aldrich</td>
			<td>93443-100ml</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>Tween-20</td>
			<td>Sigma-Aldrich</td>
			<td>P9416-100ML</td>
			<td>RNAscope</td>
		</tr>
		<tr>
			<td>Wacom bamboo pen</td>
			<td>Wacom</td>
			<td>CTL-460/K</td>
			<td>Cell size measurements</td>
		</tr>
		<tr>
			<td>Whatman prepleated qualitative filter paper, Grade 595 1/2</td>
			<td>Sigma-Aldrich</td>
			<td>WHA10311647</td>
			<td>Whole mount staining</td>
		</tr>
		<tr>
			<td>Wheat Germ Agglutinin, Alexa Fluor 633 Conjugate</td>
			<td>Thermo Fisher Scientific</td>
			<td>W21404</td>
			<td>RNAscope</td>
		</tr>
	</tbody>
</table>

location, dissection, and analysis of the murine stellate ganglion

El sistema nervioso autónomo es un conductor substancial de la electrofisiología cardiaca. Especialmente el papel de su rama comprensiva es una materia en curso de la investigación en la patofisiología de las arritmias ventriculares (VA). Las neuronas en los ganglios estrellados (SG)   – estructuras bilaterales en forma de estrella de la cadena simpática –   son un componente importante de la infraestructura simpática. Los SG son una blanco reconocida para el tratamiento vía la desnervación comprensiva cardiaca en pacientes con el VA terapia-refractario. Mientras que el remodelado neuronal y la activación glial en el SG se han descrito en pacientes con el VA, los procesos celulares y moleculares subyacentes que potencialmente preceden el inicio de la arritmia se entienden solamente escaso y se deben aclarar para mejorar la modulación autonómica. Los modelos de ratón nos permiten estudiar la remodelación neuronal simpática, pero la identificación del SG murino es un desafío para el investigador inexperto. Así, los estudios biológicos celulares y moleculares profundizados del SG murine están careciendo para muchas enfermedades cardiacas comunes. Aquí, se describe un repertorio básico para la disección y el estudio de la SG en ratones adultos para análisis a nivel de ARN (aislamiento de ARN para análisis de expresión génica, hibridación in situ), nivel de proteína (inmunofluorescente de montaje entero de tinción), y el nivel celular (morfología básica, medición del tamaño celular). Presentamos soluciones potenciales para superar los desafíos en la técnica de preparación, y cómo mejorar la tinción a través del temple de la autofluorescencia. Esto permite la visualización de neuronas, así como células gliales a través de marcadores establecidos con el fin de determinar la composición celular y los procesos de remodelación. Los métodos aquí presentados permiten caracterizar el SG para obtener más información sobre la disfunción autonómica en ratones propensos al VA y pueden complementarse con técnicas adicionales que investigan los componentes neuronales y gliales del sistema nervioso autónomo en el corazón.

La Figura 1 visualiza cómo identificar y diseccionar el SG. La Figura 1A muestra un dibujo esquemático de la ubicación, mientras que la Figura 1B presenta la vista hacia el tórax después de la eliminación del paquete corazón-pulmón. Los músculos longus colli izquierdo y derecho medial del SG y la caja torácica son puntos de referencia importantes para la orientación. La disección se realiza a lo largo de las líneas punt...

Watch this Scientific Journal Video about Location, Dissection, and Analysis of the Murine Stellate Ganglion at JoVE.com

Location, Dissection, and Analysis of the Murine Stellate Ganglion

Los cambios fisiopatológicos en el sistema nervioso autónomo cardíaco, especialmente en su rama simpática, contribuyen al inicio y mantenimiento de las arritmias ventriculares. En el actual protocolo, mostramos cómo caracterizar los ganglios estrellado murine para mejorar la comprensión de los procesos moleculares y celulares subyacentes.

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 ...

location-dissection-and-analysis-of-the-murine-stellate-ganglion

Research

JoVE Journal

Medicine

9.6K vistas.  EVK Düsseldorf, cNEP, cardiac Neuro- and Electrophysiology Research Consortium. Los cambios fisiopatológicos en el sistema nervioso autónomo cardíaco, especialmente en su rama simpática, contribuyen al inicio y mantenimiento de las arritmias ventriculares. En el actual protocolo, mostramos cómo caracterizar los ganglios estrellado murine para mejorar la comprensión de los procesos moleculares y celulares subyacentes.

