The authors have no acknowledgements.

Estos protocolos siguen las directrices del comité de nuestra institución ética de la investigación humana. 1. convencional colágeno corneal CXL (C-CXL) 1. Preparación del paciente <ol><li> 5 días antes de la cirugía, puso el 1% pilocarpina gotas dos veces al día en el ojo tratado. </li><li> En la sala de operaciones, en condiciones asépticas, mentir al paciente en su / su espalda. </li><li> Administrar anestesia tópica como oxibuprocaína 0,4%. </li><li> Limpiar el ojo y la piel alrededor del ojo con yodo antiséptico dos veces. </li><li> Utilice un espéculo tapa para mantener el ojo abierto. </li></ol> 2. Eliminación epitelial <ol><li> Marque el centro de 9,0 mm de la córnea con un círculo marcador de la córnea. </li><li> Retire las centrales 7,0 a 9,0 mm de epitelio corneal por desbridamiento mecánico utilizando una espátula roma. </li></ol> 3. La riboflavina Aplicación <ol><li> Aplicar 0.1% riboflavina con un 20% de dextrano en el th<html> e córnea cada min durante 20 min. </li></ol> La irradiación 4. UVA <ol><li> Irradiar la córnea con una longitud de onda de la luz UVA 370 nm a una irradiancia de 3 mW / cm2 (5,4 dosis en la superficie J / cm2) ya unos 5 cm de distancia de trabajo de 30 min. </li></ol><img alt="Figura 1" src="/files/ftp_upload/53119/53119fig1.jpg" /> Figura 1:. Irradiación UVA en C-CXL La córnea se irradia con una longitud de onda de luz UVA 370 nm con una irradiancia de 3 mW / cm 2 (5,4 dosis en la superficie J / cm 2) y a una distancia de trabajo de 5 cm durante 30 minutos. <a href="https://www.jove.com/files/ftp_upload/53119/53119fig1large.jpg" target="_blank">Haga clic aquí para ver una versión más grande de esta figura.</a> <ol start="2"><li> Durante la irradiación, se aplican gotas de riboflavina a la córnea cada 5 min. </li><li> Durante la irradiación, añadir anestesia tópica (oxibuprocaína 0,4%) si es necesario. </li></ol>&quot;&gt; 5. Fin de la Cirugía <ol><li> Ponga gotas antibióticas (tobramicina 0,3%) y las lágrimas artificiales (hialuronato cae 0,18%) en el ojo operado. </li><li> Colocar una lente de contacto vendaje blando al final de la cirugía hasta la reepitelización es completa. Re-epitelización tarda generalmente 3 días. </li><li> Prescribir analgésicos como el paracetamol (500 mg) más codeína (30 mg), 6 pastillas al día. </li><li> Después de la reepitelización corneal, iniciar el tratamiento tópico con esteroides (dexametasona tópica 1 mg / ml) y se continúa durante 3-4 semanas. Además, utilizar lágrimas artificiales 4 veces al día durante 1 mes. </li></ol> 2. acelerado colágeno corneal CXL (A-CXL) 1. Preparación del paciente <ol><li> 5 días antes de la cirugía, puso el 1% pilocarpina gotas dos veces al día en el ojo tratado. </li><li> En la sala de operaciones, en condiciones asépticas, mentir al paciente en su / su espalda. </li><li> Administrar anestesia tópica como oxibuprocaína 0,4%. </li><li> Limpie el correopues, y la piel alrededor del ojo con yodo antiséptico dos veces. </li><li> Utilice un espéculo tapa para mantener el ojo abierto. </li></ol> 2. Eliminación epitelial <ol><li> Marque el centro de 9,0 mm de la córnea con un círculo marcador de la córnea </li><li> Retire las centrales 7,0 a 9,0 mm de epitelio corneal por desbridamiento mecánico utilizando una espátula roma. </li></ol> 3. La riboflavina Aplicación <ol><li> Aplicar 0,1% de riboflavina sin dextrano sobre la córnea cada 2 min durante 10 min. </li></ol> La irradiación 4. UVA <ol><li> Irradiar la córnea con una longitud de onda de luz UVA 370 nm con una irradiancia de 30 mW / cm 2 (5,4 J dosis en la superficie / cm 2) y a una distancia de trabajo 5 cm por 3 min. </li><li> Durante la irradiación, añadir anestesia tópica (oxybuproca &amp; # 239; ne 