M.-A. Deutsch is supported by Dr. Rusche Forschungsprojekt (2014) Deutsche Stiftung f&#252;r Herzforschung and Deutsche Gesellschaft f&#252;r Thorax- Herz- und Gef&#228;&#223;chirurgie. M. Krane is supported by Deutsche Stiftung f&#252;r Herzforschung (F/37/11), Deutsches Zentrum f&#252;r Herz Kreislauf Forschung (DZHK B 13-050A; DZHK B 14-013SE) and Deutsche Forschungsgemeinschaft (KR3770/7-1; KR3770/9-1)

Toutes les expériences ont été approuvés par l&#39;Allemand Coeur Centre de Munich et le gouvernement de Bavière, Allemagne (Gz. 55.2-1-54-2532.3-17-13). Les animaux ont été logés dans un établissement exempt d&#39;organismes pathogènes spécifiques et ont reçu des soins humaine en conformité avec les «Principes de laboratoire Animal Care&quot; formulées par la Société nationale pour la recherche médicale et le «Guide pour le soin et l&#39;utilisation des animaux de laboratoire&quot; fournies par la National Academy des sciences et publié par les National Institutes of Health (NIH Publication no. 86-23, révisé 1985). REMARQUE: Un ensemble complet d&#39;instruments de microchirurgie stériles, y compris les deux pinces courbes et dilatateurs des vaisseaux, est nécessaire pour la procédure (Figure 2a). Antibioprophylaxie péri-opératoires ne sont pas systématiquement nécessaire mais peut être indiqué chez les animaux immunodéprimés. Les animaux ont été sacrifiés en utilisant l&#39;inhalation isoflurane terminal. Préparation des donateurs et des greffons récolte &lt;ol&gt; <li> Anesthésier l&#39;animal par voie intrapéritonéale (ip) injection de midazolam (5,0 mg / kg), de médétomidine (0,5 mg / kg) et de fentanyl (0,05 mg / kg de poids corporel). Placez la souris dans une position couchée sur le champ opératoire et de confirmer la tolérance chirurgicale par le réflexe de retrait de la pédale. </li><li> Désinfectez la peau trois fois avec de la chlorhexidine et effectuer une incision abdominale médiane. Rentrer les intestins avec gaze humide vers la gauche pour exposer la veine cave inférieure. </li><li> Injecter 50 UI d&#39;héparine dilué dans 0,4 ml de solution saline à travers la veine cave inférieure pour l&#39;anticoagulation. Inciser la cuve 1 min après héparinisation. </li><li> Ouvrez le diaphragme et effectuer une thoracotomie bilatérale pour exposer le coeur. La paroi thoracique antérieure se reflète supérieurement et maintenu en place avec une pince. </li><li> Retirez le thymus et carrément disséquer le tissu conjonctif entre l&#39;aorte et l&#39;artère pulmonaire. Inciser le droit veine cave supérieure. </li><li> Piquer l&#39;aorte ascendante mobilisées au niveau de latronc brachiocéphalique avec une aiguille 30 G monté sur une seringue de 5 ml et perfuser lentement le coeur retrogradually sans pression excessive avec 3 ml de 4 ° C à froid solution cardioplégique. Éviter l&#39;entrée d&#39;air dans la circulation coronaire. Déposez une solution saline glacée sur la greffe pour améliorer encore un arrêt cardiaque. </li><li> Transect l&#39;aorte proximale à l&#39;origine du tronc brachiocéphalique. </li><li> Ligaturer les veines caves inférieure et supérieure à la fois à proximité du cœur avec 8-0 soie et sectionner les distale pour les ligatures. </li><li> Disséquer gratuitement le tronc pulmonaire distale que possible. Ligaturer les artères pulmonaires à proximité de la bifurcation avec 8-0 soie et sectionner les distale. </li><li> Ligaturer les veines pulmonaires tous ensemble avec une seule ligature 8-0 de soie et transect eux. </li><li> Retirer le cœur du donneur et de le stocker dans une solution cardioplégique glacée jusqu&#39;à l&#39;implantation. </li></ol> 2. Bénéficiaire Préparation <ol><li> Préparerles poignets pour anastomoses en coupant le tube de polyimide avec un scalpel n ° 11 ou une paire de ciseaux au microscope. La coiffe est constituée d&#39;un corps cylindrique et un prolongement (1/3 de la circonférence) pour la mise en place de la pince vasculaire, chacune d&#39;une longueur de 1 mm (figure 2b). </li><li> Anesthésier l&#39;animal et confirmer la tolérance chirurgical comme décrit ci-dessus. Injecter 0,25 ml solution saline chaude voie sous-cutanée (sc) pour le remplacement des liquides. </li><li> Enlever les poils de la région cervicale droite en utilisant un rasoir électrique et placer l&#39;animal dans une position couchée sur un bloc chaud. Fixez