Video: Location, Dissection, and Analysis of the Murine Stellate Ganglion

A Method for Murine Islet Isolation and Subcapsular Kidney Transplantation

Since the early pioneering work of Ballinger and Reckard demonstrating that transplantation of islets of Langerhans into diabetic rodents could normalize their blood glucose levels, islet transplantation has been proposed to be a potential treatment for type 1 diabetes 1,2. More recently, advances in human islet transplantation have further strengthened this view 1,3. However, two major limitations prevent islet transplantation from being a widespread clinical reality: (a) the requirement for large numbers of islets per patient, which severely reduces the number of potential recipients, and (b) the need for heavy immunosuppression, which significantly affects the pediatric population of patients due to their vulnerability to long-term immunosuppression. Strategies that can overcome these limitations have the potential to enhance the therapeutic utility of islet transplantation.
Islet transplantation under the mouse kidney capsule is a widely accepted model to investigate various strategies to improve islet transplantation. This experiment requires the isolation of high quality islets and implantation of islets to the diabetic recipients. Both procedures require surgical steps that can be better demonstrated by video than by text. Here, we document the detailed steps for these procedures by both video and written protocol. We also briefly discuss different transplantation models: syngeneic, allogeneic, syngeneic autoimmune, and allogeneic autoimmune.

Transplantation of isolated islets has been proposed to be a potential treatment for type 1 diabetes. Here we describe a method to isolate islets from mouse pancreata and transplant them to the subcapsular space of the kidney.

Un método para el aislamiento de los islotes murinos y Trasplante Renal subcapsular

Since the early pioneering work of Ballinger and Reckard demonstrating that transplantation of islets of Langerhans into diabetic rodents could normalize ...

Investigación

Medicina

Dissection of Human Vitreous Body Elements for Proteomic Analysis

The vitreous is an optically clear, collagenous extracellular matrix that fills the inside of the eye and overlies the retina. 1,2 Abnormal interactions between vitreous substructures and the retina underlie several vitreoretinal diseases, including retinal tear and detachment, macular pucker, macular hole, age-related macular degeneration, vitreomacular traction, proliferative vitreoretinopathy, proliferative diabetic retinopathy, and inherited vitreoretinopathies. 1,2 The molecular composition of the vitreous substructures is not known. Since the vitreous body is transparent with limited surgical access, it has been difficult to study its substructures at the molecular level. We developed a method to separate and preserve these tissues for proteomic and biochemical analysis. The dissection technique in this experimental video shows how to isolate vitreous base, anterior hyaloid, vitreous core, and vitreous cortex from postmortem human eyes. One-dimensional SDS-PAGE analyses of each vitreous component showed that our dissection technique resulted in four unique protein profiles corresponding to each substructure of the human vitreous body. Identification of differentially compartmentalized proteins will reveal candidate molecules underlying various vitreoretinal diseases.

This video shows an effective technique for differentiating and dissecting the various semi-transparent structures of the human vitreous body in post mortem eyes.

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 ...

Quantitative Analysis and Characterization of Atherosclerotic Lesions in the Murine Aortic Sinus

Atherosclerosis is a disease of the large arteries and a major underlying cause of myocardial infarction and stroke. Several different mouse models have been developed to facilitate the study of the molecular and cellular pathophysiology of this disease. In this manuscript we describe specific techniques for the quantification and characterization of atherosclerotic lesions in the murine aortic sinus and ascending aorta. The advantage of this procedure is that it provides an accurate measurement of the cross-sectional area and total volume of the lesion, which can be used to compare atherosclerotic progression across different treatment groups. This is possible through the use of the valve leaflets as an anatomical landmark, together with careful adjustment of the sectioning angle. We also describe basic staining methods that can be used to begin to characterize atherosclerotic progression. These can be further modified to investigate antigens of specific interest to the researcher. The described techniques are generally applicable to a wide variety of existing and newly created dietary and genetically-induced models of atherogenesis.

We describe procedures to quantify and characterize atherosclerotic lesions in mouse models using precision sectioning of the aortic sinus and ascending aorta combined with histochemical and immunohistochemical analysis.

Atherosclerosis is a disease of the large arteries and a major underlying cause of myocardial infarction and stroke. Several different mouse models have ...

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 ...

Murine Prostate Micro-dissection and Surgical Castration

Mouse models are used extensively to study prostate cancer and other diseases. The mouse is an excellent model with which to study the prostate and has been used as a surrogate for discoveries in human prostate development and disease. Prostate micro-dissection allows consistent study of lobe-specific prostate anatomy, histology, and cellular characteristics in the absence of contamination of other tissues. Testosterone affects prostate development and disease. Androgen deprivation therapy is a common treatment for prostate cancer patients, but many prostate tumors become castration-resistant. Surgical castration of mouse models allows for the study of castration resistance and other facets of hormonal biology on the prostate. This procedure can be coupled with testosterone reintroduction, or hormonal regeneration of the prostate, a powerful method to study stem cell lineages in the prostate. Together, prostate micro-dissection and surgical castration opens up a multitude of opportunities for robust and consistent research of prostate development and disease. This manuscript describes the protocols for prostate micro-dissection and surgical castration in the laboratory mouse.

