Il finanziamento è stato fornito da Lotta per la vista. I tessuti sono stati ottenuti dalla Banca degli occhi Lions Iowa.

1. Segmento dissezione anteriore. <ol><li> La cornea viene rimosso facendo prima un&#39;incisione al limbus, in camera anteriore con una lama di 15 ° (Figura 1). Poi una lama ricurva di cornea-sclerale forbici viene inserito nella camera anteriore. Circonferenziale tagli sono fatti a cornea, appena anteriormente al limbus. </li><li> 0,12 Colibri pinze e forbici Westcott sono usati per tagliare l&#39;iride circonferenza anteriore del corpo ciliare. </li><li> Il nucleo dell&#39;obiettivo sia stato rimosso con pinza 0,12 Colibri o una lama di 15 ° (supersharp) e la corteccia e la capsula con le pinze. </li></ol> 2. Nucleo aspirazione vitreo. <ol><li> Un 23-gauge a 5-cc siringa è inserita nella metà del vitreo (Figura 1). </li><li> Circa 1 ml o più di nucleo vitreo è delicatamente aspirato. </li><li> Il campione viene quindi posto in una provetta per microcentrifuga e poi in azoto liquido. </li></ol> 3. Anteriore Hyaloid Dissection. <ol><li> Il hyaloid anteriore è visto come un semi-anello trasparente regolando la luce incidente da una luce microscopio collo d&#39;oca. Aspirazione precedente del nucleo vitreo separa il hyaloid anteriore da altre strutture per una più facile identificazione. </li><li> Il foglio di tessuto elastico viene accuratamente tirato fuori dal corpo ciliare con Colibri o pinze spogliatoio e tagliato con le forbici Vännäs. Questa manovra viene ripetuta. </li><li> Il campione viene quindi posto in una provetta per microcentrifuga e poi in azoto liquido. </li></ol> 4. Base Dissection vitreo. <ol><li> Utilizzando 0,12 pinze per stabilizzare l&#39;oculare, forbici Westcott sono usati per &quot;fiore&quot; l&#39;occhio facendo quattro equidistanti antistress tagli dal corpo ciliare verso la testa del nervo ottico. </li><li> La pars plicata del corpo ciliare (Figura 2) viene rimosso da ciascuno dei quadranti con le forbici Westcott. </li><li> La base del vitreo viene poi afferrato su entrambi i lati della serrata con una pinza il pars plana e la retina (Figura 2). </li><li> Trazione continua sulla base del vitreo è applicato con le pinze. Taglio sequenziale con le forbici Westcott estratti di un semi-trasparente tessuto con un aspetto &quot;filo di perle&quot;, come viene asportato. </li><li> Il campione viene quindi posto in una provetta per microcentrifuga e poi in azoto liquido. </li></ol> 5. Cortex rimozione vitreo. <ol><li> La pars plana è asportato da ciascuno dei quadranti Westcott con le forbici, tagliando il foglio di circa 3 mm posteriormente al serrata (Figura 2). </li><li> Tra i volantini dei tessuti, la corteccia vitreo è visualizzato come una pellicola semi trasparente. </li><li> Il film elastico può essere afferrato con pinze 0,12 o detenuti da adesione ad una Weck-Cel spugna chirurgica. La corteccia vitreo è poi tirato via dalla retina e asportato con le forbici Westcott (Figura 1). </li><li> Il campione è posto in una provetta per microcentrifuga e poi in azoto liquido. Tutti i campioni sono conservati a -80 ° C fino al momento utilizzato per la sperimentazione. </li></ol> 6. Rappresentante Risultati Campioni di tessuto possono essere elaborati da una varietà di metodi per esperimenti specifici. Nel nostro caso, i campioni sono stati sottoposti ad analisi delle proteine ​​mediante SDS-PAGE (Figura 3). <img src="/files/ftp_upload/2455/2455fig1.jpg" alt="Figura 1" /> Figura 1. Vista in sezione trasversale dell&#39;occhio umano raffigurante sottostrutture diverse del corpo vitreo. Il vitreo più anteriore è un sottile strato di collagene chiamato hyaloid anteriore. Il nucleo vitreo comprende l&#39;intera regione centrale del corpo vitreo. Questa porzione del vitreo è più acquoso in contrasto con la base del vitreo, che è abbastanza viscoso per essere afferrato con pinze ed è saldamente attaccato al corpo ciliare e la retina sottostante. Che comprende il nucleo vitreo è un guscio molto sottile collagene chiamata corteccia vitreo. <img src="/files/ftp_upload/2455/2455fig2.jpg" alt="Figura 2" /> Figura 2. Anatomia base del vitreo. La base del vitreo è un semi-trasparente sottostruttura del corpo vitreo situato lungo la serrata (frecce bianche), che è la linea di demarcazione che separa il corpo ciliare e la retina. Il margine anteriore della base del vitreo si estende su la pars plana (linea bianca) del corpo ciliare. Il bordo posteriore della base del vitreo si estende 2-3-mm posteriormente al serrata (trattino bianco). Per accise base del vitreo, pinze sono utilizzati per afferrare il tessuto e tirarlo via dal corpo ciliare e la retina sottostante. Una volta elevato, forbici Westcott sono usati per tagliare lungo la base. <img src="/files/ftp_upload/2455/2455fig3.jpg" alt="Figura 3" /> Figura 3. Monodimensionale SDS-PAGE di elementi del corpo vitreo. Concentrazioni di proteine ​​totali del hyaloid anteriore, base del vitreo, vitnucleo reous, e la corteccia vitreo sono stati 11,24, 20.1, 16,61, 14,24 mg / ml, rispettivamente. Elettroforesi su gel è stata effettuata a 200 kV per 45 minuti, colorati con Flamingo (Bio-Rad), e visualizzati utilizzando un sistema VersaDoc Imaging (Bio-Rad). I profili per i diversi tessuti mostrano diversi gruppi simili, indicando sia conservata o cross-contaminazione delle proteine, così come unica band (asterisco), indicando proteine ​​differenzialmente localizzate.