les pieds avec des brins de bande tout en évitant surexploiter les membres antérieurs, car cela peut compromettre la respiration. Couvrir les yeux à l&#39;aide pommade ophtalmique pour prévenir la sécheresse. Désinfecter le site chirurgical à trois reprises avec de la chlorhexidine. </li><li> Faire une incision de la peau transversale de l&#39;angle mandibulaire droit à la veine jugulaire. </li><li> Carrément mobiliser la posteveine jugulaire ernal. Cauterize branches latérales avec une pince bipolaire et transect eux. </li><li> Divisez la veine jugulaire externe entre ligatures au niveau de la confluence avec la veine de la glande sous-maxillaire. </li><li> Tirer la veine jugulaire externe par l&#39;intermédiaire du manchon et de manière proximale occlure le vaisseau en plaçant une pince vasculaire sur le prolongement de la manchette. Retirer la ligature et irriguer la lumière du vaisseau avec une solution saline héparinée 01h10. </li><li> Evert la paroi du vaisseau sur la manchette et le fixer avec un préréglage circonférentielle 8-0 ligature de soie. </li><li> Retirer la partie superficielle du muscle sterno. </li><li> Mobiliser le droit artère carotide commune et sectionner le navire entre ligatures en dessous de la bifurcation carotidienne. </li><li> Tirer la carotide par l&#39;intermédiaire du manchon et de manière proximale occlure le vaisseau en plaçant une pince Yasargil sur le prolongement de la manchette. Retirer la ligature et irriguer la lumière du vaisseau avec une solution saline héparinée 01h10. </li><li> Doucement Dilate la lumière artérielle avec un dilatateur des vaisseaux, Evert la paroi du vaisseau sur la manchette et le fixer avec un préréglage circonférentielle 8-0 ligature de soie. </li><li> Pour gagner assez d&#39;espace pour implantation du greffon, enlever le lobe droit de la glande sous-maxillaire. </li><li> Gardez le champ opératoire humide jusqu&#39;à l&#39;implantation. </li></ol> 3. implantation du greffon <ol><li> Placez la greffe chez le receveur région cervicale à l&#39;envers avec l&#39;aorte orienté médiane et l&#39;artère pulmonaire latéralement. </li><li> Tirez l&#39;aorte de la greffe sur le brassard avec l&#39;artère carotide et sécuriser l&#39;anastomose avec un périphérique 8-0 ligature de soie. </li><li> Inciser le tronc pulmonaire sur sa face antérieure et le tirer sur le brassard avec la veine jugulaire externe. Encore une fois sécuriser l&#39;anastomose avec un périphérique 8-0 ligature de soie. </li><li> Retirer la pince sur la veine jugulaire externe, à la suite par le desserrage de l&#39;artère carotide. Les cœurs se remplit immédiatement avec du sang. Pour améliorer reperfusion baisse de 37 ° C une solution saline chaude sur la greffe. Normalement, le coeur se développe rythme sinusal en une minute. Retirez l&#39;extension du brassard sur le site veineux. </li><li> Veiller à la position correcte de la greffe, anastomoses doit être exempte de tensions et de navires sans torsion. Ne pas les navires cautérisées de la peau, car cela pourrait nuire à la cicatrisation des plaies. Habituellement, l&#39;hémorragie arrête spontanément. Fermez la peau avec un seul 8-0 suture continue. </li></ol> 4. Soins postopératoires <ol><li> Injecter 0,25 ml chaude sc de solution saline pour le remplacement des liquides. </li><li> Pour contrer les stupéfiants injectent naloxone (1,2 mg / kg), le flumazénil (0,5 mg / kg) et atipamezol (2,5 mg / kg de poids corporel) sc laissé les animaux sur le pavé chaud jusqu&#39;à ce qu&#39;il ait repris conscience suffisante pour maintenir décubitus sternal. Observez l&#39;animal pendant au moins une heure supplémentaire pour enregistrer tout comportement anormal. Ne retournez pas un animal qui a subi une intervention chirurgicale à la compagnie d&#39;autres animaux jusqu&#39;à leur entière Recoverouge. </li><li> Fournir l&#39;analgésie préventive de la buprénorphine (0,05 mg / kg sc trois fois par jour) ou une analgésie alternatif selon un protocole institutionnel local pour une durée minimale de 72 heures après l&#39;intervention. </li><li> Le chirurgien et un vétérinaire devraient voir des animaux régulièrement. En cas de perte de poids&gt; 15%, de l&#39;apathie, de l&#39;enflure ou chirurgicales animaux d&#39;infection de côté sont euthanasiés en utilisant l&#39;inhalation isoflurane terminal. </li></ol>

