We are supported by research grants from the National Natural Science Foundation of China (NO. 81400590), National Institutes of Health (U54 CA156735) and Takeda Pharmaceutical Company Ltd. (MA-NC-D-156).

Todas as experiências com animais foram aprovados pelo Comitê de Cuidado e Uso do animal Institucional. 1. Preparação do rato <ol><li> Use camundongos mais ~ 20 g no peso corporal ou superiores a 6 semanas de idade para a cirurgia. </li><li> Antes da cirurgia, dar camundongos um laboratório de comida e água ad libitum e manter em um ciclo claro-escuro de 00:12 h. </li><li> Raspar a área cirúrgica com uma máquina de cortar cabelo. Anestesiar ratos por injecção intraperitoneal de 80 mg / kg de cetamina e 12 mg / kg de xilazina. Esta dose coloca ratos no sono em poucos minutos e fornece anestesia suficiente para os seguintes procedimentos cirúrgicos. </li><li> Confirme anesthetization adequada com base na ausência do reflexo corneal e retração membro quando a pata é comprimido. Aplicar lubrificante pomada oftálmica nos olhos para evitar o ressecamento. </li><li> Esterilizar a pele com solução de betadine. </li></ol> 2. Modelo refluxo gástrico (Figura 1B) <ol><li> Realizar uma superiorincisão na linha média de ~ 2 cm a partir da xifóide apontando para o ânus. </li><li> Abra-se a cavidade abdominal através da linha média. Remova o xifóide para aumentar a exposição com um par de tesouras. </li><li> Separa-se e cortar os tecidos conjuntivos entre o fígado e o estômago. Ligadura e cortar o feixe vascular entre o baço e do fundo, a fim de completamente livre até o fundo. </li><li> Ligue o fundo ligeiramente para a esquerda e expor o lado esquerdo da junção gastro-esofágico no campo. Adicione uma incisão longitudinal de 5 mm no músculo distai ao longo do esófago utilizando um par de tesouras para expor o epitélio. </li><li> Corte aberto do epitélio ao longo da mesma direcção (incisão 1, Figura 1B). Adicione três incisões na pança de incisão (2, 3 e 4, Figura 1B) para eliminar a maioria da pança com uma tesoura afiada operacionais. Se o estômago não está vazio, remova cuidadosamente o conteúdo gástrico. </li><li> Coloque um Sutu prolene 8-0re (com um ponto de afunilamento agulha) através de um ponto (no esôfago) e ponto A &#39;(na pança) com uma precisão de mucosa da mucosa oposição. De igual modo lugar suturas através do ponto b e Ponto b &#39;, e outros pares de pontos, respectivamente. Uniformemente espaço estas suturas 3-4 mm uns dos outros. </li><li> Lava-se a cavidade abdominal com solução salina normal para limpar o sangue e o conteúdo gástrico. Feche a parede abdominal com fio de seda e pele com grampos metálicos. </li></ol> 3. Modelo de Refluxo mistos (Figura 1C) <ol><li> Siga os passos 2.1 a 2.4 para expor a junção gastroesofágico. </li><li> Suavemente separar o lado dorsal do esófago a partir dos vasos sanguíneos por trás do esófago. Passe uma ponta de algodão pequena entre os vasos sanguíneos e esôfago. Nota: O algodão pode ser parcialmente removida a partir da ponta para reduzir o seu tamanho. Esta dica de algodão serve a dois propósitos, levantando o esôfago e proteger os vasos sanguíneos. </li><li> Faça inc dois cinco milímetros longitudinalisions cada na junção gastro-esofágico e a extremidade proximal do duodeno adjacente ao piloro com uma tesoura afiada operacionais. Para a incisão no duodeno, evitar vasos sanguíneos e coloque na fronteira anti-mesentérica. </li><li> Anastomosar as incisões com mucosa com precisão de mucosas oposição interrompidas 8-0 suturas de Prolene. Normalmente coloque 3-4 suturas na face dorsal e 2-3 suturas na parte da frente. </li><li> Retire a ponta de algodão. Lava-se a cavidade abdominal com solução salina normal e fechar a parede abdominal e a pele. </li></ol> 4. Modelo Duodenal refluxo (Figura 1D) <ol><li> Siga o passo 3.1 a 3.4 para gerar misto de refluxo. </li><li> Com cuidado, levante o estômago para expor a sua parte de trás. Um lobo do fígado podem ser abrangidos pelo tecido conjuntivo entre o fígado e parte de trás do estômago. Corte cuidadosamente os tecidos conjuntivos, corretamente proteger o fígado, e expor os vasos sanguíneos no lado dorsal do esôfago. </li><li> Ligadura ecortar os vasos sanguíneos. Ligadura e cortado o esófago na junção gastroesofágico. </li><li> Ligar o duodeno no piloro. Ligadura e cortar o mesentério. Remover todo o estômago. </li><li> Lava-se a cavidade abdominal com solução salina normal. Fechar a parede abdominal e a pele. </li></ol> 5. O tratamento pós-cirúrgico <ol><li> Após a cirurgia manter a temperatura corporal com uma almofada de aquecimento. Só depois de recuperar decúbito esternal, coloque os ratos de volta com outros animais. </li><li> Dar antibióticos e analgésicos para prevenir a infecção e aliviar a dor. Injectar Baytril (10 mg / kg, ip, qd, durante 3 dias) para prevenir a infecção, e cloridrato de buprenorfina (0,05 mg / kg, sc, bid. Dois dias) como analgésico. O jejum não é necessário após a cirurgia. A dieta líquida ou mole pode ser dada. </li><li> Avaliar o estado de saúde geral sobre basis.If diários os ratos mostram os seguintes sinais (dramática perda de peso&gt; 15%, a falta de appetite, vocalização, descarregar a partir da boca, nariz ou olhos, gesto curvado, inatividade, atividade aliciamento anormal), sacrificar os ratos. </li></ol>

