Ringraziamo i molti studenti e borsisti che hanno lavorato su questo modello per migliorarne l&#39;efficacia, e per capire i suoi meccanismi. Un ringraziamento particolare va al dott del. Mary Johnson (University of Maryland-Baltimore), Nitza Goldenberg-Cohen (Schneiderman Childrens Hospital, Petah-Tikva, Israel), Charles Zhang (Einstein Medical College, Bronx, NY), e Valerie Touitou (Hopital Salpetrie, Parigi, Francia). Questo studio è stato finanziato in parte da RO1 EY015304 per SLB.

Questo protocollo è stato approvato dalla University of Maryland Istituzionale Cura animali e del Comitato di utilizzo (IACUC, Baltimore, MD, USA) 1. set-up sperimentale <ol><li> Fare una lente a contatto personalizzato progettato da un grado ottico chiaro circolare 7 mm di diametro plexiglass, di 3 mm di spessore. Tagliare le lenti circolari con un trapano a colonna. Utilizzare una punta da trapano standard per rendere la curva interna, e, infine, lucidare le curve esterne ed interne usando una lucidatrice di lenti a contatto di grana ultrafine (1000/3000). </li><li> Preparare 2,5 mM Rosa Bengala (RB) in pH 7,4 tampone fosfato salino (PBS) in anticipo, filtro sterilizzare con un filtro a siringa da 0,45 micron e conservare 1 ml aliquote a -20 ° C in un contenitore a tenuta di luce fino a 6 mesi. NOTA: L&#39;uso di animali maschi outbred albini, come Sprague Dawley riduce al minimo lo sforzo differenze di risposta dipendono, consente una maggiore facilità di induzione e riduce la variabilità che può verificarsi con estro cycling femminile. </li><li> Impostare la frequenza raddoppiata Neodimio-ittrio granato di alluminio (Fd-YAG) laser medico oftalmico, che genera una luce laser 532 nm. Montare il laser su una lampada a fessura oftalmica Haag-Streit utilizzando un adattatore laser oftalmico standard. Questo dispositivo disponibile in commercio consente la visualizzazione simultanea di applicazione punto dell&#39;occhio e laser dell&#39;animale. Il laser medico ha anche un fascio di puntamento per una corretta messa a fuoco e centratura utilizzando lo stesso formato di punto come l&#39;induzione laser. parametri di potenza laser sono i seguenti: <ol><li> Per ratto induzione rNAION: utilizzare 500 micron punto dimensioni / 50 mW di potenza del laser / 1.000 durata msec / 1.000 ms intervallo. </li><li> Per il mouse induzione rNAION: cambiare la dimensione del punto a 300 micron per disco ottico più piccolo e lasciare gli altri parametri lo stesso come impostazione di ratto. </li><li> Ordinariamente utilizzare un misuratore di potenza laser per assicurare la potenza del laser. </li></ol></li></ol> 2. Procedura sperimentale <ol><li> Accendere la potenza del lasere impostare il parametro del laser appropriata. Riscaldare il laser per almeno cinque minuti prima dell&#39;uso. </li><li> Pesare l&#39;animale per determinare la dose appropriata per ketamina / xylazina e tintura RB. Anestetizzare l&#39;animale mediante iniezione intraperitoneale di 1 ml / kg della miscela di 80 mg / ml di ketamina e 4 mg / ml xilazina. </li><li> Lasciare l&#39;animale in una gabbia riscaldata fino a completo anestetizzato. Verificare la presenza di alcuna risposta a stimoli avversi (coda o pizzicare la punta). Controllare animali per profondità dell&#39;anestesia ogni 10 min. </li><li> Dilata allievi dell&#39;animale con 1% tropicamide e anestetizzare la superficie dell&#39;occhio con 0,5% proparacaina. Se si utilizzano gli animali pigmentate, come Long Evans, a 2,5% gocce oculari neosynephrine aumenterà la dilatazione della pupilla. </li><li> Usare le forbici per tagliare i baffi vicino al muso sul lato per essere indotti a evitare di bloccare la vista. </li><li> Mettere una goccia di 1% di metilcellulosa o di altra goccia accoppiamento oftalmica all&#39;interno della lente a contatto su misura, e quindi applicare l&#39;obiettivo sulla tha gli occhi di ratto. </li><li> Posto