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N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Assessment+of+graft+function+in+rodent+models+of+heart+transplantation.">Assessment of graft function in rodent models of heart transplantation.</a> Microsurgery. 28, 565-570 (2008).</li><li>Schramm, R., 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=The+subepicardial+microcirculation+in+heterotopically+transplanted+mouse+hearts:+an+intravital+multifluorescence+microscopy+study.">The subepicardial microcirculation in heterotopically transplanted mouse hearts: an intravital multifluorescence microscopy study.</a> J Thorac Cardiovasc Surg. 134, 210-217 (2007).</li><li>Liu, F., Kang, S. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Heterotopic+heart+transplantation+in+mice.">Heterotopic heart transplantation in mice.</a> J Vis Exp : JoVE. 238, (2007).</li><li>Gong, W., 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=Introduction+of+modified+cervical+cardiac+transplant+model+in+mice.">Introduction of modified cervical cardiac transplant model in mice.</a> Exp Clin Transplant. 10, 158-162 (2012).</li><li>Matsuura, A., Abe, T., Yasuura, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Simplified+mouse+cervical+heart+transplantation+using+a+cuff+technique.">Simplified mouse cervical heart transplantation using a cuff technique.</a> Transplantation. 51, 896-898 (1991).</li><li>Chen, H., Zhang, Y., Zheng, D., Praseedom, R. K., Dong, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Orthotopic+kidney+transplantation+in+mice:+technique+using+cuff+for+renal+vein+anastomosis.">Orthotopic kidney transplantation in mice: technique using cuff for renal vein anastomosis.</a> PloS one. 8, e77278 (2013).</li><li>Maglione, 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=A+novel+technique+for+heterotopic+vascularized+pancreas+transplantation+in+mice+to+assess+ischemia+reperfusion+injury+and+graft+pancreatitis.">A novel technique for heterotopic vascularized pancreas transplantation in mice to assess ischemia reperfusion injury and graft pancreatitis.</a> Surgery. 141, 682-689 (2007).</li><li>Sucher, R., 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=Orthotopic+hind-limb+transplantation+in+rats.">Orthotopic hind-limb transplantation in rats.</a> J Vis Exp : JoVE. , (2010).</li><li>Kamada, N., Calne, R. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Orthotopic+liver+transplantation+in+the+rat.+Technique+using+cuff+for+portal+vein+anastomosis+and+biliary+drainage.">Orthotopic liver transplantation in the rat. Technique using cuff for portal vein anastomosis and biliary drainage.</a> Transplantation. 28, 47-50 (1979).</li><li>Oldani, G., Lacotte, S., Morel, P., Mentha, G., Toso, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Orthotopic+liver+transplantation+in+rats.">Orthotopic liver transplantation in rats.</a> J Vis Exp : JoVE. , (2012).</li><li>Okazaki, 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=A+mouse+model+of+orthotopic+vascularized+aerated+lung+transplantation.">A mouse model of orthotopic vascularized aerated lung transplantation.</a> Am J Transplant. 7, 1672-1679 (2007).</li><li>Suzuki, H., Fan, L., Wilkes, D. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Development+of+obliterative+bronchiolitis+in+a+murine+model+of+orthotopic+lung+transplantation.">Development of obliterative bronchiolitis in a murine model of orthotopic lung transplantation.</a> J Vis Exp : JoVE. , (2012).</li><li>Zou, 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=Mouse+model+of+venous+bypass+graft+arteriosclerosis.">Mouse model of venous bypass graft arteriosclerosis.</a> Am J Pathol. 153, 1301-1310 (1998).</li><li>Zhou, Y., Gu, X., Xiang, J., Qian, S., Chen, Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+comparative+study+on+suture+versus+cuff+anastomosis+in+mouse+cervical+cardiac+transplant.">A comparative study on suture versus cuff anastomosis in mouse cervical cardiac transplant.</a> Exp Clin Transplant. 8, 245-249 (2010).</li><li>Niimi, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+technique+for+heterotopic+cardiac+transplantation+in+mice:+experience+of+3000+operations+by+one+surgeon.">The technique for heterotopic cardiac transplantation in mice: experience of 3000 operations by one surgeon.</a> J Heart Lung Transplant. 20, 1123-1128 (2001).</li><li>Arras, M., Autenried, P., Rettich, A., Spaeni, D., Rulicke, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Optimization+of+intraperitoneal+injection+anesthesia+in+mice:+drugs,+dosages,+adverse+effects,+and+anesthesia+depth.">Optimization of intraperitoneal injection anesthesia in mice: drugs, dosages, adverse effects, and anesthesia depth.</a> Comp Med. 51, 443-456 (2001).</li></ol>

The authors have nothing to disclose.

