The authors have no acknowledgements.

Estes protocolos seguir as orientações do comitê de ética humana de nossa instituição. 1. convencional de córnea Colágeno CXL (C-CXL) 1. Preparação do Paciente <ol><li> 5 dias antes da cirurgia, coloque 1% pilocarpina gotas duas vezes por dia no olho tratado. </li><li> Na sala de operações, em condições assépticas, deitar o paciente em seu / sua volta. </li><li> Administrar anestesia tópica, como oxibuprocaína 0,4%. </li><li> Limpe o olho ea pele ao redor dos olhos com iodo anti-séptico duas vezes. </li><li> Use um espéculo tampa para manter o olho aberto. </li></ol> 2. A remoção epitelial <ol><li> Marque a central de 9,0 milímetros da córnea com um marcador de córnea círculo. </li><li> Retire as centrais 7,0 a 9,0 mm de epitélio corneano por desbridamento mecânico usando uma espátula sem corte. </li></ol> 3. Aplicação Riboflavina <ol><li> Aplicar 0,1% riboflavina com 20% de Dextrano na th<html> e córnea cada min durante 20 min. </li></ol> A irradiação UVA 4. <ol><li> Irradiar a córnea com um comprimento de onda de luz de UVA 370 nm, a uma irradiação de 3 mW / cm2 (5,4 J / cm2 de superfície da dose) e a uma distância de trabalho de 5 cm, durante 30 min. </li></ol><img alt="figura 1" src="/files/ftp_upload/53119/53119fig1.jpg" /> Figura 1:. Irradiação UVA em C-CXL A córnea é irradiada com uma luz de UVA de comprimento de onda de 370 nm com uma irradiação de 3 mW / cm2 (5,4 J / cm2 dose superficial) e a uma distância de trabalho de 5 cm, durante 30 minutos. <a href="https://www.jove.com/files/ftp_upload/53119/53119fig1large.jpg" target="_blank">Por favor clique aqui para ver uma versão maior desta figura.</a> <ol start="2"><li> Durante a irradiação, aplicar gotas de riboflavina para a córnea a cada 5 min. </li><li> Durante a irradiação, adicionar anestesia tópica (oxibuprocaína a 0,4%), se necessário. </li></ol>&quot;&gt; 5. Término da cirurgia <ol><li> Coloque gotas de antibiótico tobramicina (0,3%) e lágrimas artificiais (hialuronato cai 0,18%) para o olho operado. </li><li> Colocar uma lente de contacto macia atadura no final da cirurgia até re-epitelização completa é. Reepitelização normalmente leva 3 dias. </li><li> Prescrever analgésicos tais como o paracetamol (500 mg) mais codeína (30 mg), 6 comprimidos por dia. </li><li> Depois da córnea reepitelização, iniciar a terapia tópica com esteróides (dexametasona tópica de 1 mg / ml) e continuar por 3-4 semanas. Além disso, usar lágrimas artificiais 4 vezes por dia durante um mês. </li></ol> 2. Aceleração Corneal Collagen CXL (A-CXL) 1. Preparação do Paciente <ol><li> 5 dias antes da cirurgia, coloque 1% pilocarpina gotas duas vezes por dia no olho tratado. </li><li> Na sala de operações, em condições assépticas, deitar o paciente em seu / sua volta. </li><li> Administrar anestesia tópica, como oxibuprocaína 0,4%. </li><li> Limpe o evós ea pele ao redor dos olhos com iodo anti-séptico duas vezes. </li><li> Use um espéculo tampa para manter o olho aberto. </li></ol> 2. A remoção epitelial <ol><li> Marque a central de 9,0 milímetros da córnea com um marcador de córnea círculo </li><li> Retire as centrais 7,0 a 9,0 mm de epitélio corneano por desbridamento mecânico usando uma espátula sem corte. </li></ol> 3. Aplicação Riboflavina <ol><li> Aplicar 0,1% de dextrano sem riboflavina na córnea cada 2 min durante 10 min. </li></ol> A irradiação UVA 4. <ol><li> Irradiar a córnea com um comprimento de onda de luz de UVA 370 nm, a uma irradiação de 30 mW / cm 2 (5,4 J / cm2 de superfície da dose) e a uma distância de trabalho de 5 cm durante 3 min. </li><li> Durante a irradiação, adicionar anestesia tópica (oxybuproca &amp; # 239; ne 0,4%) se necessário. </li></ol> 5. final da cirurgia <ol><li> Coloque gotas de antibiótico tobramicina (0,3%) e lágrimas artificiais (hialuronato de gotas 0<html> 0,18%) para dentro do olho operado. </li><li> Colocar uma lente de contacto macia atadura no final da cirurgia até re-epitelização completa é. Reepitelização normalmente leva 3 dias. </li><li> Prescrever analgésicos tais como o paracetamol (500 mg) mais codeína (30 mg), 6 comprimidos por dia. </li><li> Depois da córnea reepitelização, iniciar a terapia tópica com esteróides (dexametasona tópica de 1 mg / ml) e continuar por 3-4 semanas. Além disso, usar lágrimas artificiais 4 vezes por dia durante um mês. </li></ol> 3. A iontoforese (I-CXL) 1. Preparação do Paciente <ol><li> 5 dias antes da cirurgia, coloque 1% pilocarpina gotas duas vezes por dia no olho tratado. </li><li> Na sala de operações, em condições assépticas, deitar o paciente em seu / sua volta. </li><li> Administrar anestesia tópica, como oxibuprocaína 0,4%. </li><li> Limpe o olho ea pele ao redor dos olhos com iodo anti-séptico duas vezes. </li><li> Use um espéculo tampa para manter o olho aberto. </li></ol><html> _step &quot;&gt; 2. Posicione o dispositivo de iontoforese. <ol><li> Aplicar o eléctrodo passivo pegajoso na testa sob o campo operatório. </li><li> Aplique o eletrodo ativo, um anel de sucção, para o olho aberto. Centrar o anel de sucção na periferia da córnea antes de libertar a sucção. </li></ol><img alt="Figura 2" src="/files/ftp_upload/53119/53119fig2.jpg" /> Figura 2. dispositivo de iontoforese. O eletrodo passivo é aplicado na testa sob o campo operatório eo eletrodo ativo, um anel de sucção, é aplicada ao olho aberto. <a href="https://www.jove.com/files/ftp_upload/53119/53119fig2large.jpg" target="_blank">Por favor clique aqui para ver uma versão maior desta figura.</a> 3. Aplicação Riboflavina <ol><li> Encha o anel de sucção com hypoosmolar 0,1% riboflavina sem dextrano. </li></ol><html> jpg &quot;/&gt; Figura 3. aplicação de riboflavina em I-CXL. O anel de sucção é preenchido com hypoosmolar riboflavine 0,1% sem dextrano. <a href="https://www.jove.com/files/ftp_upload/53119/53119fig3large.jpg" target="_blank">Por favor clique aqui para ver uma versão maior desta figura.</a> <ol start="2"><li> Comece a corrente eléctrica de 0,2 mA e a aumentar gradualmente a 1,0 mA durante um tempo total de iontoforese 5 min (Figura 4). </li></ol><img alt="Figura 4" src="/files/ftp_upload/53119/53119fig4.jpg" /> Figura 4. dispositivo de iontoforese para a penetração de riboflavina. A corrente eléctrica é inicialmente de 0,2 mA e gradualmente aumentada para 1,0 mA. O tempo total de iontoforese é de 5 minutos. <a href="https://www.jove.com/files/ftp_upload/53119/53119fig4large.jpg" target="_blank">Por favor clique aqui para ver uma versão maior desta figura.</a> ep &quot;&gt; 4. A irradiação UVA <ol><li> Irradiar a córnea com um comprimento de onda de luz de UVA 370 nm, a uma irradiação de 10 mW / cm 2 (5,4 J / cm2 de superfície da dose) e a uma distância de trabalho de 5 cm durante 9 min. </li><li> Durante a irradiação, adicionar anestesia tópica (oxybuproca &amp; # 239; ne 0,4%) se necessário. </li></ol> 5. final da cirurgia <ol><li> Coloque gotas de antibiótico (tobramicina 0,3%) e lágrimas artificiais (hialuronato cai 0,18%) para o olho operado. </li><li> Prescrever analgésicos tais como o paracetamol (500 mg) mais codeína (30 mg), 6 comprimidos por dia. </li><li> Após a cirurgia, iniciar a terapia tópica com esteróides (dexametasona tópica de 1 mg / ml) e continuar por 3-4 semanas. Além disso, usar lágrimas artificiais 4 vezes por dia durante um mês. </li></ol>