0,4%) si es necesario. </li></ol> 5. Fin de la Cirugía <ol><li> Coloca gotas antibióticas (tobramicina 0,3%) y las lágrimas artificiales (hialuronato gotas 0<html> 0,18%) en el ojo operado. </li><li> Colocar una lente de contacto vendaje blando al final de la cirugía hasta la reepitelización es completa. Re-epitelización tarda generalmente 3 días. </li><li> Prescribir analgésicos como el paracetamol (500 mg) más codeína (30 mg), 6 pastillas al día. </li><li> Después de la reepitelización corneal, iniciar el tratamiento tópico con esteroides (dexametasona tópica 1 mg / ml) y se continúa durante 3-4 semanas. Además, utilizar lágrimas artificiales 4 veces al día durante 1 mes. </li></ol> 3. La iontoforesis (I-CXL) 1. Preparación del paciente <ol><li> 5 días antes de la cirugía, puso el 1% pilocarpina gotas dos veces al día en el ojo tratado. </li><li> En la sala de operaciones, en condiciones asépticas, mentir al paciente en su / su espalda. </li><li> Administrar anestesia tópica como oxibuprocaína 0,4%. </li><li> Limpiar el ojo y la piel alrededor del ojo con yodo antiséptico dos veces. </li><li> Utilice un espéculo tapa para mantener el ojo abierto. </li></ol><html> _step &quot;&gt; 2. Coloque el aparato iontoforesis. <ol><li> Aplicar el electrodo pasivo pegajosa en la frente bajo el campo operatorio. </li><li> Aplique el electrodo activo, un anillo de succión, con el ojo abierto. Centre el anillo de succión en la periferia de la córnea antes de liberar la succión. </li></ol><img alt="Figura 2" src="/files/ftp_upload/53119/53119fig2.jpg" /> Figura 2. dispositivo de iontoforesis. El electrodo pasivo se aplica en la frente bajo el campo operatorio y el electrodo activo, un anillo de succión, se aplica al ojo abierto. <a href="https://www.jove.com/files/ftp_upload/53119/53119fig2large.jpg" target="_blank">Por favor haga clic aquí para ver una versión más grande de esta figura.</a> 3. La riboflavina Aplicación <ol><li> Llene el anillo de succión con hipoosmolar 0,1% riboflavina sin dextrano. </li></ol><html> jpg &quot;/&gt; Figura 3. Aplicación riboflavina en la I-CXL. El anillo de succión está lleno de hipoosmolar 0,1% riboflavina sin dextrano. <a href="https://www.jove.com/files/ftp_upload/53119/53119fig3large.jpg" target="_blank">Haga clic aquí para ver una versión más grande de esta figura.</a> <ol start="2"><li> Comience la corriente eléctrica en 0,2 mA y aumentar gradualmente a 1,0 mA durante un tiempo iontoforesis total de 5 min (Figura 4). </li></ol><img alt="Figura 4" src="/files/ftp_upload/53119/53119fig4.jpg" /> Figura 4. dispositivo de iontoforesis para la riboflavina penetración. La corriente eléctrica es inicialmente 0,2 mA y aumentó gradualmente hasta 1,0 mA. El tiempo total de la iontoforesis es de 5 minutos. <a href="https://www.jove.com/files/ftp_upload/53119/53119fig4large.jpg" target="_blank">Por favor haga clic aquí para ver una versión más grande de esta figura.</a> ep &quot;&gt; 4. La irradiación UVA <ol><li> Irradiar la córnea con una longitud de onda de la luz UVA 370 nm a una irradiancia de 10 mW / cm2 (5,4 J dosis en la superficie / cm2) ya una distancia de trabajo de 5 cm por 9 min. </li><li> Durante la irradiación, añadir anestesia tópica (oxybuproca &amp; # 239; ne 0,4%) si es necesario. </li></ol> 5. Fin de la Cirugía <ol><li> Coloca gotas antibióticas (tobramicina 0,3%) y las lágrimas artificiales (hialuronato cae 0,18%) en el ojo operado. </li><li> Prescribir analgésicos como el paracetamol (500 mg) más codeína (30 mg), 6 pastillas al día. </li><li> Después de la cirugía, iniciar la terapia tópica con esteroides (dexametasona tópica de 1 mg / ml) y se continúa durante 3-4 semanas. Además, utilizar lágrimas artificiales 4 veces al día durante 1 mes. </li></ol>