<ol>
	<li>Kandulski, A., Malfertheiner, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gastroesophageal+reflux+disease--from+reflux+episodes+to+mucosal+inflammation.">Gastroesophageal reflux disease--from reflux episodes to mucosal inflammation.</a> Nat Rev Gastroenterol Hepatol. 9 (1), 15-22 (2012).</li><li>Orlando, R. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+integrity+of+the+esophageal+mucosa.+Balance+between+offensive+and+defensive+mechanisms.">The integrity of the esophageal mucosa. Balance between offensive and defensive mechanisms.</a> Best Pract Res Clin Gastroenterol. 24 (6), 873-882 (2010).</li><li>Stairs, D. B., Kong, J., Lynch, J. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cdx+genes,+inflammation,+and+the+pathogenesis+of+intestinal+metaplasia.">Cdx genes, inflammation, and the pathogenesis of intestinal metaplasia.</a> Prog Mol Biol Transl Sci. 96, 231-270 (2010).</li><li>Spechler, S. J., Fitzgerald, R. C., Prasad, G. A., History Wang, K. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=molecular+mechanisms,+and+endoscopic+treatment+of+Barrett's+esophagus.">molecular mechanisms, and endoscopic treatment of Barrett's esophagus.</a> Gastroenterology. 138 (3), 854-869 (2010).</li><li>Fang, Y., 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=Cellular+origins+and+molecular+mechanisms+of+Barrett's+esophagus+and+esophageal+adenocarcinoma.">Cellular origins and molecular mechanisms of Barrett's esophagus and esophageal adenocarcinoma.</a> Ann N Y Acad Sci. 1300, 187-199 (2013).</li><li>Garman, K. S., Orlando, R. C., Chen, X. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Review:+Experimental+models+for+Barrett's+esophagus+and+esophageal+adenocarcinoma.">Review: Experimental models for Barrett's esophagus and esophageal adenocarcinoma.</a> Am J Physiol Gastrointest Liver Physiol. 302 (11), G1231-G1243 (2012).</li><li>Baruah, 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=Translational+research+on+Barrett's+esophagus.">Translational research on Barrett's esophagus.</a> Ann N Y Acad Sci. 1325, 170-186 (2014).</li><li>Lechpammer, M., 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=Flavopiridol+reduces+malignant+transformation+of+the+esophageal+mucosa+in+p27+knockout+mice.">Flavopiridol reduces malignant transformation of the esophageal mucosa in p27 knockout mice.</a> Oncogene. 24 (10), 1683-1688 (2005).</li><li>Hao, J., Liu, B., Yang, C. S., Chen, X. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gastroesophageal+reflux+leads+to+esophageal+cancer+in+a+surgical+model+with+mice.">Gastroesophageal reflux leads to esophageal cancer in a surgical model with mice.</a> BMC Gastroenterol. 9, 59 (2009).</li><li>Mari, L., 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=A+pSMAD/CDX2+complex+is+essential+for+the+intestinalization+of+epithelial+metaplasia.">A pSMAD/CDX2 complex is essential for the intestinalization of epithelial metaplasia.</a> Cell Rep. 7 (4), 1197-1210 (2014).</li><li>Fang, Y., 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=Gastroesophageal+reflux+activates+the+NF-kappaB+pathway+and+impairs+esophageal+barrier+function+in+mice.">Gastroesophageal reflux activates the NF-kappaB pathway and impairs esophageal barrier function in mice.</a> Am J Physiol Gastrointest Liver Physiol. 305 (1), G58-G65 (2013).</li><li>Chen, H., 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=Nrf2+deficiency+impairs+the+barrier+function+of+mouse+oesophageal+epithelium.">Nrf2 deficiency impairs the barrier function of mouse oesophageal epithelium.</a> Gut. 63 (5), 711-719 (2014).</li></ol>

