Finansman Fight for Sight tarafından sağlanan. Dokular Iowa Lions Göz Bankası elde edildi.

1. Ön Segment Diseksiyon. <ol><li> (Şekil 1) 15 ° bir bıçak kullanarak ön odasına limbus bir kesi yaparak kornea kaldırılır. Sonra bir bıçak, kavisli kornea-skleral makas, ön kamara eklenir. Dairesel keser limbus sadece ön kornea yapılır. </li><li> 0.12 Colibri forseps ve Westcott&#39;a makas, iris, siliyer cisim dairesel ön kesmek için kullanılır. </li><li> Lens çekirdeği 0,12 Colibri forseps veya 15 ° bıçağı (supersharp) ve forseps ile korteks ve kapsül ile kaldırılır. </li></ol> 2. Vitreus Çekirdek Aspirasyon. <ol><li> 5 cc şırınga, 23-gauge iğne sokulur orta vitreus (Şekil 1). </li><li> Yaklaşık 1 ml veya daha fazla vitreus çekirdek nazikçe aspire edilir. </li><li> Örnek sonra, bir mikrofuge&#39;de tüp içinde ve daha sonra sıvı azot içine yerleştirilir. </li></ol> 3. Anterior Hyaloid Diseksiyon. <ol><li> Anterior hyaloid bir kaz boynu mikroskop ışık ışık ayarlayarak yarı saydam bir halkası olarak görülüyor. Önceki vitreus çekirdek aspirasyonu, diğer yapılardan daha kolay tespit edilebilmesi için ön hyaloid ayırır. </li><li> Colibri veya giyinme forseps ile siliyer cisim elastik doku sayfasını dikkatlice çekti ve Vannas makas ile kesilir. Bu manevra tekrarlanır. </li><li> Örnek sonra, bir mikrofuge&#39;de tüp içinde ve daha sonra sıvı azot içine yerleştirilir. </li></ol> 4. Vitreus Base Diseksiyon. <ol><li> Göz fincan stabilize 0.12 forseps kullanma, Westcott makas, siliyer cisim, optik sinir başı doğru dört eşit aralıklı stres giderici kesikler yaparak &quot;çiçek&quot; göz için kullanılır. </li><li> Siliyer cisim (Şekil 2) pars Plicata Westcott makas kullanarak kadranın her kaldırılır. </li><li> Vitreus tabanı, pars plana ve retina (Şekil 2) üzerinde forseps ile ora serrata her iki tarafta kavradı. </li><li> Vitreus tabanı sürekli çekiş forseps uygulanır. Eksize olarak Sıralı Westcott makas ile kesme &quot;inci string&quot; görünümü ile yarı saydam bir doku ayıklar. </li><li> Örnek sonra, bir mikrofuge&#39;de tüp içinde ve daha sonra sıvı azot içine yerleştirilir. </li></ol> 5. Vitreus Cortex Kaldırma. <ol><li> Pars plana ora serrata (Şekil 2) posterior yaklaşık 3 mm broşürün keserek Westcott makas ile kadranın her eksize edilir. </li><li> Vitreus korteks doku broşürler arasında, yarı saydam bir film olarak görüntülenmiştir. </li><li> Elastik film ya 0.12 forseps ile kavradı veya Weck-Cel cerrahi sünger bağlılık tarafından düzenlenen. Vitreus korteksi sonra retinanın uzak çekti ve Westcott makas (Şekil 1) ile birlikte çıkarıldı. </li><li> Örnek bir mikrofuge&#39;de tüp ve daha sonra sıvı azot içine yerleştirilir. Tüm numuneler deney için kullanılan -80 ° C kadar saklanır. </li></ol> 6. Temsilcisi Sonuçlar Doku örnekleri özel deneyler için çok çeşitli yöntemler tarafından işlenebilir. Bizim durumumuzda, örnekler, SDS-PAGE (Şekil 3) protein analizi için sunulmuştur. <img src="/files/ftp_upload/2455/2455fig1.jpg" alt="Şekil 1" /> Şekil 1. Insan gözünün kesitsel görünümü vitreus vücudun farklı altyapılardan resmeden en ön vitreus ön hyaloid denilen ince bir kollajen tabakası. Vitreus çekirdek vitreus vücudun tüm merkez bölgesi oluşur. Vitreus bu bölümü yeterli viskoz forseps ile kavranabilir ve yatan siliyer cisim ve retina sıkıca bağlı olan vitreus tabanı, aksine daha sulu. Vitreus çekirdek yakınınızda vitreus korteks denilen çok ince bir kollajen kabuk. <img src="/files/ftp_upload/2455/2455fig2.jpg" alt="Şekil 2" /> Şekil 2. Vitreus tabanı anatomisi. Vitreus tabanı, siliyer cisim ve retina ayıran hat, ora serrata (beyaz oklar), yanında bulunan vitreus yarı saydam bir altyapı . Vitreus tabanının ön sınır siliyer cisim pars plana (beyaz çizgi) boyunca uzanır. Vitreus tabanının arka sınır ora serrata (beyaz çizgi) posterior 2-3 mm uzanır. Tüketim için vitreus tabanı, forseps doku kavramak ve altta yatan siliyer cisim ve retina uzağa çekmek için kullanılır. Yükselmiş, sonra Westcott makas tabanı boyunca kesmek için kullanılır. <img src="/files/ftp_upload/2455/2455fig3.jpg" alt="Şekil 3" /> Şekil 3. Tek boyutlu Vitreus Gövde Elemanları SDS-PAGE ön hyaloid total protein konsantrasyonları, vitreus tabanı, vitreous çekirdek, ve vitreus korteksi sırasıyla 11.24, 20.1, 16.61, 14.24 mg / ml idi. Jel elektroforezi 45 dakika için 200 kV Flamingo (Bio-Rad) ile boyanmış ve bir VersaDoc Görüntüleme sistemi (Bio-Rad) kullanılarak görüntülendi. Farklı dokular için profilleri, farklı yerelleştirilmiş proteinler gösteren korunmuş (yıldız) veya protein çapraz kirlenmesine neden olan, aynı zamanda benzersiz bantları gösteren, benzer pek çok bantları göstermektedir.