<ol>
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P., Godefrooij, D. A., Imhof, S. M., Tahzib, N. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transepithelial+versus+epithelium-off+corneal+cross-linking+for+the+treatment+of+progressive+keratoconus:+a+randomized+controlled+trial.">Transepithelial versus epithelium-off corneal cross-linking for the treatment of progressive keratoconus: a randomized controlled trial.</a> Am J Ophthalmol. 159 (5), 821-828 (2015).</li></ol>

The authors have nothing to disclose.

CXL utilizando irradiación UVA y riboflavina es el tratamiento estándar para detener la progresión del queratocono. La riboflavina es un fotosensibilizador que induce químicas enlaces covalentes (enlaces cruzados) cuando se irradia con rayos UVA 3. En la córnea, este fenómeno crea enlaces cruzados entre las fibrillas de colágeno que aumentan la rigidez corneal. Aunque este fenómeno está bien descrita, hasta ahora no ha habido ninguna evidencia directa de intracorneales enlaces cruzados. Sin embargo, ...

El queratocono es una ectasia corneal bilateral y progresiva usualmente reportada en 1 en 2000 en la población general 1 resulta en la modificación de la forma de la córnea y por lo tanto disminución de la visión 2. El queratocono es generalmente presente en la pubertad temprana y progresa hasta la tercera a la cuarta década de la vida cuando la enfermedad normalmente tiende a estabilizarse, aunque la progresión puede ser variable durante toda la vida de un paciente. Al detener la progresión del queratocono, reticulación pretende posponer o evitar la queratoplastia. Hasta la fecha, el único tratamiento eficaz y seguro de queratocono progresivo demostrado en estudios clínicos es el protocolo convencional colágeno corneal entrecruzamiento (C-CXL), cuyo objetivo es aumentar la rigidez y por lo tanto, detener la progresión del queratocono 3-8. Con el fin de reducir el tiempo de operación y otros factores de posible riesgo de C-CXL, tales como queratitis infecciosa o del estroma neblina 9, varios protocolos mejorados tienenha descrito. En primer lugar, en CXL acelerado (A-CXL), una mayor irradiación de UVA se entrega a la córnea durante un tiempo reducido 10. En segundo lugar, para evitar la necesidad para el desbridamiento epitelial, se han empleado enfoques transepiteliales. Desafortunadamente, tienen un éxito limitado en comparación con el protocolo convencional 11. El método transepitelial más reciente para la entrega riboflavina corneal durante CXL es iontoforesis (I-CXL), pero la evaluación rigurosa de este tratamiento aún no se ha realizado 12. La iontoforesis es una técnica no invasiva en la que se aplica una pequeña corriente eléctrica para mejorar la penetración de un fármaco ionizado a través de un tejido. En CXL por iontoforesis, la riboflavina se ioniza para penetrar la córnea a través del epitelio. In vivo microscopía confocal (IVCM) es un método de formación de imágenes de la córnea que pueden resaltar los cambios celulares anormales en las córneas de enfermedades tales como el queratocono 13. De hecho, IVCMha demostrado alteraciones a todas las capas de la córnea en el queratocono con una reducción en particular de la densidad de los nervios del plexo sub-basal y del estroma 13-15 de queratocitos. Además, IVCM ha demostrado ser altamente conveniente para el análisis microestructural de la córnea después de C-16 CXL. La línea de demarcación de la córnea se describe como una línea hiperreflectivo visto en segmento anterior tomografía de coherencia óptica (AS octubre) 1 mes después de la C-CXL a una profundidad de 300 m 17,18. IVCM siguiente C-CXL proporciona información acerca de alteraciones estructurales de la córnea, incluyendo la ausencia de queratocitos de la córnea hasta una profundidad de 300 m. La profundidad de esta zona acelular, así como la profundidad de la línea de demarcación dentro del estroma corneal revelaron en AS PTU, parece estar asociado con el profundidad efectiva del tratamiento CXL 19, y medición de la profundidad de la línea de demarcación de la córnea en AS 01 de octubre meses después de CXL ha sido propuesta como un clínico eficientemétodo para la evaluación de la eficacia CXL 18. En el presente estudio se investiga la eficacia de los tres protocolos diferentes de reticulación del colágeno corneal (convencionales, acelerada y iontoforesis) mediante la medición de la línea de demarcación del estroma corneal por AS octubre y microscopía confocal. Además Utilizamos IVCM analizar cuantitativamente los cambios de microestructura de la córnea después de los tres tratamientos.

