The authors acknowledge the following funding sources, Whittier Foundation, Harrington Foundation, National Institutes of Health and Research to Prevent Blindness.

Todos os olhos enucleados foram obtidos a partir de um matadouro e todos os experimentos foram realizados de acordo com as leis de biossegurança institucionais. <ol><li> Fixe olho enucleado para baixo em uma superfície. Fazer isso colocando os pinos do tecido através do excesso de tecido em torno do olho e prendendo-o para baixo em uma placa de isopor. </li><li> Dissecar e retire conjuntiva perilimbal de olho. <ol><li> Use uma pinça fina (0,3 fórceps) e microtesoura (tesoura Westcott) para inciso conjuntiva no limbo e sem rodeios dissecá-lo fora da esclera. Cortar a conjuntiva ao longo do limbo como a dissecção romba prossegue para permitir posterior dissecação. </li><li> Remover conjuntiva a toda a volta do olho (360 graus) para expor tanto quanto possível esclera. NOTA: É útil para deixar uma pequena quantidade de conjuntiva para facilitar a fixação do olho com os pinos de uma pinça cirúrgica ou durante o resto do procedimento. </li></ol></li><li> Criar um retalho escleral de espessura total ao longo de umlado do olho (lâmina de bisturi 69). <ol><li> Adicione uma ~ 5 milímetros incisão escleral paralela ao limbo e 3 mm posterior ao limbo cortando suavemente para dentro da esclerótica com um bisturi (lâmina de bisturi 11) até coróide pigmentadas é visível. Tenha cuidado para não inciso a própria coróide. </li><li> Cuidadosamente dissecar ao longo do plano entre a esclera e coróide, ou com tesouras (utilizando uma dissecção romba) ou com o bisturi, até que seja possível aumentar o espaço de potencial entre estes tecidos, empurrando suavemente para dentro da coróide. </li><li> Fazer uma outra incisão escleral perpendicular à primeira incisão escleral com microtesouras afiadas, criando uma incisão em forma de T. </li><li> Em seguida, continuar a dissecar bruscamente de um modo circunferencial para remover o tecido a partir da coróide esclerótica subjacente e empurre o coróide longe da esclera como mencionado acima. </li><li> Ampliar a retalho escleral, conforme necessário. <ol><li> Suavemente sem corte dissecar a coróide longe da escleradurante todo o processo. </li><li> Aumentar a aba até que a incisão feita perpendicularmente ao limbo atinge o nervo óptico e a incisão feita paralela ao limbo é, pelo menos, um terço da circunferência do olho (45 °). </li><li> Use a retalho escleral como fonte de alavancagem para manipular o olho durante a dissecção romba. O contra-tração na aba torna mais fácil para dissecção romba. </li></ol></li><li> Repita o passo anterior no outro retalho escleral. </li></ol></li><li> Cortaram os retalhos esclerais para expor uma grande área de coróide. </li><li> Continuar a incisão feita no passo 3 em torno da circunferência do olho (360 graus). </li><li> Remover o tecido coróide-restante retina. <ol><li> Dentro da região exposta, use um algodão-ponta para escovar suavemente o tecido coróide-retina restante. Alternativamente, agarram delicadamente o tecido com uma pinça e retire-a do corpo vítreo subjacente. </li><li> Se for necessário fazer uma incisão na coróide starting do nervo óptico. Em seguida, retire a coróide suavemente para expor a retina eo córtex vítreo. </li></ol></li><li> Continuar sem corte dissecar a coróide longe da esclera e, em seguida, retire a coróide obter todo o vítreo intacta. </li><li> Utilizar o aro escleral fixada na córnea para posicionar todo o vítreo em local desejado. Neste ponto, o vítreo é ligada à esclerótica anterior e lente. </li><li> Blunt dissecar o vítreo a partir do interior da esclerótica, à volta da lente, a remoção de todos esclera. </li><li> Use uma ferramenta sem corte para colher a lente de distância do vítreo se for necessário. Use o exemplo para várias experiências vitreodynamic. <ol><li> Colocar a amostra num copo de vidro com uma área superficial conhecida exposta ao ar. Coloque sonda sensível ao oxigénio no bordo da amostra. Usar um micro-manipulador para mover a sonda para uma distância conhecida (R) na amostra. </li><li> Use Lei de Fick para obter a equação de difusão teórico. Com a data recolhidas no passo 10.1, obtenha o termo coeficiente de difusão experimental / reação etc. </li></ol></li></ol>