One of the authors (XC) received funding that was provided by Takeda Pharmaceutical Company Ltd which supports basic research associated with gastroesophageal reflux disease. None of the reagents or instruments used in this article is produced by this company.

Vários modelos cirúrgicos foram criados para imitar gástrica, duodenal e misto de refluxo em roedores. Estes três procedimentos aqui descritos são adequados para experimentos de longo prazo com taxas razoáveis ​​de sobrevivência pós-operatória. Um pesquisador com experiência cirúrgica deve ser capaz de compreender a técnica, após um curto período de prática. A hemorragia pode resultar de injecção intraperitoneal de anestésicos antes da cirurgia, laceração do fígado d...

Gastroesophageal reflux disease (GERD) is a chronic disorder caused by the prolonged exposure of distal esophagus to gastric or gastroduodenal contents1. Prolonged exposure to these noxious refluxates impairs the intrinsic defenses within the esophageal epithelium and thus results in esophagitis2. Barrett&#8217;s esophagus arises in the setting of chronic reflux, and is a premalignant lesion with increased risk of esophageal adenocarcinoma3,4. Despite the clinical importance, the mechanisms of GERD, Barrett&#8217;s esophagus and adenocarcinoma have not been well understood. 
Animal models are essential for research on etiology, pathology, molecular mechanisms, prevention and treatment of human diseases. Up to date, various animal models of GERD, Barrett&#8217;s esophagus and adenocarcinoma have been developed using model animals5,6. Mouse esophagus is lined with stratified squamous epithelium which is histologically similar to that in human esophagus. Although a mouse esophagus is different from human esophagus in terms of keratinization and the absence of submucosal glands, the mouse is still an appealing model animal because of its relatively low cost of maintenance and its potential of sophisticated genetic modifications. Two approaches are commonly used to model GERD, Barrett&#8217;s esophagus and adenocarcinoma in mice: reflux surgery and genetic modification. Reflux surgery is the best way to induce reflux and genetic modifications mimics molecular alterations5,7. Reflux surgery can be combined with genetic modifications to further understand disease mechanisms8. 
Many surgical procedures have been reported by us and others6,9: (1) gastric reflux: pyloric ligation, pyloric constriction with forestomach ligation, Wendel cardioplasty, and esophagogastric anastomosis; (2) mixed reflux: esophagogastroduodenal anastomosis, esophagoduodenostomy (or esophagojejunostomy); (3) duodenal reflux: esophagogastroduodenal anastomosis plus gastrectomy; (4) reflux of chemical components: bilious reflux, pancreatic reflux, esophageal perfusion; and (5) esophageal transplantation5. Recently a microsurgical mouse model was reported to produce jejunal reflux via an esophagojejunostomy with magnets10. These surgical models have advantages over in vitro cell culture or organotypic culture models. In vitro, esophageal cells cannot tolerate a medium with high acidity or high concentrations of bile acids. Unconjugated bile acids which are commonly used to produce changes in esophageal epithelial cells in vitro are usually not present in the duodenal refluxate in vivo. Thus conclusions drawn from such in vitro studies should be taken with caution.
Surgery on the mouse esophagus remains a technical challenge because of its small size. A low rate of postoperative survival does not allow experiments which require certain sample size to reach statistically sound conclusions. In the past we have successfully developed and characterized surgical models of gastric reflux, mixed reflux, duodenal reflux with mice in long-term experiments9,11,12. We have also provided consultation to several other groups in their mouse surgery. Herein, we describe three surgical procedures in mice in order to help the community to establish these models in their labs.