<ol>
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S., 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=Pharmacokinetics+and+safety+of+transcorneal+iontophoresis+of+tobramycin+in+the+rabbit.">Pharmacokinetics and safety of transcorneal iontophoresis of tobramycin in the rabbit.</a> Invest Ophthalmol Vis Sci. 29 (9), 1397-1401 (1998).</li><li>Vinciguerra, P., 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=Transepithelial+iontophoresis+corneal+collagen+cross-linking+for+progressive+keratoconus:+initial+clinical+outcomes.">Transepithelial iontophoresis corneal collagen cross-linking for progressive keratoconus: initial clinical outcomes.</a> J Refract Surg. 30 (11), 746-753 (2014).</li><li>Caporossi, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Riboflavin-UVA-induced+corneal+collagen+cross-linking+in+pediatric+patients.">Riboflavin-UVA-induced corneal collagen cross-linking in pediatric patients.</a> Cornea. 31 (3), 227-231 (2012).</li><li>Buzzonetti, L., Petrocelli, G., Valente, P., Larossi, G., Ardia, R., Petroni, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Iontophoretic+transepithelial+corneal+cross-linking+to+halt+keratoconus+in+pediatric+cases:+15-month+follow-up.">Iontophoretic transepithelial corneal cross-linking to halt keratoconus in pediatric cases: 15-month follow-up.</a> Cornea. 34 (5), 512-515 (2015).</li><li>Baiocchi, S., Mazzotta, C., Cerretani, D., Caporossi, T., Caporossi, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Corneal+crosslinking:+riboflavin+concentration+in+corneal+stroma+exposed+with+and+without+epithelium.">Corneal crosslinking: riboflavin concentration in corneal stroma exposed with and without epithelium.</a> J Cataract Refract Surg. 35 (5), 893-899 (2009).</li><li>Wollensak, G., Iomdina, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biomechanical+and+histological+changes+after+corneal+crosslinking+with+and+without+epithelial+debridement.">Biomechanical and histological changes after corneal crosslinking with and without epithelial debridement.</a> J Cataract Refract Surg. 35 (3), 540-546 (2009).</li><li>Soeters, N., Wisse, R. 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 usando irradiação UVA e riboflavina é o tratamento padrão para deter a progressão do ceratocone. A riboflavina é um fotossensibilizador que induz ligações covalentes (ligações cruzadas) químicos quando irradiado com UVA 3. Na córnea, este fenómeno cria ligações cruzadas entre as fibras de colagénio corneal que aumentam a rigidez. Embora esse fenômeno é bem descrito, até agora não houve nenhuma evidência direta de intracorneais ligações cruzadas. No entanto, vários estudos têm rela...