<table>
	<tbody>
		<tr>
			<td></td>
			<td></td>
			<td>Riboflavin&#160;&#160;&#160;&#160;&#160;&#160;&#160; Product number</td>
			<td></td>
		</tr>
		<tr>
			<td>C-CXL</td>
			<td>Sooft SPA, Montegiorgio, Italy</td>
			<td>Ricrolin&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160; 468465-6</td>
			<td></td>
		</tr>
		<tr>
			<td>A-CXL</td>
			<td>Avedro Inc, Waltham, Massachusetts</td>
			<td>VibeX&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160; 520-01863-006</td>
			<td></td>
		</tr>
		<tr>
			<td>I-CXL</td>
			<td>Sooft SPA, Montegiorgio, Italy</td>
			<td>Ricrolin+&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160; 975481-6</td>
			<td>Passive electrode: PROTENS ELITE 4848LE/ Active electrode: IONTOFOR CXL</td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td>UVA Machine</td>
			<td></td>
		</tr>
		<tr>
			<td>X-Vega</td>
			<td></td>
			<td></td>
			<td>UVA: 3 mW/cm2 30 min</td>
		</tr>
		<tr>
			<td>KXL System</td>
			<td></td>
			<td></td>
			<td>UVA: 30 mW/cm2 10 min</td>
		</tr>
		<tr>
			<td>X-Vega</td>
			<td></td>
			<td></td>
			<td>UVA: 10 mW/cm2 9 min</td>
		</tr>
	</tbody>
</table>

three different protocols of corneal collagen crosslinking in keratoconus: conventional, accelerated and iontophoresis

El queratocono es una ectasia corneal bilateral y progresiva. Con el fin de frenar su progresión, colágeno entrecruzamiento corneal (CXL) recientemente se ha introducido como una opción de tratamiento eficaz. En las ciencias biológicas y químicas, la reticulación se refiere a nuevos enlaces químicos formados entre moléculas reactivas. Por lo tanto, el objetivo de CXL colágeno corneal es aumentar sintéticamente la formación de enlaces cruzados entre las fibrillas de colágeno en el estroma corneal. A pesar del hecho de que la eficiencia del protocolo CXL convencional (C-CXL) ya se ha demostrado en varios estudios clínicos, podría beneficiarse de las mejoras en la duración del procedimiento y la eliminación del epitelio corneal. Por lo tanto, con el fin de proporcionar una evaluación coherente de dos nuevas y optimizadas protocolos CXL, se estudiaron pacientes con queratocono que se habían sometido a uno de los tres tratamientos CXL: iontoforesis (I-CXL), acelerada CXL (A-CXL) y CXL convencional ( C-CXL). A-CXL es 6 veces más rápido procedimiento de CXL ucantar una irradiación UVA diez veces mayor, pero aún incluyendo una eliminación epitelio. La iontoforesis es una técnica no invasiva transepitelial en que se aplica una pequeña corriente eléctrica para mejorar la penetración de riboflavina en toda la córnea. El uso de la tomografía de coherencia óptica del segmento anterior (AS octubre) e in vivo microscopía confocal (IVCM), llegamos a la conclusión de que con respecto a la profundidad de la penetración del tratamiento, el protocolo CXL convencional sigue siendo el estándar para el tratamiento de queratocono progresivo. Acelerado CXL parece ser una alternativa rápida, eficaz y segura para el tratamiento de córneas delgadas. El uso de la iontoforesis todavía está siendo investigado y debe considerarse con mayor cautela.

<html> La línea de demarcación de la córnea fue visible en AS octubre en el 92% de los casos, a una profundidad media de 301,6 mm (SD, 73.6) <img alt="Figura 5" src="/files/ftp_upload/53119/53119fig5.jpg" /> Figura 5. Línea de Demarcación después de C-CXL. De alta resolución del segmento anterior de la córnea tomografía de coherencia óptica (AS octubre) la visualización de la línea de demarcación del estroma de la córne...

Watch this Scientific Journal Video about Three Different Protocols of Corneal Collagen Crosslinking in Keratoconus: Conventional, Accelerated and Iontophoresis at JoVE.com

Three Different Protocols of Corneal Collagen Crosslinking in Keratoconus: Conventional, Accelerated and Iontophoresis

Corneal collagen cross-linking (CXL) is the only conservative treatment currently available to halt keratoconus progression by improving the biomechanical rigidity of the corneal stroma. The aim of this manuscript is to highlight the methods of three different protocols of CXL: conventional CXL (C-CXL), accelerated CXL (A-CXL), and iontophoresis CXL (I-CXL).