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>Instruments</td>
			<td>Source</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Dumont #1 Forceps Dumostar Tip&#160;</td>
			<td>Roboz Surgical Instrument Co. (Gaithersburg, MD)</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Micro Clip Applying Forceps 5.5&#34;</td>
			<td>Roboz Surgical Instrument Co. (Gaithersburg, MD)</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Bonn Scissors 3.5&#34; Straight 15mm Sharp/Sharp Tungsten Carbide Blades</td>
			<td>Roboz Surgical Instrument Co. (Gaithersburg, MD)</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Operating Scissors 5.5&#34; Straight Sharp/Sharp SureCut</td>
			<td>Roboz Surgical Instrument Co. (Gaithersburg, MD)</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>4-0 Silk Black Braid 100 Yard Spool</td>
			<td>Roboz Surgical Instrument Co. (Gaithersburg, MD)</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Surgeon&#39;s Needle 1/2 Circle Cutting Edge Size 12 (25 mm Chord Length) Pack 12</td>
			<td>Roboz Surgical Instrument Co. (Gaithersburg, MD)</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Halsey Needle Holder 5&#34; Smooth</td>
			<td>Roboz Surgical Instrument Co. (Gaithersburg, MD)</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Micro Needle Holder 5.125&#34; Curved Lock .6mm</td>
			<td>Roboz Surgical Instrument Co. (Gaithersburg, MD)</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Reflex 9mm Wound Clip Applier</td>
			<td>Roboz Surgical Instrument Co. (Gaithersburg, MD)</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Reflex 9mm Wound Clips Box Of 100</td>
			<td>Roboz Surgical Instrument Co. (Gaithersburg, MD)</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>PRONOVA Poly (hexafluoropropylene-VDF) Suture 8-0</td>
			<td>Ethicon US, LLC</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Reagents</td>
			<td>Source</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Ringer&#39;s solution</td>
			<td>Henry Schein, Inc.</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>ketamine</td>
			<td>Henry Schein, Inc.</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>xylazine</td>
			<td>Henry Schein, Inc.</td>
			<td></td>
			<td></td>
		</tr>
	</tbody>
</table>

surgical models of gastroesophageal reflux with mice

Multiple surgical procedures have been reported to induce gastroesophageal reflux in animals. Herein, we report three surgical models with mice aiming to induce reflux of gastric contents, duodenal contents or mixed contents. Surgical procedures and general principles have been described in detail. A researcher with surgical experience should be able to grasp the technique after a short period of practice. After surgery, most mice can survive and develop reflux esophagitis similar to that in humans. However, it should be noted that histological differences between mouse and human esophagus are the inherent limitations of these surgical models. If used for research on Barrett&#8217;s esophagus and adenocarcinoma, these procedures may need to be combined with genetic modifications.

A maioria dos ratinhos (&gt; 95%) pode sobreviver à cirurgia. Durante o período perioperatório, as principais causas de morte incluem overdose de anestésicos, sangramento e razões desconhecidas. Quatro semanas após a cirurgia,&gt; 90% de ratinhos com refluxo gástrico ou misto de refluxo e&gt; 80% de ratinhos com refluxo duodenal podem sobreviver. Durante este período, os ratos morrem principalmente de estenose esofágica e incapacidade para comer. Estes ratos mostram sinais de estres...

Watch this Scientific Journal Video about Surgical Models of Gastroesophageal Reflux with Mice at JoVE.com

Surgical Models of Gastroesophageal Reflux with Mice

This article demonstrates surgical procedures of gastroesophageal reflux with mice. These models are useful tools for research on mechanisms and treatment of gastroesophageal reflux disease and potentially Barrett&#8217;s esophagus and esophageal adenocarcinoma.

Modelos cirúrgicos de refluxo gastroesofágico com Ratos

Multiple surgical procedures have been reported to induce gastroesophageal reflux in animals. Herein, we report three surgical models with mice aiming to ...

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10.0K visualizações.  Ningxia Medical University General Hospital. This article demonstrates surgical procedures of gastroesophageal reflux with mice. These models are useful tools for research on mechanisms and treatment of gastroesophageal reflux disease and potentially Barrett’s esophagus and esophageal adenocarcinoma.