<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>

Nessun conflitto di interessi dichiarati.

Il corpo vitreo è un gel semitrasparente cui composizione molecolare è poco conosciuta, soprattutto a livello delle sue sottostrutture: la base del vitreo, core, corteccia, e hyaloid anteriore. Il nucleo vitreo contiene collagene II, V, IX e XI, insieme con proteoglicani condroitin solfato, solfato di eparan proteoglicani e acido ialuronico. 1,2 biomarcatori proteine ​​nel nucleo vitreo sono stati associati a malattie come la retinopatia diabetica. 3-5 Come queste proteine ​​sono differenzialmente espressi in ciascuna delle sottostrutture, e in molti casi l&#39;identità specifica proteina, non sono noti. Questi dettagli può dare qualche idea per l&#39;origine delle proteine ​​associate a specifiche patologie vitreo-retinica e contribuire a future terapie target. Nonostante l&#39;ottimo intervallo post mortem per la dissezione dei tessuti non è stata determinata, la degradazione delle proteine ​​possono influenzare gli esperimenti a valle. Per esempio, l&#39;immunoistochimica è influenzato in 12 ore gli occhi post mortem ed alcune specifiche attività enzimatiche può essere ridotta entro poche ore (osservazione inedita). Tutti i tessuti in questo studio sono stati raccolti tra 2 e 8 ore di morte senza significativi cambiamenti nell&#39;espressione delle proteine ​​o idoneità per analsyis proteomica. Il metodo di congelamento azoto liquido di conservazione viene scelto più di fissaggio per evitare piccoli cambiamenti nella struttura delle proteine ​​causata da fissativo reticolazione, che è stato dimostrato in altri tessuti da LC-MS/MS. 6 studi di proteomica dipenderà dalla capacità di precisione sezionare i diversi compartimenti del vitreo, come dimostrato in questo esperimento video. Abbiamo convalidato la tecnica di dissezione con 1-dimensionale SDS-PAGE. Come i nostri risultati suggeriscono, ci sono proteine ​​differenzialmente espressi nelle diverse sottostrutture corpo vitreo. Identificare queste proteine ​​fornirà una comprensione più dettagliata di compartimentazione vitreo.