Tres diferentes Protocolos de colágeno corneal entrecruzamiento en queratocono: convencional, acelerado y iontoforesis

Keratoconus is a bilateral and progressive corneal ectasia. In order to slow down its progression, corneal collagen cross-linking (CXL) has recently been ...

three-different-protocols-corneal-collagen-crosslinking-keratoconus

Research

JoVE Journal

Medicine

19.8K vistas.  Quinze-Vingts National Ophthalmology Hospital. Corneal collagen cross-linking (CXL) is the only conservative treatment currently available to halt keratoconus progression by improving the biomechanical rigidity of the corneal stroma. The aim of this manuscript is to highlight the methods of three different protocols of CXL: conventional CXL (C-CXL), accelerated CXL (A-CXL), and iontophoresis CXL (I-CXL).

Video: Three Different Protocols of Corneal Collagen Crosslinking in Keratoconus: Conventional, Accelerated and Iontophoresis

Staining Protocols for Human Pancreatic Islets

Estimates of islet area and numbers and endocrine cell composition in the adult human pancreas vary from several hundred thousand to several million and beta mass ranges from 500 to 1500 mg 1-3. With this known heterogeneity, a standard processing and staining procedure was developed so that pancreatic regions were clearly defined and islets characterized using rigorous histopathology and immunolocalization examinations. 
Standardized procedures for processing human pancreas recovered from organ donors are described in part 1 of this series. The pancreas is processed into 3 main regions (head, body, tail) followed by transverse sections. Transverse sections from the pancreas head are further divided, as indicated based on size, and numbered alphabetically to denote subsections. This standardization allows for a complete cross sectional analysis of the head region including the uncinate region which contains islets composed primarily of pancreatic polypeptide cells to the tail region.
The current report comprises part 2 of this series and describes the procedures used for serial sectioning and histopathological characterization of the pancreatic paraffin sections with an emphasis on islet endocrine cells, replication, and T-cell infiltrates. Pathology of pancreatic sections is intended to characterize both exocrine, ductular, and endocrine components. The exocrine compartment is evaluated for the presence of pancreatitis (active or chronic), atrophy, fibrosis, and fat, as well as the duct system, particularly in relationship to the presence of pancreatic intraductal neoplasia4. Islets are evaluated for morphology, size, and density, endocrine cells, inflammation, fibrosis, amyloid, and the presence of replicating or apoptotic cells using H&amp;E and IHC stains. 
The final component described in part 2 is the provision of the stained slides as digitized whole slide images. The digitized slides are organized by case and pancreas region in an online pathology database creating a virtual biobank. Access to this online collection is currently provided to over 200 clinicians and scientists involved in type 1 diabetes research. The online database provides a means for rapid and complete data sharing and for investigators to select blocks for paraffin or frozen serial sections.

This video demonstrates procedures for characterization of human pancreatic islets using hematoxylin and eosin (H&E) and immunohistochemistry (IHC). Pancreatic sections from head, body, and tail regions are stained by both H&amp;E and IHC to determine islet endocrine composition (insulin, glucagon, and pancreatic polypeptide), cell replication (Ki67), and inflammatory infiltrates (H&E, CD3). The uncinate region is localized using IHC for pancreatic polypeptide.

Protocolos de tinción de islotes pancreáticos humanos

Estimates of islet area and numbers and endocrine cell composition in the adult human pancreas vary from several hundred thousand to several million and ...

Investigación

Medicina

An Alkali-burn Injury Model of Corneal Neovascularization in the Mouse

Under normal conditions, the cornea is avascular, and this transparency is essential for maintaining good visual acuity. Neovascularization (NV) of the cornea, which can be caused by trauma, keratoplasty or infectious disease, breaks down the so called &lsquo;angiogenic privilege' of the cornea and forms the basis of multiple visual pathologies that may even lead to blindness. Although there are several treatment options available, the fundamental medical need presented by corneal neovascular pathologies remains unmet. In order to develop safe, effective, and targeted therapies, a reliable model of corneal NV and pharmacological intervention is required. Here, we describe an alkali-burn injury corneal neovascularization model in the mouse. This protocol provides a method for the application of a controlled alkali-burn injury to the cornea, administration of a pharmacological compound of interest, and visualization of the result. This method could prove instrumental for studying the mechanisms and opportunities for intervention in corneal NV and other neovascular disorders.

Neovascularization (NV) of the cornea can complicate multiple visual pathologies. Utilizing a controlled, alkali-burn injury model, a quantifiable level of corneal NV can be produced for mechanistic study of corneal NV and evaluation of potential therapies for neovascular disorders.

Una Lesión álcalis quemar modelo de neovascularización corneal en el ratón

Under normal conditions, the cornea is avascular, and this transparency is essential for maintaining good visual acuity. Neovascularization (NV) of the ...