Nous avons réalisé plus de 200 greffes cardiaques avec un taux de réussite de plus de 90% dans divers paramètres expérimentaux. Avec la technique décrite ici, la procédure complète peut être effectuée dans les 60 minutes par un chirurgien expérimenté. En préparant les vaisseaux cervicaux avant le prélèvement d&#39;organes, le temps total greffon ischémie peut être limité à 20 min. Le temps d&#39;implantation (temps d&#39;ischémie chaude) est significativement plus courte en utilisant la technique de ...

Le modèle de la transplantation cardiaque est fréquemment utilisé pour enquêter sur I / R-blessure, le rejet d&#39;allogreffe, la vasculopathie post-transplantation et l&#39;efficacité des agents immunomodulateurs. Des modèles de souris combinent de nombreux avantages comme le immunologique connu et origines génétiques, la disponibilité de nombreuses souches consanguines et transgéniques expérimentales et les coûts relativement faibles. En 1973, une technique de abdominale transplantation cardiaque chez la souris a été décrite par Corry et al 1. Dans ce modèle greffons sont revascularisés par anastomoses de l&#39;aorte ascendante à l&#39;aorte abdominale du destinataire et de l&#39;artère pulmonaire principale à la veine cave inférieure end-to-côté. En 1991, une technique de suture du col pour une transplantation cardiaque chez la souris a été décrit par Chen et al deux. Dans ce modèle, une perfusion de l&#39;artère coronaire rétrograde est établie par anastomose de l&#39;artère carotide du bénéficiaire de l&#39;aorte ascendante du grarrière. Les drains veineux via le sinus coronaire de sang dans l&#39;oreillette droite, le ventricule droit et est éjecté par l&#39;artère pulmonaire dans le receveur veine jugulaire externe (Figure 1). Par rapport à la greffe implantation abdominale le modèle col de l&#39;utérus est moins invasive et l&#39;emplacement superficiel de la greffe permet une accessibilité très facile pour les examens de suivi supplémentaires (par exemple, la palpation, l&#39;échocardiographie, la microscopie à fluorescence intravitale) 3,4. Malgré les améliorations techniques 5,6, une utilisation plus généralisée de ce modèle a été limitée en raison de difficultés techniques pour réaliser des anastomoses suturées de petits navires avec une incidence élevée de fuite de l&#39;anastomose et la thrombose. En 1991, Matsuura et al. introduit la technique de manchon dans lequel la paroi du vaisseau est éversée sur un cylindre synthétique, ce qui facilite grandement le modèle de transplantation cardiaque du col 7. La technique était fucomplémen adopté pour différents modèles de transplantation expérimentales dont 8 rénale, pancréatique 9, branche 10, 11,12 hépatique, du poumon et de la transplantation 13,14 vasculaire 15. Par rapport à la technique de suture de la durée requise pour anastomoses et, par conséquent, le temps d&#39;ischémie chaude est plus courte et il ya des complications vasculaires beaucoup moins en utilisant la technique de la coiffe 16. En outre, il permet aux chirurgiens avec peu de formation en microchirurgie pour effectuer l&#39;opération avec un taux de réussite élevé. Un inconvénient de la technique du brassard est la ligature préalable de l&#39;artère carotide commune, ce qui peut modifier la perfusion cérébrale. Dans ce papier, nous présentons une vidéo technique de brassard améliorée et simplifiée pour cervicale transplantation cardiaque chez la souris.

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>Forceps JFC-7.2</td>
			<td>S&#38;T</td>
			<td>P-00036</td>
			<td>Curved tip 0.20 mm</td>
		</tr>
		<tr>
			<td>Vessel dilatator D-5a</td>
			<td>S&#38;T</td>
			<td>S-00124</td>
			<td></td>
		</tr>
		<tr>
			<td>Adventita scissor</td>
			<td>S&#38;T</td>
			<td>S-00102</td>
			<td></td>
		</tr>
		<tr>
			<td>Vascular clamp B-1V</td>
			<td>S&#38;T</td>
			<td>S-00396</td>
			<td></td>
		</tr>
		<tr>
			<td>Yasargil Clip</td>
			<td>Peter Lazic</td>
			<td>65,097</td>
			<td>3.5 mm lenght</td>
		</tr>
		<tr>
			<td>Bipolar forceps</td>
			<td>Micromed</td>
			<td>148-100-011</td>
			<td>tip 0.25 mm</td>
		</tr>
		<tr>
			<td>Polyimide tubes</td>
			<td>River Tech Medical</td>
			<td></td>
			<td>19-24 gauge</td>
		</tr>
		<tr>
			<td>8.0 silk ligatures</td>
			<td>Catgut</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Custodiol HTK solution</td>
			<td>Essential Pharmaceuticals</td>
			<td></td>
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heterotopic cervical heart transplantation in mice