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	<li>Siggers, J. H., Ethier, C. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fluid+Mechanics+of+the+Eye.">Fluid Mechanics of the Eye.</a> Annual Review of Fluid Mechanics. 44 (1), 347-372 (2012).</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=Age-related+changes+in+human+vitreous+structure.">Age-related changes in human vitreous structure.</a> Graefes Arch Clin Exp Ophthalmol. 225 (2), 89-93 (1987).</li><li>Grignolo, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fibrous+components+of+the+vitreous+body.">Fibrous components of the vitreous body.</a> AMA Arch Ophthalmol. 47 (6), 760-774 (1952).</li><li>Gisladottir, S., Loftsson, T., Stefansson, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Diffusion+characteristics+of+vitreous+humour+and+saline+solution+follow+the+Stokes+Einstein+equation.">Diffusion characteristics of vitreous humour and saline solution follow the Stokes Einstein equation.</a> G Graefes Arch Clin Exp Ophthalmol. 247 (12), 1677-1684 (2009).</li><li>Xu, J., Heys, J. J., Barocas, V. H., Randolph, T. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Permeability+and+diffusion+in+vitreous+humor:+implications+for+drug+delivery.">Permeability and diffusion in vitreous humor: implications for drug delivery.</a> Pharm Res. 17 (6), 664-669 (2000).</li><li>Watts, F., Tan, L. E., Wilson, C. G., Girkin, J. M., Tassieri, M., Wright, A. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Investigating+the+micro-rheology+of+the+vitreous+humor+using+an+optically+trapped+local+probe.">Investigating the micro-rheology of the vitreous humor using an optically trapped local probe.</a> Journal of Optics. 16 (1), 015301 (2014).</li><li>Skeie, J. M., Mahajan, V. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dissection+of+human+vitreous+body+elements+for+proteomic+analysis.">Dissection of human vitreous body elements for proteomic analysis.</a> J Vis Exp. (47), e2455 (2011).</li><li>Abdallah, W., Ameri, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Vitreal+oxygenation+in+retinal+ischemia+reperfusion.">Vitreal oxygenation in retinal ischemia reperfusion.</a> Invest Ophthalmol Vis Sci. 52 (2), 1035-1042 (2011).</li><li>Goldenberg, D., Trese, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pharmacologic+vitreodynamics:+what+is+it?+Why+is+it+important.">Pharmacologic vitreodynamics: what is it? Why is it important.</a> Expert Review of Ophthalmology. 3 (3), 273-277 (2008).</li><li>Choonara, Y. E., Pillay, V., Danckwerts, M. P., Carmichael, T. R., du Toit, L. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+review+of+implantable+intravitreal+drug+delivery+technologies+for+the+treatment+of+posterior+segment+eye+diseases.">A review of implantable intravitreal drug delivery technologies for the treatment of posterior segment eye diseases.</a> J Pharm Sci. 99 (5), 2219-2239 (2010).</li><li>Balachandran, R. K., Barocas, V. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Computer+modeling+of+drug+delivery+to+the+posterior+eye:+effect+of+active+transport+and+loss+to+choroidal+blood+flow.">Computer modeling of drug delivery to the posterior eye: effect of active transport and loss to choroidal blood flow.</a> Pharm Res. 25 (11), 2685-2696 (2008).</li><li>Smith, C. a., Newson, T. a., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+framework+for+modeling+ocular+drug+transport+and+flow+through+the+eye+using+micro-CT.">A framework for modeling ocular drug transport and flow through the eye using micro-CT.</a> Phys Med Biol. 57 (19), 6295-6307 (2012).</li><li>Quiram, P. A., Leverenz, V. R., Baker, R. M., Dang, L., Giblin, F. J., Trese, M. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microplasmin-induced+posterior+vitreous+detachment+affects+vitreous+oxygen+levels.">Microplasmin-induced posterior vitreous detachment affects vitreous oxygen levels.</a> Retina. 27 (8), 1090-1096 (2007).</li><li>Shui, Y., Holekamp, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+gel+state+of+the+vitreous+and+ascorbate-dependent+oxygen+consumption:+relationship+to+the+etiology+of+nuclear+cataracts.">The gel state of the vitreous and ascorbate-dependent oxygen consumption: relationship to the etiology of nuclear cataracts.</a> Arch Ophthalmol. 127 (4), 475-482 (2009).</li><li>Burk, S. E., Da Mata, A. P., Snyder, M. E., Schneider, S., Osher, R. H., Cionni, R. 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The authors have nothing to disclose.