A Rat Model of Ventricular Fibrillation and Resuscitation by Conventional Closed-chest Technique

A rat model of electrically-induced ventricular fibrillation followed by cardiac resuscitation using a closed chest technique that incorporates the basic components of cardiopulmonary resuscitation in humans is herein described. The model was developed in 1988 and has been used in approximately 70 peer-reviewed publications examining a myriad of resuscitation aspects including its physiology and pathophysiology, determinants of resuscitability, pharmacologic interventions, and even the effects of cell therapies. The model featured in this presentation includes: (1) vascular catheterization to measure aortic and right atrial pressures, to measure cardiac output by thermodilution, and to electrically induce ventricular fibrillation; and (2) tracheal intubation for positive pressure ventilation with oxygen enriched gas and assessment of the end-tidal CO2. A typical sequence of intervention entails: (1) electrical induction of ventricular fibrillation, (2) chest compression using a mechanical piston device concomitantly with positive pressure ventilation delivering oxygen-enriched gas, (3) electrical shocks to terminate ventricular fibrillation and reestablish cardiac activity, (4) assessment of post-resuscitation hemodynamic and metabolic function, and (5) assessment of survival and recovery of organ function. A robust inventory of measurements is available that includes &#8211; but is not limited to &#8211; hemodynamic, metabolic, and tissue measurements. The model has been highly effective in developing new resuscitation concepts and examining novel therapeutic interventions before their testing in larger and translationally more relevant animal models of cardiac arrest and resuscitation.

This article describes a rat model of electrically-induced ventricular fibrillation and resuscitation by chest compression, ventilation, and delivery of electrical shocks that simulates an episode of sudden cardiac arrest and conventional cardiopulmonary resuscitation. The model enables gathering insights on the pathophysiology of cardiac arrest and exploration of new resuscitation strategies.

Un modelo de rata de la fibrilación ventricular y Reanimación por Convencional cerrado pecho Técnica

A rat model of electrically-induced ventricular fibrillation followed by cardiac resuscitation using a closed chest technique that incorporates the basic ...

Echocardiographic Approaches and Protocols for Comprehensive Phenotypic Characterization of Valvular Heart Disease in Mice

The aim of this manuscript and accompanying video is to provide an overview of the methods and approaches used for imaging heart valve function in rodents, with detailed descriptions of the appropriate methods for anesthesia, the echocardiographic windows used, the imaging planes and probe orientations for image acquisition, the methods for data analysis, and the limitations of emerging technologies for the evaluation of cardiac and valvular function. Importantly, we also highlight several future areas of research in cardiac and heart valve imaging that may be leveraged to gain insights into the pathogenesis of valve disease in preclinical animal models. We propose that using a systematic approach to evaluating cardiac and heart valve function in mice can result in more robust and reproducible data, as well as facilitate the discovery of previously underappreciated phenotypes in genetically-altered and/or physiologically-stressed mice.

This protocol provides a detailed description of the echocardiographic approach for comprehensive phenotyping of heart and heart valve function in mice.

Enfoques y Protocolos para la Integral ecocardiográficos Caracterización fenotípica en las valvulopatías en ratones

The aim of this manuscript and accompanying video is to provide an overview of the methods and approaches used for imaging heart valve function in ...

Induction of Accelerated Atherosclerosis in Mice: The "Wire-Injury" Model

Atherosclerosis is a proliferative fibro-inflammatory disease developing in the arterial wall, inducing a deficient blood flow or a lack of blood flow. Moreover, by rupture of the defective vascular wall, atherosclerosis induces occlusive thrombus formation, which represents the main cause of myocardial infarction or stroke and the most frequent cause of death. Despite the advances in the cardiovascular field, many questions remain unanswered, and additional basic research is essential to improve our understanding of the molecular mechanisms during atherosclerosis and its effects. Due to limited clinical studies, there is a need for representative animal models recreating atherosclerotic conditions such as neointima formation after stent implantation, balloon angioplasty, or endarterectomy. Since the mouse presents many advantages and is the most frequently used model for studying molecular processes, the current study proposes an invasive procedure of endothelial denudation, also known as the wire-injury model, which is representative of the human condition of neointima formation in arteries after revascularization procedures.

Induction of Accelerated Atherosclerosis in Mice: The &quot;Wire-Injury&quot; Model

This study describes an invasive procedure for the induction of accelerated atherosclerosis in mice. In comparison to other methods using electric- or cryo-induced injury, mechanical-induced injury mimics the human condition of restenosis after revascularization therapies and is ideal for the study of the molecular mechanisms involved.