l&#39;animale su una piattaforma regolato all&#39;altezza della lampada a fessura. Impostare testa dell&#39;animale con un angolo di 45 ° in modo che l&#39;occhio è perpendicolare alla lampada a fessura e laser. </li><li> Visualizza l&#39;occhio attraverso la lampada a fessura oftalmica. Assicurarsi che il fascio di puntamento è la dimensione giusta, così come messo a fuoco e centrato direttamente sul nervo ottico visualizzato. Fotografare il fondo della retina con una fotocamera digitale con alta ASA (1.200 - 2.000) di velocità montato su uno dei pezzi occhio della lampada a fessura con un adattatore su misura. NOTA: La capacità di vedere i vasi coroideali rivela la trasparenza della retina. Questo è un segnale importante per essere in grado di rilevare successivamente ischemia retinica, che possono confondere l&#39;interpretazione se si verifica. rNAION è ottico ischemia del nervo, che si traduce in perdita isolato RGC, mentre i risultati ischemia retinica in danni alla retina che colpisce tutte le cellule degli strati retinici interni. </li><li> Facoltativamente, ulteriormente immagine retina enervo ottico utilizzando una spettrale dominio a coerenza ottica tomografo (SD-8 ottobre) per valutare l&#39;occhio non-indotta. Scansione en face (Figura 3B) e 7 scansioni trasversali attraverso la retina (Figura 3C). L&#39;SD-OCT di imaging utilizza la stessa lente a contatto utilizzato per l&#39;induzione laser. </li><li> Iniettare 1 ml / kg per via endovenosa attraverso RB vena della coda, e attendere 30 secondi, quindi attivare la potenza del laser. Questa temporizzazione consente RB di distribuire uniformemente nella circolazione. Usiamo un impulso laser di 50 mW a una sec / impulso. energie maggiori (≥ 60 MW) possono danneggiare la retina o causare ischemia vascolare retinica. Attenzione: Assicurarsi che ognuno di noi ha un paio di occhiali filtranti sicurezza laser della lunghezza d&#39;onda di blocco appropriata per evitare che la luce laser diffusa entra l&#39;occhio del ricercatore. NOTA: La somministrazione laser deve essere rapidamente dopo l&#39;iniezione IV RB poiché il colorante viene eliminato rapidamente dalla circolazione. Più a lungo l&#39;induzione laser, il più grave l&#39;ottica ischemia del nervo. Generalmente, gli animali sono date 7 - 12 impulsi sec in rapida successione. Il contatto della luce laser con il colorante circola dà vasi nervo ottico bella bagliore dorato che può essere visto attraverso la lampada a fessura (Figura 4B). Questo dimostra il colorante è stato iniettato per via sistemica e distribuita nel flusso sanguigno. Se la luce è debole, o del tutto assenti (Figura 4A), il colorante non è stato iniettato per via endovenosa. In questo caso, non danno una seconda iniezione immediatamente, poiché l&#39;animale dovrà recuperare per almeno due giorni prima reiniezione. </li><li> Subito dopo induzione, rimuovere le lenti a contatto. Coprire entrambi gli occhi con antibiotico oftalmico tripla (Neosporin / polimixina / bacitracina) pomata con desametasone, e mettere l&#39;animale in un riscaldamento pad 37 ° C in una sola gabbia alloggiata sotto stretta osservazione fino al pieno recupero. <ol><li> Pulire la lente a contatto con acqua distillata wAter e asciugare con un panno non abrasivo per un utilizzo futuro. </li><li> Due giorni dopo l&#39;induzione valutare l&#39;edema nervo ottico sia da fotografia a colori del fondo oculare e SD OCT-analisi 8. NOTA: Confrontando il grado di edema nervo ottico fornisce una stima della gravità della ischemia nervo ottico. In due giorni, il margine del disco nervo ottico è offuscata, e le vene della retina sono leggermente dilatati, rispetto con l&#39;occhio (non-indotta) controlaterale. Eseguire elettroretinografia (ERG) e flash visivo potenziali evocati (VEP 11) a due e quattro settimane inviare ad induzione per l&#39;analisi elettrofisiologia. </li></ol></li></ol>