<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>

Vitreus tabanı, çekirdek, korteks, anterior hyaloid: vitreus moleküler bileşimini özellikle altyapılardan düzeyinde yeterince anlaşılır bir yarı saydam bir jel. Vitreus çekirdek diyabetik retinopati gibi hastalıklar ile ilişkili olduğu, kondroitin sülfat proteoglikanlar, heparan sülfat proteoglikanların, ve vitreus çekirdek hyaluronan 1,2 Protein biyobelirteçleri ile birlikte kolajenlerin II, V, IX, ve XI içerir. 3-5 nasıl Bu proteinlerin farklı altyapılardan, her ifade ve özel protein kimlikler, birçok durumda bilinmemektedir. Bu ayrıntıları belirli Vitreoretinal hastalıklar ile ilişkili proteinlerin kökenini fikir vermek ve hedef gelecekte tedaviler yardımcı olabilir. Doku diseksiyonu için optimum post mortem aralığı tespit edilmemiş olmasına rağmen, protein bozulması downstream deneyler etkileyebilir. Örneğin, 12 saatlik postmortem gözünde immünohistokimya etkilenir ve bazı özel enzim aktiviteleri (basılmamış gözlem), birkaç saat içinde azalabilir. Bu çalışmada tüm dokuların protein ekspresyonu veya proteomik analsyis uygunluğu önemli değişiklikler olmadan ölüm 2 ve 8 saat arasında toplanmıştır. Korunması sıvı nitrojen dondurma yöntemi fiksatif çapraz bağlama, diğer dokularda LC-MS/MS tarafından kanıtlanmıştır neden protein yapısındaki küçük değişiklikler önlemek için fiksasyon üzerinde seçilen 6 Proteomik çalışmalar doğru bir şekilde yeteneğine bağlıdır Bu video deneyde ortaya vitreus farklı bölmeleri, teşrih. SDS-PAGE 1-boyutlu diseksiyon tekniği kullanılarak valide var. Sonuçlar, farklı çeşitli vitreus altyapılar proteinlerin ifade edilir. Bu proteinlerin belirlenmesi vitreus bölümlendirme daha ayrıntılı bir anlayış sağlayacak.

<table border="1"><tbody><tr><td> Isim </td><td> Şirket </td><td> Katalog Numarası </td></tr><tr><td> 0.12 forseps </td><td> Storz Ophtalmics&#39;in </td><td> E1502 </td></tr><tr><td> 5-cc şırınga </td><td> Becton-Dickinson </td><td> 309603 </td></tr><tr><td> Çentikler ile Düz Giyinme Forseps </td><td> Storz Ophtalmics&#39;in </td><td> E1400 </td></tr><tr><td> 23 iğne </td><td> Becton-Dickinson </td><td> 305145 </td></tr><tr><td> Colibri forseps </td><td> Storz Ophtalmics&#39;in </td><td> 2 / 132 </td></tr><tr><td> Castroviejo açılı kornea makası </td><td> Storz Ophtalmics&#39;in </td><td> E3223 </td></tr><tr><td> Vannas Kavisli kapsülotomi Makas </td><td> Storz Ophtalmics&#39;in </td><td> E3387 </td></tr><tr><td> Weck-Cel cerrahi spears </td><td> Medtronic </td><td> 0008680 </td></tr><tr><td> Westcott Kavisli tenotomisi Makas </td><td> Storz Ophtalmics&#39;in </td><td> E3320 </td></tr></tbody></table>