The heterotopic cervical heart transplantation in mice is a valuable tool in transplant and cardiovascular research. The cuff technique greatly simplifies this model by avoiding challenging suture anastomoses of small vessels thereby reducing warm ischemia time. In comparison to abdominal graft implantation the cervical model is less invasive and the implanted graft is easily accessible for further follow-up examinations. Anastomoses are performed by pulling the ascending aorta of the graft over the cuff with the recipient&#8217;s common carotid artery and by pulling the main pulmonary artery over the cuff with the external jugular vein. Selection of appropriate cuff size and complete mobilization of the vessels are important for successful revascularization. Ischemia-reperfusion (I/R) injury can be minimized by perfusing the graft with a cardioplegic solution and by hypothermia. In this article, we provide technical details for a simplified and improved cuff technique, which should allow surgeons with basic microsurgical skills to perform the procedure with a high success rate.

Notre équipe pourrait effectuer avec succès la technique décrite après environ 20 chirurgies de formation. La partie la plus difficile est de retourner la paroi de la cuve de l&#39;artère carotide sur le brassard, parce qu&#39;il retourne facilement en arrière et de multiples tentatives peut conduire à la déchirure de la paroi. Après la période de formation, le temps opératoire total moyen était de 60 ± 8 min. Environ 20 minutes sont nécessaires pour la récolte des donateurs, 25 min pour la préparation d...

Watch this Scientific Journal Video about Heterotopic Cervical Heart Transplantation in Mice at JoVE.com

Heterotopic Cervical Heart Transplantation in Mice

The cuff technique shortens and facilitates the model of heterotopic cervical heart transplantation in mice by avoiding technically challenging suture anastomoses of small vessels. The technical details shown in this video paper should allow researches to establish this model in their laboratories.

Heterotopic col de l&#39;utérus chez la souris Transplantation cardiaque

The heterotopic cervical heart transplantation in mice is a valuable tool in transplant and cardiovascular research. The cuff technique greatly simplifies ...

heterotopic-cervical-heart-transplantation-in-mice

Research

JoVE Journal

Medicine

11.1K vues.  German Heart Center Munich, Technische Universität München. The cuff technique shortens and facilitates the model of heterotopic cervical heart transplantation in mice by avoiding technically challenging suture anastomoses of small vessels. The technical details shown in this video paper should allow researches to establish this model in their laboratories.

Vidéo: Heterotopic Cervical Heart Transplantation in Mice

Controlled Cervical Laceration Injury in Mice

Use of genetically modified mice enhances our understanding of molecular mechanisms underlying several neurological disorders such as a spinal cord injury (SCI). Freehand manual control used to produce a laceration model of SCI creates inconsistent injuries often associated with a crush or contusion component and, therefore, a novel technique was developed. Our model of cervical laceration SCI has resolved inherent difficulties with the freehand method by incorporating 1) cervical vertebral stabilization by vertebral facet fixation, 2) enhanced spinal cord exposure, and 3) creation of a reproducible laceration of the spinal cord using an oscillating blade with an accuracy of &plusmn;0.01 mm in depth without associated contusion. Compared to the standard methods of creating a SCI laceration such as freehand use of a scalpel or scissors, our method has produced a consistent lesion. This method is useful for studies on axonal regeneration of corticospinal, rubrospinal, and dorsal ascending tracts.

A novel technique to create a reproducible in vivo model of cervical spinal cord laceration injury in the mouse is described. This technique is based on spine stabilization by fixation of the cervical facets and laceration of the spinal cord using an oscillating blade with an accuracy of &plusmn;0.01 mm.

Contrôlé blessures de lacération du col chez la souris

Use of genetically modified mice enhances our  understanding of molecular mechanisms underlying several neurological disorders  such as a spinal cord ...