O ceratocone é uma ectasia corneana bilateral progressiva e geralmente relatada em 1 em 2.000 na população em geral, resultando em uma modificação da forma da córnea e, portanto, diminuição da visão 2. Ceratocone é normalmente presentes no início da puberdade e progride até a terceira para a quarta década de vida, quando a doença normalmente tende a se estabilizar, embora a progressão pode ser variável ao longo da vida de um paciente. Por travar a progressão do ceratocone, reticulação visa adiar ou evitar a ceratoplastia. Até o momento, o único tratamento eficiente e seguro de ceratocone progressivo comprovado em estudos clínicos é o (C-CXL) protocolo convencional colágeno corneal de reticulação, que visa aumentar a rigidez e, portanto, deter a progressão do ceratocone 3-8. A fim de reduzir o tempo de operação e outros factores de risco possível de C-CXL, tais como ceratite infecciosa ou estromal neblina 9, vários protocolos melhorados têmForam descritos. Em primeiro lugar, em CXL acelerado (A-CXL), uma irradiação UVA é mais elevado de entregues na córnea durante um tempo reduzido 10. Em segundo lugar, para evitar a necessidade de desbridamento epitelial, transepiteliais abordagens têm sido empregadas. Infelizmente, eles têm sucesso limitado quando comparado com o protocolo convencional 11. O método mais recente transepitelial para entrega riboflavina corneana durante a iontoforese é CXL (I-CXL), mas uma avaliação rigorosa de este tratamento ainda não foi realizado 12. A iontoforese é uma técnica não-invasiva, em que é aplicada uma pequena corrente eléctrica para melhorar a penetração de uma droga ionizada através de um tecido. Em CXL por iontoforese, a riboflavina é ionizado para penetrar na córnea através do epitélio. Na microscopia confocal in vivo (IVCM) é um método de imagiologia da córnea que pode destacar as alterações celulares anormais de córneas em doenças tais como ceratocone 13. Com efeito, IVCMtem demonstrado alterações para todas as camadas da córnea no ceratocone com uma redução em particular na densidade do plexo do nervo sub-basal e ceratócitos de estroma 13-15. Além disso, IVCM tem provado ser altamente conveniente para análise microestrutural da córnea após C-16 CXL. A linha de demarcação da córnea é descrito como uma linha hyperreflective visto no segmento anterior tomografia de coerência óptica (AS OCT) 1 mês após o C-CXL a uma profundidade de 300 mm 17,18. IVCM seguinte C-CXL fornece informação sobre as alterações estruturais da córnea, incluindo a ausência de ceratócitos de córnea a uma profundidade de 300 um. A profundidade desta zona acelular, bem como a profundidade da linha de demarcação dentro do estroma da córnea revelou em AS PTU, parece estar associada com a profundidade efectiva de tratamento CXL 19, e medição da profundidade linha de demarcação da córnea em AS PTU 1 mês depois CXL tem sido proposto como um clínico eficientemétodo para a avaliação da eficácia CXL 18. No presente estudo investigamos a eficiência de três diferentes protocolos de crosslinking de colágeno corneano (convencionais, acelerado, e iontoforese), utilizando a medição da linha do estroma corneal demarcação por AS outubro e microscopia confocal. Nós, além disso, utilizado IVCM analisar quantitativamente mudanças microestruturais da córnea após os três tratamentos.