dissection of human vitreous body elements for proteomic analysis

Vitreus, göz ve retina üzerinde yer içini doldurur optik açık bir kollajenöz ekstrasellüler matriks, vitreus altyapılar ve retina arasında 1,2 Anormal etkileşimleri retina yırtığı ve dekolmanı, maküler pucker, makula deliği dahil olmak üzere birçok vitreoretinal hastalıkların , altında yaşa bağlı makula dejenerasyonu, vitreomaküler çekiş, proliferatif vitreoretinopati, proliferatif diyabetik retinopati ve miras Vitreoretinopatilerde. 1,2 vitreus altyapılardan moleküler kompozisyonu bilinmemektedir. Vitreus sınırlı cerrahi erişim ile şeffaf olduğundan, moleküler düzeyde altyapılar incelemek için zor olmuştur. Proteomik ve biyokimyasal analiz için ayrı ve bu dokuları korumak için bir yöntem geliştirdi. Bu deneysel video diseksiyon tekniği, vitreus tabanı, anterior hyaloid, vitreus çekirdek ve postmortem insan gözü vitreus korteks izole etmek için nasıl gösterir. Her vitreus bileşeni tek boyutlu SDS-PAGE analizleri bizim diseksiyon tekniği, insan vitreus vücudunun her bir altyapı karşılık gelen dört benzersiz protein profilleri yol açtığını gösterdi. Farklı bölümlere proteinlerin belirlenmesi, aday molekülleri çeşitli vitreoretinal hastalıkların altında yatan ortaya koyacaktır.

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

Bu video post-mortem gözünde, insan vitreus vücudunun çeşitli yarı saydam yapıların ayrılması ve diseksiyon için etkili bir teknik gösterir.

Proteomik Analizi İnsan Vitreus Gövde Elemanları diseksiyonu

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

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31.7K Görüntüleme.  University of Iowa. Bu video post-mortem gözünde, insan vitreus vücudunun çeşitli yarı saydam yapıların ayrılması ve diseksiyon için etkili bir teknik gösterir.

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.

Vitreodynamic Analiz Tüm Vitreus Mizah Diseksiyon

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

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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.

RNA Analizi İnsan Prostat Epitelinin Lazer yakalama mikrodiseksiyon

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.

Fare pankreas diseksiyon histolojik analiz ve metabolik profil oluşturma

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.

Diseksiyon ve Explant kültür Allantoic eki Vitro analizi için fare Allantois ve

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.

Vücut kompozisyonu ve metabolik Caging Analizi yüksek yağ Fed farelerde

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.

Aortopathy aort anevrizması β-aminopropionitrile-indüklenen ve diseksiyon, fare modeli nicel mikro-CT Analizi

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

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.

Slice-O-Matic ve Horos Kullanarak Vücut Kompozisyon Analizi için Segmentasyon ve Doğrusal Ölçüm

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.

Murine Stellate Ganglion'un Yeri, Diseksiyonu ve Analizi

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.

Sıçan Dizinin Döngüsel Yüklemeye Bağlı Eklem İçi Kıkırdak Lezyon Modelini Oluşturmak için Non-İnvaziv Bir Yöntem

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

Flow Cytometric Analysis of Biomarkers for Detecting Human Sperm Functional Defects

The conventional semen parameter analysis is widely used to assess male fertility. However, studies have found that ~15% of infertile patients show no abnormalities in conventional semen parameters. Additional technologies are needed to explain the idiopathic infertility and detect subtle sperm defects. Currently, biomarkers of sperm function, including sperm apoptosis, mitochondrial membrane potential (MMP), and DNA damage, reveal sperm physiology at the molecular level and are capable of predicting male fertility.
With flow cytometry (FCM) techniques, each of these markers can be rapidly, accurately, and precisely measured in human semen samples, but time costs substantially increase and results could be obstructed if all the biomarkers need to be tested with a single cytometer. In this protocol, after collection and immediate incubation at 37 &#176;C for liquefication, semen samples were further analyzed for sperm apoptosis using Annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) staining. The MMP was labeled with 5,5&#8242;,6,6&#8242;-tetrachloro-1,1&#8242;,3,3&#8242;-tetraethyl-benzimidazolylcarbocyanine iodide (JC-1) probe, and DNA damage was assessed using the sperm chromatin structure assay (SCSA) with acridine orange (AO) staining. Thus, flow cytometric analysis of sperm function markers can be a practical and reliable toolkit for the diagnosis of infertility and evaluation of sperm function at both bench and bed.

The present protocol provides a practical solution that allows for the measurement of apoptosis, mitochondrial membrane potential, and DNA damage in human sperm with a single cytometer.

İnsan Sperm Fonksiyonel Kusurlarını Tespit Etmek için Biyobelirteçlerin Akış Sitometrik Analizi

The conventional semen parameter analysis is widely used to assess male fertility. However, studies have found that ~15% of infertile patients show no ...