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D., 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=PGJ2+Provides+Prolonged+CNS+Stroke+Protection+by+Reducing+White+Matter+Edema.">PGJ2 Provides Prolonged CNS Stroke Protection by Reducing White Matter Edema.</a> PLoS One. 7 (12), (2012).</li><li>Goldenberg-Cohen, N., 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=Oligodendrocyte+Dysfunction+Following+Induction+of+Experimental+Anterior+Optic+Nerve+Ischemia.">Oligodendrocyte Dysfunction Following Induction of Experimental Anterior Optic Nerve Ischemia.</a> Invest Ophthalmol Vis Sci. 46, 2716-2725 (2005).</li><li>Bernstein, S. L., Guo, Y., Kelman, S. E., Flower, R. W., Johnson, M. 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Gli autori non hanno nulla da rivelare.

Mentre ci sono un certo numero di modelli di danno del nervo ottico (nervo ottico crush 12, ottico resezione del nervo 13, e PION 14), il modello rNAION è umano, adattabile a entrambi i ratti e topi. Assomiglia più da vicino le condizioni cliniche umana di NAION. Questa condizione include progressiva anteriore del nervo ottico edema, una sindrome ottico scompartimento del nervo anteriore, focale ischemia assonale, isolato retina cellule gangliari danno assonale e la perdita di più di ...

Nonarteritic neuropatia ottica ischemica anteriore (NAION) è una lesione ischemica focale della porzione anteriore del nervo ottico (ON) 1. NAION è la causa più comune di improvvisa fibre nervose perdita della vista ottico in soggetti di età superiore a 50 2. Il meccanismo è ritenuta essere una sindrome vano che provoca edema intraneurale, e provoca la compressione dei capillari forniscono gli assoni nel nervo ottico 3. Dal momento che l&#39;ON è in realtà un tratto del sistema nervoso centrale (CNS), la NAION roditore modello (rNAION) può essere utilizzato per studiare i meccanismi e le risposte di isolati colpi della sostanza bianca del sistema nervoso centrale. Il modello rNAION può quindi essere utile in sezionare molti problemi connessi con danni relativi corsa di materia bianca. Può essere usato per valutare diverse strategie neuroprotettive e agenti in bianco ictus materia. Una delle caratteristiche più interessanti del modello è che èun indolore, procedura non invasiva. La potenza del laser può essere regolata per produrre vari gradi di danno ischemico. Un&#39;altra caratteristica è che si basa su laser indotta radicali superossido danneggiare l&#39;endotelio capillare, producendo una disfunzione capillare progressiva. È questa disfunzione e progressiva edema che si ritiene essere notevolmente simile al meccanismo che causa NAION. La ricerca ha dimostrato che non provoca la coagulazione capillare diretta, ma agisce attraverso almeno due meccanismi: superossido morte indotta e stripping di alcuni capillare cellule endoteliali 4, e NFkB (fattore nucleare kappa-light-chain-enhancer di cellule B attivate ) associato regolazione infiammatorio in endotelio restante, con un aumento del trasporto di liquidi attraverso le membrane cellulari nell&#39;interstizio 5. La chiusura dei capillari del nervo ottico e compressione causate da interstiziale risultato accumulo di liquido in ottica ischemia testa del nervo. Un quadro schematico è mostrato inFigura 2. Il modello rNAION può essere utilizzato in entrambe le specie di ratti e topi 6,7, e può essere variata nel livello della sua gravità, da una lesione lieve a una distruzione completa, ma indolore del nervo ottico e la retina, tale come occlusione dell&#39;arteria centrale della retina (CRAO).