Existem dois passos críticos que devem ser cuidadosamente realizados durante a dissecção vítreo. Passo 3, a criação de uma aba escleral de espessura total, é crucial para toda a dissecção. Cuidados devem ser tomados para não cortar na coróide ao criar a espessura total retalho escleral. O outro passo crítico é dissecar afastado da esclera da coróide. Esta etapa deve ser feito com cuidado para evitar a criação de vários buracos na coróide de que o vítreo pode derramar. Existe uma maneira de modificar o...

O objectivo deste método é a de detalhe uma técnica para isolar todo um corpo vítreo, intacta, com o núcleo vítreo e córtex intacto, a partir de um olho de cadáver, para efeitos de análise vitreodynamic. Como o campo da fisiologia vítreo tem crescido, pesquisadores multidisciplinares, tais como mecânica dos fluidos pesquisadores, estão a investigar as propriedades físicas e biomecânicas do vítreo 1. Para este fim, é essencial detalhe uma técnica para isolar todo o, corpo vítreo intacta para ajudar os investigadores multidisciplinares. Sebag et al. 2 e 3 outros realizados inteiras elegantes dissecações de cadáveres vítreas em olhos humanos e mostrou ilustrações dos resultados. No entanto, a técnica utilizada não foi descrito em pormenor e não especialistas não seria capaz de se replicar independentemente do método. Outros estudos têm colhido vítreo de cadáveres olhos usando métodos mais simples, como aspiração ou dissecação parcial,ambas as quais não resultam em conjunto um corpo vítreo, intacta. Gisladottis et al. 4 e Xu et al. 5 investigar permeabilidade no humor vítreo colhida a partir de olhos de cadáveres. No entanto, uma vez que nenhum método de extracção vítreo foi descrito, assumiu-se que eles aspirado do humor vítreo com uma seringa. Watts et al. 6 foi um passo adiante, descrevendo um método de isolamento de coelho humor vítreo com uma técnica cirúrgica. No entanto, este método resulta num isolamento de apenas o núcleo vítreo e não no córtex vítreo. Skeie et al. 7 posteriormente organizou o vítreo em 4 regiões únicas e elegantemente descrito um método para dissecar cada parte para análise. Esta técnica no entanto, não resultar numa vítreo intacto como um todo. A técnica actual foi desenvolvido para facilitar experiências biofísicas que actualmente apenas são realizadas em cadáveres olhos. Os métodos anteriores, como descrito umbove, são limitados porque 1) nenhuma isolar completamente todo o corpo vítreo, núcleo e córtex vítreo 2) colhidas são homogeneizados, 3) estrutura anatômica vítreo não é mantido, ou 4) técnicas de dissecação não são adequadamente detalhados para a replicação por pesquisadores em outros campos . Além disso, devido à opacidade da esclera e coróide, visualização do corpo vítreo é limitado no globo ocular intacto. Isto limita a precisão das medições de viabilidade e que podem ser feitas dentro de todo o olho. Além disso, as estruturas anatómicas circundantes do vítreo pode confundir o estudo das propriedades bioquímicas e físicas do vítreo. Nos últimos anos, o corpo vítreo da ciência tem crescido muito e não há razão para acreditar que todo o corpo vítreo tem propriedades diferentes do que suas partes individuais. Há um crescente interesse em investigar as propriedades físicas, biomecânicos e químicas do vítreo para vitreodynamics research, que tem aplicações na medicina clínica, tais como a entrega de drogas, oxigenação intravítrea 8 e vitrectomia. Vitreodynamics farmacológicos, que utiliza agentes farmacológicos para manipular o vítreo, pode ser utilizada para melhorar os resultados de vitrectomia 9. As propriedades biomecânicas são utilizados para modelar o fluxo de fluido vítreo, que pode ser usado para melhorar as tecnologias de entrega de drogas intravítrea 10-12. Propriedades físicas de vários segmentos da vítreo são cruciais para a compreensão de transporte de oxigênio vítreo-retiniana 13. A técnica de dissecção do vítreo proposto pode ser utilizado para estudar diversas propriedades do humor vítreo intacta. Ele permite que os experimentos de bancada a ser feito em, corpos vítreos intactas inteiros com melhor visualização. Em resumo, os métodos actuais para o estudo do vítreo ou não são adequadamente descritos, ou resultar em um isolamento completo do núcleo vítreo e no córtex. Portanto, existe uma necessidade de executar experiments em um modelo de olho transparente, mantendo a anatomia do vítreo existente no olho cadáver.