<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

O ceratocone é uma ectasia corneana bilateral e progressiva. A fim de retardar sua progressão, córnea colágeno cross-linking (CXL) foi recentemente introduzido como uma opção de tratamento eficaz. Em ciências biológicas e químicas, reticulação refere-se a novas ligações químicas formadas entre moléculas reativas. Assim, o objectivo da LCX colagénio corneal é aumentar sinteticamente a formação de ligações cruzadas entre as fibras de colagénio no estroma da córnea. Apesar do facto de a eficiência do (C-CXL) CXL protocolo convencional já foi demonstrado em vários estudos clínicos, que poderia beneficiar de melhorias na duração do processo e remoção do epitélio da córnea. Assim, a fim de fornecer uma avaliação coerente de dois protocolos novos e otimizados CXL, estudamos pacientes com ceratocone que tinham sido submetidos a um dos três tratamentos CXL: iontoforese (I-CXL), acelerou CXL (A-CXL) e CXL convencional ( C-CXL). A-6 CXL é um tempo de procedimento mais rápido CXL ucantar uma irradiação UVA dez vezes maior, mas ainda incluindo uma remoção epitélio. A iontoforese é uma técnica não-invasiva transepitelial em que é aplicada uma pequena corrente eléctrica para melhorar a penetração de riboflavina para a córnea. Usando segmento de tomografia de coerência óptica anterior (AS OCT) e microscopia confocal in vivo (IVCM), conclui-se que em relação à profundidade de penetração tratamento, protocolo CXL convencional continua sendo o padrão para o tratamento de ceratocone progressivo. CXL Acelerado parece ser uma alternativa rápida, segura e eficaz para tratar córneas finas. A utilização de iontoforese ainda está a ser investigada e devem ser consideradas com cautela maior.

<html> A linha de demarcação da córnea era visível no AS outubro em 92% dos casos, a uma profundidade média de 301,6 mm (SD, 73,6) <img alt="Figura 5" src="/files/ftp_upload/53119/53119fig5.jpg" /> Figura 5. Linha de Demarcação após C-CXL. De alta resolução segmento anterior da córnea de coerência óptica tomografia (AS OCT) visualizar a linha do estroma corneal demarcação a uma profundidade média de 358 mm (seta branc...

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).

Três diferentes protocolos de Corneal Collagen reticulação em Ceratocone: convencional, com aceleração e Iontoforese

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 visualizações.  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).

Vídeo: 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 coloração de ilhotas pancreáticas humanas

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

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.

Um ferimento Modelo Alkali queima de córnea neovascularização no Rato

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.

Um modelo de rato de Fibrilação Ventricular e Reanimação por Técnica de tórax fechado convencional

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.

Abordagens ecocardiograma e protocolos de Comprehensive Caracterização fenotípica de doença cardíaca valvular em ratos

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.

Indução de Aterosclerose Acelerada em Camundongos: O Modelo de "Lesão por Fio"

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.

Cicatrização de feridas abrasão epitelial da córnea com Burr Ocular como um modelo para 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.

Ressonância magnética cardíaca para avaliação de suspeita de trombo cardíaco: técnicas convencionais e 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.

Monitoramento simultâneo de PH do Esofágico Laryngophangeal e 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.

Implantação de andaimes de matriz de colágeno tridimensional como estratégia de regeneração 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 Animais Ex Vivo e In Vivo para Lesões Mecânicas e Químicas do Epitélio da Córnea

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