<table border="0" cellpadding="0" cellspacing="0" width="600">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">50mW 532nm laser</td>
			<td style="width:99px;">Iridex</td>
			<td style="width:119px;"></td>
			<td style="width:167px;">Standard Ophthalmic Laser</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">0-100mW 532nm laser</td>
			<td style="width:99px;">Laserglow technologies</td>
			<td style="width:119px;"></td>
			<td>Substitute for iridex</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Laser slit lamp adapter</td>
			<td style="width:99px;">Iridex</td>
			<td style="width:119px;"></td>
			<td>SMA coupled adapter for laser output</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Cpherent Fieldmate laser meter with thermopile sensor</td>
			<td style="width:99px;">Coherent</td>
			<td style="width:119px;"></td>
			<td>others also appropriate</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Ophthalmic Examing Slit lamp biomicroscope</td>
			<td style="width:99px;">VArious</td>
			<td style="width:119px;"></td>
			<td>Haag-Streit is the best; cheaper versions available on ebay</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Rose Bengal</td>
			<td style="width:99px;">Sigma</td>
			<td style="width:119px;">330000-1G</td>
			<td>Photoinducing agent</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Fundus Contact lens or glass cover slip</td>
			<td style="width:99px;">custom/Cantor and Nissel (UK)</td>
			<td style="width:119px;"></td>
			<td>Custom designed planoconvex plastic lens for eye exam and induction</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Tropicamide 1%</td>
			<td style="width:99px;"></td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Tamiya polishing compound</td>
			<td style="width:99px;">Tamiya, INC</td>
			<td align="right" style="width:119px;">87068</td>
			<td>polishing contact lens</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">2.5% Hypromellose (Goniovisc)/1% Methycellulose</td>
			<td style="width:99px;">HUB Pharmaceuticals</td>
			<td style="width:119px;"></td>
			<td>contact lens coupling agent</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">2.5% Neosynephrine Ophthalmic drops</td>
			<td style="width:99px;">Alcon labs</td>
			<td style="width:119px;"></td>
			<td>pupil dilating agent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Tropicamide 1%</td>
			<td style="width:99px;">Alcon labs</td>
			<td style="width:119px;"></td>
			<td>pupil dilating agent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">0.5% Proparacaine</td>
			<td style="width:99px;">Alcon labs</td>
			<td style="width:119px;"></td>
			<td>topical Anesthetic</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">30ga fused needle insulin syringe</td>
			<td style="width:99px;">Various</td>
			<td style="width:119px;">Various</td>
			<td>for intravenous injection of rose bengal</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Ophthamic Antibiotic ointment with dexamethasone added (Triple antibiotic ointment)</td>
			<td style="width:99px;">Various</td>
			<td style="width:119px;">Various</td>
			<td>Apply after induciton to minimize corneal scarring</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Heidelberg Corporation Spectral domain-Optical Coherence Tomograph</td>
			<td style="width:99px;">Heidelberg Corportion</td>
			<td style="width:119px;"></td>
			<td>For Optical coherence measurements baseline and post-induction; not essential for induction</td>
		</tr>
	</tbody>
</table>

the rodent model of nonarteritic anterior ischemic optic neuropathy (rnaion)

Nonarteritic anterior ischemic optic neuropathy (NAION) is a focal ischemic lesion of the optic nerve that affects 1/700 individuals throughout their lifetime. NAION results in optic nerve edema, selective loss of the retinal ganglion cell neurons (RGCs) and atrophy of the optic nerve. A rodent model of NAION that expresses most NAION features and sequelae has been developed, which is applicable to both rats and mice. This model utilizes a focal laser application of 532 nm wavelength to illuminate a photoactive dye, Rose Bengal (RB), to cause capillary damage and leakage at the targeted anterior optic nerve (the laminar region). After rNAION induction, there is an early optic nerve ischemia, optic nerve edema, and intraneural inflammation, followed by selective RGC and axonal loss. Since the optic nerve is a CNS white matter tract, the rNAION model is applicable to mechanistic studies of selective white matter ischemia, as well as neuroprotective analyses and short and long-term mechanisms of glial and neuronal response to ischemia.

<html> La visualizzazione lente a contatto abilitato centrale retina (Figura 1). Il punto laser focale illumina il disco ottico sul retro della retina (Figura 2). Il normale retina non-indotta è mostrato ripreso dalla lampada a fessura bio-microscopio (figura 3A) e SD-OCT (Figura 3B e 3C). Durante l&#39;induzione laser, quando nessun colorante è presente nella circolazione, luce laser non comporta recipiente e fluoresce...

Watch this Scientific Journal Video about The Rodent Model of Nonarteritic Anterior Ischemic Optic Neuropathy rNAION at JoVE.com

The Rodent Model of Nonarteritic Anterior Ischemic Optic Neuropathy rNAION

The following report describes how to replicate the rodent model of nonarteritic anterior ischemic optic neuropathy (rNAION), using the appropriate dye, contact lens and laser parameters. We also reveal the appropriate steps for evaluating the rNAION lesion in vivo.