Inducción de la Aterosclerosis Acelerada en Ratones: El Modelo de "Lesiones por Alambre"

Atherosclerosis is a proliferative fibro-inflammatory disease developing in the arterial wall, inducing a deficient blood flow or a lack of blood flow. ...

Corneal Epithelial Abrasion with Ocular Burr As a Model for Cornea Wound Healing

The murine cornea provides an excellent model to study wound healing. The cornea is the outermost layer of the eye, and thus is the first defense to injury. In fact, the most common type of eye injury found in clinic is a corneal abrasion. Here, we utilize an ocular burr to induce an abrasion resulting in removal of the corneal epithelium in vivo on anesthetized mice. This method allows for targeted and reproducible epithelial disruption, leaving other areas intact. In addition, we describe the visualization of the abraded epithelium with fluorescein staining and provide concrete advice on how to visualize the abraded cornea. Then, we follow the timeline of wound healing 0, 18, and 72 h after abrasion, until the wound is re-epithelialized. The epithelial abrasion model of corneal injury is ideal for studies on epithelial cell proliferation, migration and re-epithelialization of the corneal layers. However, this method is not optimal to study stromal activation during wound healing, because the ocular burr does not penetrate to the stromal cell layers. This method is also suitable for clinical applications, for example, pre-clinical test of drug effectiveness.

This protocol describes a method to inflict an abrasion to the ocular surface of the mouse, and to follow the wound healing process thereafter. The protocol takes advantage of an ocular burr to partially remove the surface epithelium of the eye in anaesthetized mice.

Cicatrización de heridas de abrasión epitelial corneal con rebabas Ocular como un modelo de córnea

The murine cornea provides an excellent model to study wound healing. The cornea is the outermost layer of the eye, and thus is the first defense to ...

Cardiac Magnetic Resonance for the Evaluation of Suspected Cardiac Thrombus: Conventional and Emerging Techniques

We present the conventional cardiac magnetic resonance (CMR) protocol for evaluating a suspected thrombus and highlight emerging techniques.&#160;The appearance of a mass on certain magnetic resonance (MR) sequences can help differentiate a thrombus from competing diagnoses such as a tumor. T1 and T2 signal characteristics of a thrombus are related to the evolution of hemoglobin properties. A thrombus typically does not enhance following contrast administration, which also helps differentiation from a tumor. We also highlight the emerging role of T1 mapping in the evaluation of a thrombus, which can add another level of support in diagnosis. Prior to any CMR exam, patient screening and interviews are critical to ensure safety and to optimize patient comfort. Effective communication during the exam between the technologist and the patient promotes proper breath holding technique and higher quality images. Volumetric post processing and structured reporting are helpful to ensure that the radiologist answers the ordering services&#39; question and communicates these results effectively. Optimal pre-MR safety evaluation, CMR exam execution, and post exam processing and reporting allow for delivery of high quality radiological service in the evaluation of a suspected cardiac thrombus.

The goal of this article is to describe how cardiac magnetic resonance can be used for the evaluation and diagnosis of a suspected cardiac thrombus. The method presented will describe data acquisition as well as the pre-procedure and post-procedure protocol.

Resonancia Magnética Cardíaca para la Evaluación del Trombo Cardíaco Sospechoso: Técnicas Convencionales y Emergentes

We present the conventional cardiac magnetic resonance (CMR) protocol for evaluating a suspected thrombus and highlight emerging techniques.&#160;The ...

Simultaneous Laryngopharyngeal and Conventional Esophageal pH Monitoring

In addition to typical reflux symptoms, many patients with gastroesophageal reflux disease (GERD) present with extraesophageal symptoms such as cough, hoarseness or asthma, which can be caused by laryngopharyngeal reflux (LPR). Due to their multifactorial origin, those symptoms can be a great diagnostic and therapeutic challenge. Esophageal pH-monitoring is commonly used to determine abnormal esophageal acid exposure and confirm the diagnosis of GERD. However, for better evaluation of acid exposure above the upper esophageal sphincter, a new laryngopharyngeal pH measurement system is now available and may lead to more reliable results in patients with predominantly extraesophageal symptoms. This article aims to present a standardized protocol for simultaneous pH measurement using esophageal and laryngopharyngeal pH probes in order to obtain acid exposure scores from both measurements.

Laryngopharyngeal pH monitoring has been specifically designed to measure acid exposure above the upper esophageal sphincter and complements diagnostic evaluation in patients that present with mainly extraesophageal reflux symptoms. Patients with suspected laryngopharyngeal reflux (LPR) were evaluated using distal esophageal and laryngopharyngeal pH testing simultaneously.