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			<td>0.3 forceps</td>
			<td>Storz Opthalmics</td>
			<td>E1793</td>
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			<td>Westcott Tenotomy Scissors Curved Right</td>
			<td>Storz Opthalmics</td>
			<td>E3320 R</td>
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			<td>Scalpel Blade, #11, for #3 Handle</td>
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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.

Seguindo o protocolo irá levar a uma dissecção do vítreo bem sucedida com o núcleo e o córtex (Figura 3) intactos. Isto é evidente a partir dos pedaços residuais de retina aderiram ao córtex vítreo. Todo intacta humor vítreo pode ser usado de várias maneiras para experiências vitreodynamic específicos. No nosso caso, a taxa de difusão do oxigénio no humor vítreo intacto e é constante de tempo correspondente foi estudada (Figura 2). Vítreo que foi dissecado (núcleo e ...

Watch this Scientific Journal Video about Whole Vitreous Humor Dissection for Vitreodynamic Analysis at JoVE.com

Whole Vitreous Humor Dissection for Vitreodynamic Analysis

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.

Whole humor vítreo Dissecção de Análise Vitreodynamic

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

whole-vitreous-humor-dissection-for-vitreodynamic-analysis

Research

JoVE Journal

Medicine

11.3K visualizações.  University of Southern California. 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.

Vídeo: Whole Vitreous Humor Dissection for Vitreodynamic Analysis

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.

Dissecção da Human Elements Corpo Vítreo para Análise Proteômica

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

Medicina

Protocol for Long Duration Whole Body Hyperthermia in Mice

Hyperthermia is a general term used to define the increase in core body temperature above normal. It is often used to describe the increased core body temperature that is observed during fever. The use of hyperthermia as an adjuvant has emerged as a promising procedure for tumor regression in the field of cancer biology. For this purpose, the most important requirement is to have reliable and uniform heating protocols. We have developed a protocol for hyperthermia (whole body) in mice. In this protocol, animals are exposed to cycles of hyperthermia for 90 min followed by a rest period of 15 min. During this period mice have easy access to food and water. High body temperature spikes in the mice during first few hyperthermia exposure cycles are prevented by immobilizing the animal. Additionally, normal saline is administered in first few cycles to minimize the effects of dehydration. This protocol can simulate fever like conditions in mice up to 12-24 hr. We have used 8-12 weeks old BALB/Cj female mice to demonstrate the protocol.