Il modello di roditore di nonarteritic Neuropatia ottica ischemica anteriore (rNAION)

Nonarteritic anterior ischemic optic neuropathy (NAION) is a focal ischemic lesion of the optic nerve that affects 1/700 individuals throughout their ...

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The Rodent Model of Nonarteritic Anterior Ischemic Optic Neuropathy (rNAION)

8.8K Views.  University of Maryland-Baltimore School of Medicine. The following report describes how to replicate the rodent model of nonarteritic anterior ischemic optic neuropathy (rNAION), using the appropriate dye, contact lens and laser parameters. We also reveal the appropriate steps for evaluating the rNAION lesion in vivo.

Video: The Rodent Model of Nonarteritic Anterior Ischemic Optic Neuropathy rNAION

Gene Transfer for Ischemic Heart Failure in a Preclinical Model

Various emerging technologies are being developed for patients with heart failure. Well-established preclinical evaluations are necessary to determine their efficacy and safety.
Gene therapy using viral vectors is one of the most promising approaches for treating cardiac diseases. Viral delivery of various different genes by changing the carrier gene has immeasurable therapeutic potential.
In this video, the full process of an animal model of heart failure creation followed by gene transfer is presented using a swine model. First, myocardial infarction is created by occluding the proximal left anterior descending coronary artery. Heart remodeling results in chronic heart failure. Unique to our model is a fairly large scar which truly reflects patients with severe heart failure who require aggressive therapy for positive outcomes. After myocardial infarct creation and development of scar tissue, an intracoronary injection of virus is demonstrated with simultaneous nitroglycerine infusion. Our injection method provides simple and efficient gene transfer with enhanced gene expression. This combination of a myocardial infarct swine model with intracoronary virus delivery has proven to be a consistent and reproducible methodology, which helps not only to test the effect of individual gene, but also compare the efficacy of many genes as therapeutic candidates.

A method of gene transfer for the treatment of ischemic heart failure is described using a swine model of myocardial infarction. Our simple and reproducible method enables us to readily evaluate the efficacy of various gene transfers with a very simple and reproducible way.

Trasferimento Genico per insufficienza cardiaca ischemica in un modello preclinico

Various emerging technologies are being developed for patients with heart failure. Well-established preclinical evaluations are necessary to determine ...

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Medicina

A Simplified Technique for Producing an Ischemic Wound Model

One major obstacle in current diabetic wound research is a lack of an ischemic wound model that can be safely used in diabetic animals. Drugs that work well in non-ischemic wounds may not work in human diabetic wounds because vasculopathy is one major factor that hinders healing of these wounds. We published an article in 2007 describing a rabbit ear ischemic wound model created by a minimally invasive surgical technique. Since then, we have further simplified the procedure for easier operation. On one ear, three small skin incisions were made on the vascular pedicles, 1-2 cm from the ear base. The central artery was ligated and cut along with the nerve. The whole cranial bundle was cut and ligated, leaving only the caudal branch intact. A circumferential subcutaneous tunnel was made through the incisions, to cut subcutaneous tissues, muscles, nerves, and small vessels. The other ear was used as a non-ischemic control. Four wounds were made on the ventral side of each ear. This technique produces 4 ischemic wounds and 4 non-ischemic wounds in one animal for paired comparisons. After surgery, the ischemic ear was cool and cyanotic, and showed reduced movement and a lack of pulse in the ear artery. Skin temperature of the ischemic ear was 1-10 &deg;C lower than that on the normal ear and this difference was maintained for more than one month. Ear tissue high-energy phosphate contents were lower in the ischemic ear than the control ear. Wound healing times were longer in the ischemic ear than in the non-ischemic ear when the same treatment was used. The technique has now been used on more than 80 rabbits in which 23 were diabetic (diabetes time ranging from 2 weeks to 2 years). No single rabbit has developed any surgical complications such as bleeding, infection, or rupture in the skin incisions. The model has many advantages, such as little skin disruption, longer ischemic time, and higher success rate, when compared to many other models. It can be safely used in animals with reduced resistance, and can also be modified to meet different testing requirements.

We have developed a minimally invasive technique to create a rabbit ischemic ear wound model by dividing the central artery and nerve and the cranial neurovascular bundle. A subcutaneous tunnel then cuts all subcutaneous tissues. This procedure causes minimal skin disruption and can be safely used in diabetic animals.