<table border="1"><tbody><tr><td> Nome </td><td> Azienda </td><td> Numero di catalogo </td></tr><tr><td> 0,12 forcipe </td><td> Storz oftalmica </td><td> E1502 </td></tr><tr><td> 5-cc siringa </td><td> Becton Dickinson- </td><td> 309603 </td></tr><tr><td> Pinza diritta spogliatoio con dentellature </td><td> Storz oftalmica </td><td> E1400 </td></tr><tr><td> 23 gauge </td><td> Becton Dickinson- </td><td> 305145 </td></tr><tr><td> Colibri forcipe </td><td> Storz oftalmica </td><td> 2 / 132 </td></tr><tr><td> Castroviejo forbici angolata corneale </td><td> Storz oftalmica </td><td> E3223 </td></tr><tr><td> Vännäs Forbici capsulotomia curvo </td><td> Storz oftalmica </td><td> E3387 </td></tr><tr><td> Weck-Cel chirurgico lance </td><td> Medtronic </td><td> 0008680 </td></tr><tr><td> Forbici Tenotomia Westcott curvo </td><td> Storz oftalmica </td><td> E3320 </td></tr></tbody></table>

dissection of human vitreous body elements for proteomic analysis

Il vitreo è una otticamente chiaro, matrice di collagene extracellulare che riempie l&#39;interno dell&#39;occhio e sovrappone la retina. 1,2 interazioni anomale tra sottostrutture vitreo e la retina alla base di diverse patologie vitreo-retinica, tra cui rottura retinica e distacco, pucker maculare, foro maculare, età degenerazione maculare, la trazione vitreomacular, proliferazione vitreoretinica, retinopatia diabetica proliferativa, e vitreoretinopathies ereditato. 1,2 La composizione molecolare delle sottostrutture vitreo non è noto. Poiché il corpo vitreo è trasparente con limitato accesso chirurgico, è stato difficile studiare le sue sottostrutture a livello molecolare. Abbiamo sviluppato un metodo per separare e conservare questi tessuti per l&#39;analisi proteomica e biochimica. La tecnica di dissezione in questo video sperimentale mostra come isolare base del vitreo, hyaloid anteriore, centrale vitreo, e la corteccia vitreo da occhi umani post-mortem. Monodimensionale SDS-PAGE analisi di ogni componente vitreo ha dimostrato che la nostra tecnica dissezione portato a quattro profili proteici uniche corrispondenti a ciascuna sottostruttura del corpo vitreo umano. Identificazione di proteine ​​differenzialmente compartimenti rivelerà molecole candidate sottostante varie malattie vitreo-retinica.

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

Questo video mostra una tecnica efficace per differenziare e sezionare le varie semitrasparente strutture del corpo vitreo in post mortem occhi.

Dissezione di Human Elements corpo vitreo per l&#39;analisi proteomica

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 Views.  University of Iowa. Questo video mostra una tecnica efficace per differenziare e sezionare le varie semitrasparente strutture del corpo vitreo in post mortem occhi.

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.

Tutta vitreo Umore dissezione per l&#39;analisi Vitreodynamic

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

Ricerca

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-capture Microdissezione di Human prostatica Epitelio per l&#39;analisi di RNA

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.

Dissezione del Pancreas del Mouse per l'analisi istologica e Profiling metabolico

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.

Dissezione e cultura Explant della Allantois murino per l'analisi In Vitro di allantoico allegato

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.

Analisi metabolica ingabbiamento in topi alta Fed grasso e della composizione corporea

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.

Analisi quantitativa di Micro-CT di Aortopathy in un modello murino di indotta da β-aminopropionitrile aneurisma e dissezione

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.

Tutto il corpo e quantificazione regionale dell'essere umano attivo marrone tessuto adiposo con 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.

Segmentazione e misurazione lineare per l'analisi della composizione corporea mediante Slice-O-Matic e 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.

Posizione, dissezione e analisi del Ganglio Stellato Murine

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 metodo non invasivo per generare il modello di lesione cartilaginea intra-articolare indotta dal carico ciclico del ginocchio di ratto

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