This paper describes a protocol for whole body hyperthermia in mice that can stimulate fever like conditions up to 12-24 hr.

Protocolo para Longa Duração Hipertermia corpo inteiro em Ratos

Hyperthermia is a general term used to define the increase in core body temperature above normal. It is often used to describe the increased core body ...

3D Whole-heart Myocardial Tissue Analysis

Cardiac regenerative therapies aim to protect and repair the injured heart in patients with ischemic heart disease. By injecting stem cells or other biologicals that enhance angio- or vasculogenesis into the infarct border zone (IBZ), tissue perfusion is improved, and the myocardium can be protected from further damage. For maximum therapeutic effect, it is hypothesized that the regenerative substance is best delivered to the IBZ. This requires accurate injections and has led to the development of new injection techniques. To validate these new techniques, we have designed a validation protocol based on myocardial tissue analysis. This protocol includes whole-heart myocardial tissue processing that enables detailed two-dimensional (2D) and three-dimensional (3D) analysis of the cardiac anatomy and intramyocardial injections. In a pig, myocardial infarction was created by a 90-min occlusion of the left anterior descending coronary artery. Four weeks later, a mixture of a hydrogel with superparamagnetic iron oxide particles (SPIOs) and fluorescent beads was injected in the IBZ using a minimally-invasive endocardial approach. 1 h after the injection procedure, the pig was euthanized, and the heart was excised and embedded in agarose (agar). After the solidification of the agar, magnetic resonance imaging (MRI), slicing of the heart, and fluorescence imaging were performed. After image post-processing, 3D analysis was performed to assess the IBZ targeting accuracy. This protocol provides a structured and reproducible method for the assessment of the targeting accuracy of intramyocardial injections into the IBZ. The protocol can be easily used when the processing of scar tissue and/or validation of the injection accuracy of the whole heart is desired.

This protocol describes a novel method for the 3D comparison of whole-heart myocardial tissue with MRI. This is designed for the accurate assessment of intramyocardial injections in the infarct border zone of a chronic porcine model of myocardial infarction.

Análise Tecidos do Miocárdio Whole-coração 3D

Cardiac regenerative therapies aim to protect and repair the injured heart in patients with ischemic heart disease. By injecting stem cells or other ...

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.

Dissecação do pâncreas Mouse para análise histológica e perfis metabólicos

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

Discovering Middle Ear Anatomy by Transcanal Endoscopic Ear Surgery: A Dissection Manual

The middle ear is located in the center of the temporal bone and bears a highly complex anatomy. The recently introduced exclusively endoscopic transcanal approach to the middle ear is a minimally invasive technique sparing the bone and mucosa of the mastoid bone, since the middle ear is accessed through the external auditory canal. This emerging method has several advantages over the traditional (microscopic) approaches to the middle ear such as the panoramic wide-angle views of the anatomy, the possibility to approach and magnify tiny structures, and the possibility of looking around the corner using angled endoscopes.
The cadaveric dissection method presented here consists of an overview on the technical requirements and a precise description of a step-by-step protocol to discover the anatomy of the middle ear. Each step and anatomical structure is carefully described in order to provide a comprehensive guide to endoscopic ear anatomy. In our opinion, this is particularly important to any novice in endoscopic ear surgery as it provides thorough anatomical knowledge and may improve surgical skills.

The aim of this article is to describe the methodology of exclusively endoscopic cadaveric middle ear dissection. Moreover, we aim to provide a comprehensive guide to endoscopic middle ear anatomy.

Descobrindo a anatomia do ouvido médio por cirurgia endoscópica de ouvido de Transcanal: um Manual de dissecação

The middle ear is located in the center of the temporal bone and bears a highly complex anatomy. The recently introduced exclusively endoscopic transcanal ...

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.