Una tecnica semplificata per produrre un modello di lesione ischemica

One major obstacle in current diabetic wound research is a lack of an ischemic wound model that can be safely used in diabetic animals. Drugs that work ...

Angiogenesis in the Ischemic Rat Lung

The adult lung is perfused by both the systemic bronchial artery and the entire venous return flowing through the pulmonary arteries. In most lung pathologies, it is the smaller systemic vasculature that responds to a need for enhanced lung perfusion and shows robust neovascularization. Pulmonary vascular ischemia induced by pulmonary artery obstruction has been shown to result in rapid systemic arterial angiogenesis in man as well as in several animal models. Although the histologic assessment of the time course of bronchial artery proliferation in rats was carefully described by Weibel 1, mechanisms responsible for this organized growth of new vessels are not clear. We provide surgical details of inducing left pulmonary artery ischemia in the rat that leads to bronchial neovascularization. Quantification of the extent of angiogenesis presents an additional challenge due to the presence of the two vascular beds within the lung. Methods to determine functional angiogenesis based on labeled microsphere injections are provided.

The lung is perfused by both the systemic bronchial artery and pulmonary arteries. In most lung pathologies, it is the smaller systemic vasculature that shows robust neovascularization. Cessation of pulmonary blood flow promotes brisk bronchial angiogenesis. We provide surgical details of inducing left pulmonary artery ischemia that promotes bronchial neovascularization.

L&#39;angiogenesi nel polmone del ratto ischemica

The  adult lung is perfused by both the systemic bronchial artery and the entire  venous return flowing through the pulmonary arteries. In most lung ...

A Murine Model of Myocardial Ischemia-reperfusion Injury through Ligation of the Left Anterior Descending Artery

Acute or chronic myocardial infarction (MI) are cardiovascular events resulting in high morbidity and mortality. Establishing the pathological mechanisms at work during MI and developing effective therapeutic approaches requires methodology to reproducibly simulate the clinical incidence and reflect the pathophysiological changes associated with MI. Here, we describe a surgical method to induce MI in mouse models that can be used for short-term ischemia-reperfusion (I/R) injury as well as permanent ligation. The major advantage of this method is to facilitate location of the left anterior descending artery (LAD) to allow for accurate ligation of this artery to induce ischemia in the left ventricle of the mouse heart. Accurate positioning of the ligature on the LAD increases reproducibility of infarct size and thus produces more reliable results. Greater precision in placement of the ligature will improve the standard surgical approaches to simulate MI in mice, thus reducing the number of experimental animals necessary for statistically relevant studies and improving our understanding of the mechanisms producing cardiac dysfunction following MI. This mouse model of MI is also useful for the preclinical testing of treatments targeting myocardial damage following MI.

We introduce a surgical method to induce experimental ischemia/reperfusion (I/R) injury to simulate myocardial infarction (MI) in mouse models that allows for more clarity in positioning of the ligation on the left anterior descending artery (LAD) to increase the reproducibility of MI experiments in mice.

Un modello murino di ischemia miocardica e riperfusione lesioni da legatura della arteria discendente anteriore sinistra

Acute or chronic myocardial infarction (MI) are cardiovascular events resulting in high morbidity and mortality. Establishing the pathological mechanisms ...

Rat Model of Photochemically-Induced Posterior Ischemic Optic Neuropathy

Posterior Ischemic optic neuropathy (PION) is a sight-devastating disease in clinical practice. However, its pathogenesis and natural history have&#160;remained poorly understood. Recently, we developed a reliable, reproducible animal model of PION and tested the treatment effect of some neurotrophic factors in this model1. The purpose of this video is to demonstrate our photochemically induced model of posterior ischemic optic neuropathy, and to evaluate its effects with retrograde labeling of retinal ganglion cells. Following surgical exposure of the posterior optic nerve, a photosensitizing dye, erythrosin B, is intravenously injected and a laser beam is focused onto the optic nerve surface. Photochemical interaction of erythrosin B and the laser during irradiation damages the vascular endothelium, prompting microvascular occlusion mediated by platelet thrombosis and edematous compression. The resulting ischemic injury yields a gradual but pronounced retinal ganglion cell dieback, owing to a loss of axonal input - a remote, injury-induced and clinically relevant outcome. Thus, this model provides a novel platform to study the pathophysiologic course of PION,&#160;and can be further optimized for testing therapeutic approaches for optic neuropathies as well as other CNS ischemic diseases.