Recherche

Médecine

Murine Cervical Heart Transplantation Model Using a Modified Cuff Technique

Mouse models are of special interest in research since a wide variety of monoclonal antibodies and commercially defined inbred and knockout strains are available to perform mechanistic in vivo studies. While heart transplantation models using a suture technique were first successfully developed in rats, the translation into an equally widespread used murine equivalent was never achieved due the technical complexity of the microsurgical procedure. In contrast, non-suture cuff techniques, also developed initially in rats, were successfully adapted for use in mice1-3. This technique for revascularization involves two major steps I) everting the recipient vessel over a polyethylene cuff; II) pulling the donor vessel over the formerly everted recipient vessel and holding it in place with a circumferential tie. This ensures a continuity of the endothelial layer, short operating time and very high patency rates4.
Using this technique for vascular anastomosis we performed more than 1,000 cervical heart transplants with an overall success rate of 95%. For arterial inflow the common carotid artery and the proximal aortic arch were anastomosed resulting in a retrograde perfusion of the transplanted heart. For venous drainage the pulmonary artery of the graft was anastomosed with the external jugular vein of the recipient5.
Herein, we provide additional details of this technique to supplement the video.

The murine cervical heart transplantation model is well suited for immunological as well as ischemia reperfusion injury studies. We modified the procedure using a non-suture cuff technique and performed more than 1,000 successful transplants with this approach.
Herein, we provide additional details of this technique to supplement the video.

Murin col de l&#39;utérus Cœur, la transplantation en utilisant une technique de modification de manchette

Mouse models are of special interest in research since a wide variety of monoclonal antibodies and commercially defined inbred and knockout strains are ...

A Modified Method for Heterotopic Mouse Heart Transplantion

Mice are often used as heart transplant donors and recipients in studies of transplant immunology due to the wide range of transgenic mice and reagents available. A difficulty is presented due to the small size of the animal and the considerable technical challenges of the microsurgery involved in heart transplantation. In particular, a high rate of technical failure early after transplantation may result from recipient death and post-operative complications such as hind limb paralysis or a non-beating heart. Here, the complete technique for heterotopic mouse heart transplantation is demonstrated, involving harvesting the donor heart and its subsequent implantation into a recipient mouse. The donor heart is harvested immediately following in situ perfusion with cold heparinized saline and transection of the ascending aorta and pulmonary artery. The recipient operation involves preparation of the abdominal aorta and inferior vena cava (IVC), followed by end-to-side anastomosis of the donor aorta with the recipient aorta using a single running 10-0 microsuture and a similar anastomosis of the donor pulmonary artery with the recipient IVC. Following the operation the animal is injected with 0.6 ml normal saline subcutaneously and allowed to recover on a 37&#160;&#176;C heating pad. The results from 227 mouse heart transplants are summarized with a success rate at 48 hr of 86.8%. Of the 13.2% failures within 48 hr, 5 (2.2%) experienced hind limb paralysis, 10 (4.4%) had a non-beating heart due to graft ischemic injury and/or thrombosis, while 15 (6.6%) died within 48 hr.

A technique is demonstrated for the microsurgical procedure for heterotopic transplantation of hearts in mice, including simplified methods for donor harvesting and recipient vessel anastomosis.

Une méthode modifiée de Heterotopic souris coeur la transplantation

Mice are often used as heart transplant donors and recipients in studies of transplant immunology due to the wide range of transgenic mice and reagents ...

Murine Heterotopic Heart Transplant Technique

It is now over forty years since this technique was first reported by Corry, Wynn and Russell. Although it took some years for other labs to become proficient in and utilize this technique, it is now widely used by many laboratories around the world. A significant refinement to the original technique was developed and reported in 2001 by Niimi. Described here are the techniques that have evolved over more than a decade in the hands of three surgeons (Plenter, Grazia, Pietra) in our center. These techniques are now being passed on to a younger generation of surgeons and researchers.
Based largely on the Niimi experience, the procedures used have evolved in the fine details - details which we will endeavor to relate here in such a way that others may be able to use this very useful model. Like Niimi, we have found that a video aid to learning is a priceless resource for the beginner.

The manuscript describes the steps required to perform the heterotopic heart transplant in the mouse.

Murin de transplantation cardiaque hétérotopique Technique

It is now over forty years since this technique was first reported by Corry, Wynn and Russell. Although it took some years for other labs to become ...

Transplantation of Pulmonary Valve Using a Mouse Model of Heterotopic Heart Transplantation

Tissue engineered heart valves, especially decellularized valves, are starting to gain momentum in clinical use of reconstructive surgery with mixed results. However, the cellular and molecular mechanisms of the neotissue development, valve thickening, and stenosis development are not researched extensively. To answer the above questions, we developed a murine heterotopic heart valve transplantation model. A heart valve was harvested from a valve donor mouse and transplanted to a heart donor mouse. The heart with a new valve was transplanted heterotopically to a recipient mouse. The transplanted heart showed its own heartbeat, independent of the recipient&#8217;s heartbeat. The blood flow was quantified using a high frequency ultrasound system with a pulsed wave Doppler. The flow through the implanted pulmonary valve showed forward flow with minimal regurgitation and the peak flow was close to 100 mm/sec. This murine model of heart valve transplantation is highly versatile, so it can be modified and adapted to provide different hemodynamic environments and/or can be used with various transgenic mice to study neotissue development in a tissue engineered heart valve.