Monitoreo simultáneo del pH esofágico laringofaríngeo y convencional

In addition to typical reflux symptoms, many patients with gastroesophageal reflux disease (GERD) present with extraesophageal symptoms such as cough, ...

Three-Dimensional Collagen Matrix Scaffold Implantation as a Liver Regeneration Strategy

Liver diseases are the leading cause of death worldwide. Excessive alcohol consumption, a high-fat diet, and hepatitis C virus infection promote fibrosis, cirrhosis, and/or hepatocellular carcinoma. Liver transplantation is the clinically recommended procedure to improve and extend the life span of patients in advanced disease stages. However, only 10% of transplants are successful, with organ availability, presurgical and postsurgical procedures, and elevated costs directly correlated with that result. Extracellular matrix (ECM) scaffolds have emerged as an alternative for tissue restoration. Biocompatibility and graft acceptance are the main beneficial characteristics of those biomaterials. Although the capacity to restore the size and correct function of the liver has been evaluated in liver hepatectomy models, the use of scaffolds or some kind of support to replace the volume of the extirpated liver mass has not been assessed.
Partial hepatectomy was performed in a rat liver with the xenoimplantation of a collagen matrix scaffold (CMS) from a bovine condyle. Left liver lobe tissue was removed (approximately 40%), and an equal proportion of CMS was surgically implanted. Liver function tests were evaluated before and after the surgical procedure. After days 3, 14, and 21, the animals were euthanized, and macroscopic and histologic evaluations were performed. On days 3 and 14, adipose tissue was observed surrounding the CMS, with no clinical evidence of rejection or infection, as was vessel neoformation and CMS reabsorption at day 21. There was histologic evidence of an insignificant inflammation process and migration of adjacent cells to the CMS, observed with the hematoxylin and eosin (H&#38;E) and Masson&#39;s trichrome staining. The CMS was shown to perform well in liver tissue and could be a useful alternative for studying tissue regeneration and repair in chronic liver diseases.

Liver diseases are induced by many causes that promote fibrosis or cirrhosis. Transplantation is the only option for recovering health. However, given the scarcity of transplantable organs, alternatives must be explored. Our research proposes the implantation of collagen scaffolds in liver tissue from an animal model.

Implantación de andamio de matriz de colágeno tridimensional como estrategia de regeneración hepática

Liver diseases are the leading cause of death worldwide. Excessive alcohol consumption, a high-fat diet, and hepatitis C virus infection promote fibrosis, ...

Ex Vivo and In Vivo Animal Models for Mechanical and Chemical Injuries of Corneal Epithelium

Corneal injury to the ocular surface, including chemical burn and trauma, may cause severe scarring, symblepharon, corneal limbal stem cells deficiency, and result in a large, persistent corneal epithelial defect. Epithelial defect with the following corneal opacity and peripheral neovascularization result in irreversible visual impairment and hinder future management, especially keratoplasty. Since the animal model can be used as an effective drug development platform, models of corneal injury to the mouse and alkali burn to rabbit corneal epithelium are developed here. New Zealand white rabbit is used in the alkali burn model. Different concentrations of sodium hydroxide can be applied onto the central circular area of the cornea for 30 s under intramuscular and topical anesthesia. After copious isotonic normal saline irrigation, residual loose corneal epithelium was removed with corneal burr deep down to the Bowman's layer within this circular area. Wound healing was documented by fluorescein staining under Cobalt blue light. C57BL/6 mice were used in the traumatic model of murine corneal epithelium. The murine central cornea was marked using a skin punch, 2 mm in diameter, and then debrided by a corneal rust ring remover with a 0.5 mm burr under a stereomicroscope. These models can be prospectively used to validate the therapeutic effect of eye drops or mixed agents such as stem cells, which potentially facilitate corneal epithelial regeneration. By observing corneal opacity, peripheral neovascularization, and conjunctival congestion with stereomicroscope and imaging software, therapeutic effects in these animal models can be monitored.

Ex Vivo and In Vivo Animal Models for Mechanical and Chemical Injuries of Corneal Epithelium

Here, animal models based on mouse and rabbit are developed for mechanical and chemical injury of corneal epithelium to screen new therapeutics and the underlying mechanism.

Modelos animales ex vivo e in vivo para lesiones mecánicas y químicas del epitelio corneal

Corneal injury to the ocular surface, including chemical burn and trauma, may cause severe scarring, symblepharon, corneal limbal stem cells deficiency, ...