The goal of this protocol is to photochemically induce ischemic injury to the posterior optic nerve in rat. This model is critical to studies of the pathophysiology of posterior ischemic optic neuropathy, and therapeutic approaches for this and other optic neuropathies, as well as of other CNS ischemic diseases.

Rat Modello di fotochimica-Induced posteriore Neuropatia ottica ischemica

Posterior Ischemic optic neuropathy (PION) is a sight-devastating disease in clinical practice. However, its pathogenesis and natural history ...

A Piglet Model of Neonatal Hypoxic-Ischemic Encephalopathy

Birth asphyxia, which causes hypoxic-ischemic encephalopathy (HIE), accounts for 0.66 million deaths worldwide each year, about a quarter of the world&#8217;s 2.9 million neonatal deaths. Animal models of HIE have contributed to the understanding of the pathophysiology in HIE, and have highlighted the dynamic process that occur in brain injury due to perinatal asphyxia. Thus, animal studies have suggested a time-window for post-insult treatment strategies. Hypothermia has been tested as a treatment for HIE in pdiglet models and subsequently proven effective in clinical trials. Variations of the model have been applied in the study of adjunctive neuroprotective methods and piglet studies of xenon and melatonin have led to clinical phase I and II trials1,2. The piglet HIE model is further used for neonatal resuscitation- and hemodynamic studies as well as in investigations of cerebral hypoxia on a cellular level. However, it is a technically challenging model and variations in the protocol may result in either too mild or too severe brain injury. In this article, we demonstrate the technical procedures necessary for establishing a stable piglet model of neonatal HIE. First, the newborn piglet (&#60; 24 hr old, median weight 1500 g) is anesthetized, intubated, and monitored in a setup comparable to that found in a neonatal intensive care unit. Global hypoxia-ischemia is induced by lowering the inspiratory oxygen fraction to achieve global hypoxia, ischemia through hypotension and a flat trace amplitude integrated EEG (aEEG) indicative of cerebral hypoxia. Survival is promoted by adjusting oxygenation according to the aEEG response and blood pressure. Brain injury is quantified by histopathology and magnetic resonance imaging after 72 hr.

Hypoxic-ischemic encephalopathy following perinatal asphyxia can be studied using animal models. We demonstrate the procedures necessary for establishing a piglet model of neonatal hypoxic-ischemic encephalopathy.

Un modello di Piglet neonatale Hypoxic-ischemica Encefalopatia

Birth asphyxia, which causes hypoxic-ischemic encephalopathy (HIE), accounts for 0.66 million deaths worldwide each year, about a quarter of the ...

Demonstration of the Rat Ischemic Skin Wound Model

The propensity for chronic wounds in humans increases with ageing, disease conditions such as diabetes and impaired cardiovascular function, and unrelieved pressure due to immobility. Animal models have been developed that attempt to mimic these conditions for the purpose of furthering our understanding of the complexity of chronic wounds. The model described herein is a rat ischemic skin flap model that permits a prolonged reduction of blood flow resulting in wounds that become ischemic and resemble a chronic wound phenotype (reduced vascularization, increased inflammation and delayed wound closure). It consists of a bipedicled dorsal flap with 2 ischemic wounds placed centrally and 2 non-ischemic wounds lateral to the flap as controls. A novel addition to this ischemic skin flap model is the placement of a silicone sheet beneath the flap that functions as a barrier and a splint to prevent revascularization and reduce contraction as the wounds heal. Despite the debate of using rats for wound healing studies due to their quite distinct anatomic and physiologic differences compared to humans (i.e., the presence of a panniculus carnosus muscle, short life-span, increased number of hair follicles, and their ability to heal infected wounds) the modifications employed in this model make it a valuable alternative to previously developed ischemic skin flap models.

The rat, due to its size, availability, and rather docile behavior, has been utilized as a research model for many years. The goal of this protocol is to utilize the rat as an ischemic skin wound healing model to provide valuable insight into the pathophysiology of chronic wounds.

Dimostrazione del Topo ischemica pelle ferita Modello

The propensity for chronic wounds in humans increases with ageing, disease conditions such as diabetes and impaired cardiovascular function, and ...