Dissecção e cultura de explante de murino alantoide para a análise In Vitro de fixação alantoico

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

Whole-brain Segmentation and Change-point Analysis of Anatomical Brain MRI&#8212;Application in Premanifest Huntington's Disease

Recent advances in MRI offer a variety of useful markers to identify neurodegenerative diseases. In Huntington&#39;s disease (HD), regional brain atrophy begins many years prior to the motor onset (during the &#34;premanifest&#34; period), but the spatiotemporal pattern of regional atrophy across the brain has not been fully characterized. Here we demonstrate an online cloud-computing platform, &#34;MRICloud&#34;, which provides atlas-based whole-brain segmentation of T1-weighted images at multiple granularity levels, and thereby, enables us to access the regional features of brain anatomy. We then describe a regression model that detects statistically significant inflection points, at which regional brain atrophy starts to be noticeable, i.e. the &#34;change-point&#34;, with respect to a disease progression index. We used the CAG-age product (CAP) score to index the disease progression in HD patients. Change-point analysis of the volumetric measurements from the segmentation pipeline, therefore, provides important information of the order and pattern of structural atrophy across the brain. The paper illustrates the use of these techniques on T1-weighted MRI data of premanifest HD subjects from a large multicenter PREDICT-HD study. This design potentially has wide applications in a range of neurodegenerative diseases to investigate the dynamic changes of brain anatomy.

Whole-brain Segmentation and Change-point Analysis of Anatomical Brain MRI&amp;#8212;Application in Premanifest Huntington&#039;s Disease

This paper describes a statistical model for volumetric MRI data analysis, which identifies the &#34;change-point&#34; when brain atrophy begins in premanifest Huntington&#39;s disease. Whole-brain mapping of the change-points is achieved based on brain volumes obtained using an atlas-based segmentation pipeline of T1-weighted images.

Todo cérebro-segmentação e análise de ponto de mudança de anatômico cérebro MRI — aplicação em Premanifest a doença de Huntington

Recent advances in MRI offer a variety of useful markers to identify neurodegenerative diseases. In Huntington&#39;s disease (HD), regional brain atrophy ...

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.

Análise quantitativa de Micro-CT de Aortopathy em um modelo do rato de β-aminopropionitrile-induzida de aneurisma da aorta e dissecação

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

Imaging and Analysis of Oil Red O-Stained Whole Aorta Lesions in an Aneurysm Hyperlipidemia Mouse Model

Apolipoprotein E (Apoe)- or low density lipoprotein receptor (Ldlr)-deficient hyperlipidemic mice are the two most commonly used models for atherosclerosis research. They are used to study the impact of a various genetic factors and different cell types on atherosclerotic lesion formation and as well as test the development of new therapies. Isolation, excision of the whole aorta, and quantification of Oil Red O-stained atherosclerotic lesions are basic morphometric methods used to evaluate atherosclerotic burden. The goal of this protocol is to describe an optimized, step-by-step surgical method to dissect, perfuse-fix, isolate, stain, image and analyze atherosclerotic lesions in mouse aortas with Oil Red O. Because atherosclerotic lesions can form anywhere in the entire aortic tree, this whole aorta Oil Red O staining method has the advantage of evaluating lipid-laden plaques in the entire aorta and all branches in a single mouse. In addition to Oil Red O staining, fresh isolated whole aortas can be used for variety of in vitro and in vivo experiments and cell isolations.

This protocol provides a step-by-step procedure to analyze atherosclerotic burden in mice. Investigators can use this protocol to compare the abundance, location, and size of atherosclerotic lesions in different animals.

Imagem e Análise de Lesões inteiras de aorta manchadas de óleo em um modelo de rato de hiperlipidemia aneurisma

Apolipoprotein E (Apoe)- or low density lipoprotein receptor (Ldlr)-deficient hyperlipidemic mice are the two most commonly used models for ...

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.

Localização, Dissecção e Análise do Gânglio Murine Stellate

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