In order to understand the cellular and molecular mechanisms underlying neotissue formation and stenosis development in tissue engineered heart valves, a murine model of heterotopic heart valve transplantation was developed. A pulmonary heart valve was transplanted to recipient using the heterotopic heart transplantation technique.

Transplantation de valve pulmonaire en utilisant un modèle de souris de transplantation cardiaque hétérotopique

Tissue engineered heart valves, especially decellularized valves, are starting to gain momentum in clinical use of reconstructive surgery with mixed ...

Rat Heterotopic Abdominal Heart/Single-lung Transplantation in a Volume-loaded Configuration

Herein, we describe a novel technique for heterotopic abdominal heart-lung transplantation (HAHLT) in rats. The configuration of the transplant graft involves anastomosis of donor inferior vena cava (IVC) to recipient IVC, and donor ascending aorta (Ao) to recipient abdominal Ao. The right upper and middle lung lobes are preserved and function as conduits for blood flow from right heart to left heart.
There are several advantages to using this technique, and it lends itself to a broad range of applications. Because the graft is transplanted in a configuration that allows for dyamic volume-loading, cardiac function may be directly assessed in vivo. The use of pressure-volume conductance catheters permits characterization of load-dependent and load-independent hemodynamic parameters. The graft may be converted to a loaded configuration by applying a clamp to the recipient&#8217;s infra-hepatic IVC. We describe modified surgical techniques for both donor and recipient operations, and an ideal myocardial protection strategy. Depending on the experimental aim, this model may be adapted for use in both acute and chronic studies of graft function, immunologic status, and variable ventricular loading conditions. The conducting airways to the transplanted lung are preserved, and allow for acute lung re-ventilation. This facilitates analysis of the effects of the mixed venous and arterial blood providing coronary perfusion to the graft.
A limitation of this model is its technical complexity. There is a significant learning curve for new operators, who should ideally be mentored in the technique. A surgical training background is advantageous for those wishing to apply this model. Despite its complexity, we aim to present the model in a clear and easily applicable format. Because of the physiologic similarity of this model to orthotopic transplantation, and its broad range of study applications, the effort invested in learning the technique is likely to be worthwhile.

We describe a novel technique for heterotopic abdominal heart-lung transplantation (HAHLT) in rats. The transplant configuration results in a partially loaded graft circulation, allowing direct functional assessment. This model may be employed for acute or chronic studies of function and immunologic status of the transplanted graft.

Rat Heterotopic abdominale Coeur / Transplantation pulmonaire simple dans une configuration de volume chargé

Herein, we describe a novel technique for heterotopic abdominal heart-lung transplantation (HAHLT) in rats. The configuration of the transplant graft ...

A Novel Microsurgical Model for Heterotopic, En Bloc Chest Wall, Thymus, and Heart Transplantation in Mice

Exploration of novel strategies in organ transplantation to prolong allograft survival and minimizing the need for long-term maintenance immunosuppression must be pursued. Employing vascularized bone marrow transplantation and co-transplantation of the thymus have shown promise in this regard in various animal models.1-11 Vascularized bone marrow transplantation allows for the uninterrupted transfer of donor bone marrow cells within the preserved donor microenvironment, and the incorporation of thymus tissue with vascularized bone marrow transplantation has shown to increase T-cell chimerism ultimately playing a supportive role in the induction of immune regulation. The combination of solid organ and vascularized composite allotransplantation can uniquely combine these strategies in the form of a novel transplant model. Murine models serve as an excellent paradigm to explore the mechanisms of acute and chronic rejection, chimerism, and tolerance induction, thus providing the foundation to propagate superior allograft survival strategies for larger animal models and future clinical application. Herein, we developed a novel heterotopic en bloc chest wall, thymus, and heart transplant model in mice using a cervical non-suture cuff technique. The experience in syngeneic and allogeneic transplant settings is described for future broader immunological investigations via an instructional manuscript and video supplement.

To study combined solid organ and vascularized composite allotransplantation, we describe a novel heterotopic en bloc chest wall, thymus, and heart transplant model in mice using a cervical non-suture cuff technique.

A Novel Microsurgical Modèle pour Heterotopic, en bloc de la paroi thoracique, Thymus, et Transplantation cardiaque chez la souris

Exploration of novel strategies in organ transplantation to prolong allograft survival and minimizing the need for long-term maintenance immunosuppression ...