This manuscript describes the protocols for prostate micro-dissection and surgical castration in the laboratory mouse. We also depict representative results produced by these protocols. Finally, we discuss the advantages and utilization of these protocols.

Murino de próstata micro-disección y la castración quirúrgica

Mouse models are used extensively to study prostate cancer and other diseases. The mouse is an excellent model with which to study the prostate and has ...

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 ...

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 ...

Robot-assisted Total Mesorectal Excision and Lateral Pelvic Lymph Node Dissection for Locally Advanced Middle-low Rectal Cancer

Since their approval for clinical use, da Vinci surgical robots have shown great advantages in gastrointestinal surgical operations, especially in complex procedures. The high-quality 3-D visual, multijoint arm and natural tremor filtration allow the surgeon to expose and dissect more accurately with minimal invasion. Total mesorectal excision is the standard surgical technique for the treatment of resectable rectal cancer. To reduce the lateral recurrence rate, lateral pelvic lymph node dissection can be performed, as it is a safe and feasible procedure for locally advanced middle-low rectal cancer with a high possibility of metastasis to the lateral lymph nodes. However, the complexity of the anatomic structures and the high postoperative complication rate limit its application. Recently, several surgeons have increasingly used robotic techniques for total mesorectal excision and lateral pelvic lymph node dissection. Compared with open and laparoscopic surgery, the robotic technique has several advantages, such as less blood loss, fewer blood transfusions, minimal trauma, shorter postoperative hospitalization, and quicker recovery. A robotic approach is generally regarded as a reasonable alternative for complicated procedures such as lateral pelvic lymph node dissection, although there are a limited number of high-quality prospective randomized controlled studies reporting direct evidence. Here, we provide the detailed steps of robot-assisted total mesorectal excision and lateral pelvic lymph node dissection performed at the First Affiliated Hospital of Xi&#39;an Jiaotong University.

The robotic technique described herein aims to detail a stepwise approach to robot-assisted total mesorectal excision and lateral pelvic lymph node dissection for locally advanced (T3/T4) rectal cancer located below the peritoneal reflection.

Escisión mesorrectal total asistida por robot y disección lateral de ganglios linfáticos pélvicos para el cáncer de recto medio-bajo localmente avanzado

Since their approval for clinical use, da Vinci surgical robots have shown great advantages in gastrointestinal surgical operations, especially in complex ...

Robotic Left Hepatectomy using Indocyanine Green Fluorescence Imaging for an Intrahepatic Complex Biliary Cyst

Biliary cysts (BC) are rare congenital dilatations of intra- and extrahepatic parts of the biliary tract and bear a significant risk of carcinogenesis. Surgery is the cornerstone treatment for patients with BC. While total BC excision and Roux-Y hepaticojejunostomy is the treatment method of the choice in patients with extrahepatic BC (i.e., Todani I-IV), patients with intrahepatic BC (i.e., Todani V) benefit the most from a surgical liver resection. In recent years, minimally invasive liver surgery (MILS) including robotic MILS has gained more acceptance as a feasible, safe, and effective procedure for the treatment of both benign and malignant indications. Robotic major MILS is still considered technically demanding and a detailed description of the technical approach during robotic major MILS has only been limitedly discussed in the literature. The current article describes the main steps for a robotic left hepatectomy in a patient with a large BC Todani Type V. The patient is in French position with 5 trocars placed (4 robotic, 1 laparoscopic assistant). After mobilizing the left hemiliver, the left and right hepatic artery are dissected carefully followed by a cholecystectomy. Intraoperative ultrasound is performed to confirm localization and margins of the BC. The Left hepatic artery and left portal vein are isolated, clipped, and divided. Indocyanine green (ICG) fluorescence imaging is used regularly during the entire procedure to visualize and confirm biliary tract anatomy and the BC. Parenchymal transection is performed with robotic cautery hook for the superficial part and robotic cautery spatula, bipolar cautery, and vessel sealer for the deeper parenchyma. The postoperative course was uncomplicated. A robotic left hepatectomy is technically demanding, yet a feasible and safe procedure. ICG-fluorescence imaging aids in delineating the BC and bile duct anatomy. Further, comparative studies are needed to confirm clinical benefits of robotic MILS for benign and malignant indications.

Robotic liver surgery has gained more acceptance as a feasible, safe, and effective procedure for the treatment of both benign and malignant indications. However, robotic left hepatectomy is still technically demanding. We describe our surgical technique of a robotic left hepatectomy using indocyanine green fluorescence imaging for a large biliary cyst.

Hepatectomía izquierda robótica con imágenes de fluorescencia verde de indocianina para un quiste biliar complejo intrahepático

Biliary cysts (BC) are rare congenital dilatations of intra- and extrahepatic parts of the biliary tract and bear a significant risk of carcinogenesis. ...