Isometric Contractility Measurement of the Mouse Mesenteric Artery Using Wire Myography

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and drugs. It is widely used in many studies on the physiological functions of vascular smooth muscle, the pathogenesis of vascular diseases such as hypertension, and the development of smooth muscle relaxant drugs. The mouse is a widely used model animal with a large pool of disease models and genetically modified strains. We introduced this method to measure the isometric contraction of mouse mesenteric artery in detail. A 1.4-mm segment of mouse mesenteric resistance artery was isolated and mounted on a myograph chamber by passing two steel wires through its lumen. After equilibration and normalization steps, the vessel segment was potentiated by a high-K+ solution twice prior to the contraction assay. As an example of the application of this method in drug development, we measured the relaxant effect of a novel natural substance, neoliensinine, isolated from a Chinese herb, embryos of the lotus seed (Nelumbo nucifera Gaertn.) on mouse mesenteric arteries. The vessel segments mounted on the myograph chamber were stimulated with a high-K+ solution. When the force tension reached a stable sustained phase, cumulative doses of neoliensinine were added to the chamber. We found that neoliensinine had a dose-dependent relaxant effect on smooth muscle contraction, thus suggesting that it bears potential activity against hypertension. In addition, as the vessel segment can survive at least 4 hours after mounting and maintain contractility induced by the high-K+ solution for many times, we suggest that the wire myograph system may be used for the time-consuming process of drug screening.

The wire myograph technique is used to investigate vascular smooth muscle functions and screen new drugs. We report a detailed protocol for measuring the isometric contractility of the mouse mesenteric artery and for screening new relaxants of vascular smooth muscle.

Misurazione di contrattilità isometrica dell'arteria mesenterica Mouse con filo Myography

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and ...

An Intact Pericardium Ischemic Rodent Model

This protocol has shown that the pericardium and its contents play an essential anti-fibrotic role in the ischemic rodent model (coronary ligation to induce myocardial injury). The majority of pre-clinical myocardial infarction models require the disruption of pericardial integrity with loss of the homeostatic cellular milieu. However, recently a methodology has been developed by us to induce myocardial infarction, which minimizes pericardial damage and retains the heart&#39;s resident immune cell population. An improved cardiac functional recovery in mice with an intact pericardial space following coronary ligation has been observed. This method provides an opportunity to study inflammatory responses in the pericardial space following myocardial infarction. Further development of the labeling techniques can be combined with this model to understand the fate and function of pericardial immune cells in regulating the inflammatory mechanisms that drive remodeling in the heart, including fibrosis.

This protocol outlines the steps for inducing myocardial infarction in mice while preserving the pericardium and its contents.

Un modello di roditore ischemico del pericardio intatto

This protocol has shown that the pericardium and its contents play an essential anti-fibrotic role in the ischemic rodent model (coronary ligation to ...

Large Animal Model for Evaluating the Efficacy of the Gene Therapy in Ischemic Heart

Coronary artery disease is one of the significant causes of mortality and morbidity worldwide. Despite the progression of current therapeutics, a considerable proportion of coronary artery disease patients remain symptomatic. Gene therapy-mediated therapeutic angiogenesis offers a novel therapeutic method for improving myocardial perfusion and relieving symptoms. Gene therapy with different angiogenic factors has been studied in few clinical trials. Due to the novelty of the method, the progress of myocardial gene therapy is a continuous path from bench to bedside. Therefore, large animal models are needed for evaluating the safety and efficacy. The more the large animal model identifies the original disease and the endpoints used in clinics, the more predictable outcomes are from clinical trials. Here, we introduce a large animal model for evaluating the efficacy of the gene therapy in the ischemic porcine heart. We use clinically relevant imaging methods such as ultrasound imaging and 15H2O-PET. For targeting the gene transfers into the desired area, electroanatomical mapping is used. The aim of this method is: (1) to mimic chronic coronary artery disease, (2) to induce therapeutic angiogenesis at hypoxic areas of the heart, and (3) to evaluate the safety and efficacy of the gene therapy by using relevant endpoints.

Myocardial gene therapy for ischemic heart disease holds great promise for future therapeutics. Here, we introduce a large animal model for evaluating the efficacy of gene therapy in the ischemic heart.