A Simplified Model for Heterotopic Heart Valve Transplantation in Rodents

There is an urgent clinical need for heart valve replacements that can grow in children. Heart valve transplantation is proposed as a new type of transplant with the potential to deliver durable heart valves capable of somatic growth with no requirement for anticoagulation. However, the immunobiology of heart valve transplants remains unexplored, highlighting the need for animal models to study this new type of transplant. Previous rat models for heterotopic aortic valve transplantation into the abdominal aorta have been described, though they are technically challenging and costly. For addressing this challenge, a renal subcapsular transplant model was developed in rodents as a practical and more straightforward method for studying heart valve transplant immunobiology. In this model, a single aortic valve leaflet is harvested and inserted into the renal subcapsular space. The kidney is easily accessible, and the transplanted tissue is securely contained in a subcapsular space that is well vascularized and can accommodate a variety of tissue sizes. Furthermore, because a single rat can provide three donor aortic leaflets and a single kidney can provide multiple sites for transplanted tissue, fewer rats are required for a given study. Here, the transplantation technique is described, providing a significant step forward in studying the transplant immunology of heart valve transplantation.

This protocol describes a simple and efficient method for the transplantation of aortic valve leaflets under the renal capsule to allow for the study of alloreactivity of heart valves.

Un modèle simplifié pour la transplantation valvulaire cardiaque hétérotopique chez les rongeurs

There is an urgent clinical need for heart valve replacements that can grow in children. Heart valve transplantation is proposed as a new type of ...

A Porcine Heterotopic Heart Transplantation Protocol for Delivery of Therapeutics to a Cardiac Allograft

Cardiac transplantation is the gold standard treatment for end-stage heart failure. However, it remains limited by the number of available donor hearts and complications such as primary graft dysfunction and graft rejection. The recent clinical use of an ex vivo perfusion device in cardiac transplantation introduces a unique opportunity for treating cardiac allografts with therapeutic interventions to improve function and avoid deleterious recipient responses. Establishing a translational, large-animal model for therapeutic delivery to the entire allograft is essential for testing novel therapeutic approaches in cardiac transplantation. The porcine, heterotopic heart transplantation model in the intraabdominal position serves as an excellent model for assessing the effects of novel interventions and the immunopathology of graft rejection. This model additionally offers long-term survival for the pig, given that the graft is not required to maintain the recipient's circulation. The aim of this protocol is to provide a reproducible and robust approach for achieving ex vivo delivery of a therapeutic to the entire cardiac allograft prior to transplantation and provide technical details to perform a survival heterotopic transplant of the ex vivo perfused heart.

We present a protocol for utilizing a normothermic ex vivo sanguinous perfusion system for the delivery of therapeutics to an entire cardiac allograft in a porcine heterotopic heart transplant model.

Protocole de transplantation cardiaque hétérotopique porcine pour l’administration de produits thérapeutiques à une allogreffe cardiaque

Cardiac transplantation is the gold standard treatment for end-stage heart failure. However, it remains limited by the number of available donor hearts ...

A Modified Cuff Technique for Mouse Cervical Heterotopic Heart Transplantation Model

Cardiac allograft rejection limits the long-term survival of patients after heart transplantation. A mouse heart transplantation model is ideal for investigating the mechanism of cardiac allograft rejection in preclinical studies because of their high homology with human genes. This understanding would help develop unique approaches to improving patients&#39; long-term survival treated with cardiac allografts. In a mouse model, abdominal donor heart implantation is commonly performed with an end-to-end&#160;anastomosis to the recipient&#39;s aorta and inferior vena cava using stitches. In this model, the donor&#39;s heart is implanted by end-to-end anastomosis to the recipient&#39;s carotid artery and jugular vein by the modified-Cuff technique. The transplantation surgery is performed without stitching and thus may increase the survival of the recipient since there is no interference with the blood supply and venous reflux of the lower body. This mouse model would help investigate the mechanisms underlying the immunological and pathological (acute/chronic) rejection of cardiac allografts.

In the present protocol, a mouse heart transplantation model is used for investigating the mechanism of cardiac allograft rejection. In this heterotopic heart transplantation model, operation efficiency is improved, and the survival of cardiac grafts is ensured by a cervical end-to-end anastomosis of heart implantation using a modified Cuff technique.

Une technique de brassard modifiée pour un modèle de transplantation cardiaque hétérotopique cervicale de souris

Cardiac allograft rejection limits the long-term survival of patients after heart transplantation. A mouse heart transplantation model is ideal for ...