マーク·J·カーンズは（医師とカナダの外科医のロイヤルカレッジオブを通じて受賞）UBC臨床医研究者プログラムからの支援を受け、UBC 4YF博士フェローシップ。 ジョン·H·ボイドは、ナショナルSanitorium協会と保健研究奨学生のためのマイケルスミス財団です。 CIHRを通じて資金調達。著者らは、灌流機器のセットアップやメンテナンスとの支援のために博士M.アラードとリチャードWamboldtに感謝したいと思います。

すべての動物を飼育し、実験動物の管理と使用のための国家と制度ガイドラインに従って世話しました。このプロトコルの倫理承認は、ブリティッシュコロンビア州の動物管理委員会の大学によって付与されました。 300との間体重の雄、SDラット - 450 gが、このプロトコルのために使用しました。 1.ドナー操作<ol><li> 3方活栓によって、長い静脈内（IV）カテーテル管に接続された円筒形フラスコに心臓麻痺（RT）の約100ミリリットルを持っています。重力によって心停止提供を可能にする、約80センチメートル作用表面の上にフラスコを上昇させるために、スタンドを使用してください。 </li><li>適切な構造を可視化するために、外科手術用ルーペのペアまたは解剖顕微鏡のどちらかを使用します。 注：我々は現在、3.4で双眼手術用顕微鏡を使用する - 21.3X倍率。 </li><li>麻酔室にドナーを置き、4で麻酔を誘導 - 5％イソフルラン。 </lI&gt; <li>オペレーティングプラットフォームにラットを移し、1とノーズコーンにより麻酔を維持 - 2％イソフルラン。乾燥を防ぐために、動物の目に獣医軟膏を適用します。 25 G針でミダゾラム（2ミリグラム/ kg）を腹腔内に投与します。 </li><li>手術用バリカンを使用して、剣状突起から下顎にドナーを剃ります。 （その後のIVアクセスのための）左鼠径部の小さな領域を剃ります。作用面に脱毛剤を塗布、約5分間待機し、ガーゼで髪を削除します。 </li><li>準備ポビドンヨードまたはchlorexidineベースのソリューションで手術部位（我々は唯一のクロルヘキシジンを使用します）。 0.5％リドカイン皮下 - 0.1と切開部位に潜入。 </li><li>麻酔深度とバイタルサインのモニタリングのための無料の右後肢を残して、粘着テープを使用してオペレーティング·プラットフォームに前肢と左後肢を固定します。 </li><li>ペダルピンチすることにより、適切な麻酔深度を確認した後、使用して、左鼠径部の折り目に切開を作ります22ブレードメス。鈍的切開によって、左大腿静脈を露出させ、次のように静脈アクセスを取得します。 <ol><li>静かに大腿静脈を覆う組織を撤回し、24のG IVで静脈にカニューレを挿入。生理食塩水で満たされたIVチューブの短い長さにIVを接続し、粘着テープで所定の位置にチューブを固定します。 注：ラットにおける大腿静脈カニューレ挿入のための手順はイェスペルセンや同僚14によって他の箇所に記載されています。 </li><li> IVチューブに生理食塩水を充填した10mLのシリンジを接続し、静かにIVの正確な位置決めを確実にするために血液を吸引します。 </li><li> IVチューブを通して未分画ヘパリンの500 IU、その後3でチューブをフラッシュ - - 300を注入し、生理食塩水5mlの。 </li></ol></li><li>次に、以下のようにドナーをtracheotomize： <ol><li> 22ブレードメスを用いて頸静脈ノッチと下顎の間の軟組織に正中切開を行います。 Metzenを使用して、正中線に甲状腺のカプセルに浸透バウムはさみ、および鈍的切開を使用して、そのローブを分離します。 </li><li>鈍的切開を使用して、気管の前面を露出させるために正中線にネックストラップの筋肉を分離します。 </li><li>ぶっきらぼうに気管の周りに周面を分析するために、湾曲鉗子を使用してください。 4-0絹のネクタイと気管を取り囲みます。 </li><li>アイリスはさみを使用して、前方気管内に横切開を行い、甲状軟骨の約5ミリメートル劣ります。静かに気管カニューレ（14 G IV）を導入し、4-0絹のネクタイを使用して所定の位置に固定します。 </li><li>人工呼吸器に気管カニューレを接続します。人工呼吸器回路を通る酸素およびイソフルランの流れをリダイレクトし、速度、重量13によって予測された一回換気量でドナーを換気。 </li></ol></li><li>胸の正中線で切開を行います（22ブレードメスを用いて）、xipisternum以下に頸静ノッチで切開部を拡張します。 </li><li>正中線での滞在、骨のカッターを用いて胸骨正中切開を行います。自己保持リトラクターと胸骨のエッジを後退させます。心膜および胸膜腔を入力します。 </li><li>胸腺摘出を行います。それは最初ぶっきらぼうに正中線で胸腺を分割し、その後鈍、鋭い切開の組み合わせを使用して、周囲の構造から分離するのが最も簡単です。 注：離れて優れた胸骨縁から胸腺を解剖時に内胸動脈の起源は、けがをする恐れがあります。出血を防ぐために、止血クリップは、これらの点で、胸腺を除去する前に適用してもよいです。 </li><li> Metzenbaumはさみを使用して、および/またはシャープラウアー、下大静脈（IVC）から離れた脂肪ペリ大静脈解剖。 IVCは、下方ダイヤフラムに、上方カーボ心房接合部から脂肪と結合組織の比較的自由であることを確認してください。 </li><li>シャープラウアーを使用して、円周方向に上大静脈（SVC）を解放し、4-0絹のネクタイとそれを取り囲みます。 </li><li>次に、解剖周囲の構造のない大静脈を離れ、近位および遠位4-0シルクネクタイとそれを連結します。左鎖骨下動脈を露出させるために静脈の介在部分を切除します。 </li><li>円周方向に鋭いラウアーを使用して、大動脈弓の血管を解放します。腕頭動脈の近位および遠位の外科手術用クリップを適用して、クリップの間にそれを分けます。左総頸動脈と左鎖骨下動脈の未クリップされたままにしておきます。 </li><li>次に、24 G IVカテーテルでSVCにカニューレを挿入。以前に配置された4-0絹のネクタイでカテーテルを固定します。 </li><li>好都合が、穏やかなグラフト収穫の準備をします。 </li><li>鋭いハサミを使用して、横隔膜面にだけ優れたIVCを分割。近くの気道構造が負傷されていないことを確認して、そのカニューレ挿入部位への優れたSVCを分割します。 </li><li>人工呼吸器の電源をオフにして、気管カニューレを外します。近位に気管を横断。 </li><li>鉗子で横に切開気管を持ち、心を削除肺エン圏。心肺移植片が除去されるように、これは鋭い解剖で穏やかなトラクションが必要になります。基本となる食道から気管を分離します。それの長い部分は収穫後そのまま残るように、下行大動脈を傷つけないように注意してください。 </li></ol>心肺移植片の調製注：この手順のこの部分を完了しながら、アシスタントが受信者のげっ歯類を麻酔し、移植片移植の準備をする必要があります。 <ol><li>下に向けて、心臓の前面に手術用ガーゼに心肺移植片を配置します。鋭い解剖を使用して、後部気道への損傷を回避すること、任意の残留食道を削除します。 </li><li>降順胸部大動脈の位置を確認し、16 Gの鈍先端のカニューレを挿入します。 4-0絹のネクタイで所定の位置にカニューレを固定します。 </li><li>大動脈弓の血管を見つけ、左鎖骨下動脈に外科用クリップを適用します。サブ国連クリップされ、左頸動脈を残しますシークエントの脱気。 </li><li> 16 G大動脈カニューレに心臓麻痺チューブを接続することにより、心停止の最初の用量を投与。頸動脈は一時的に心臓麻痺の適切な供給を確保するために鉗子で閉塞されなければなりません。 注：添付の大動脈カニューレを有することの利点は、所望のように心臓麻痺を連続的および/または断続的に投与され得ることです。 45秒 - 30以上の5ミリリットルのボーラスで15分 - 間欠投与のために、我々は、典型的には、すべての10心筋保護を提供します。 </li><li> Metzenbaumはさみおよび/または鋭いラウアーを使用して、離れて大動脈から周囲の大動脈脂肪を分析。大動脈は、その挿管サイトに左鎖骨下動脈（遠位弓）からの脂肪および結合組織の比較的自由であることを確認してください。 </li><li>次に、鋭いラウアーを使用して、左主気管支を露出し、4-0絹のネクタイと近位にそれを連結。アイリスのはさみを使用して、合字の左気管支の先端を横断。左肺を連結することにより、左肺全摘術を行います4-0シルクネクタイと動脈と静脈。合字に遠位横断し、左肺を削除します。 </li><li>上記のように、右上と中央ローブの例外を除いて、他のすべての肺葉を削除します。 lobectomiesを行う過程でIVCを負傷しないようにします。 </li><li>大動脈カニューレに心臓麻痺チューブを接続し、受信者が用意されている間、連続心臓麻痺を管理します。滅菌容器（ 例えば、ビーカー）で心肺移植片を配置します。 </li></ol> 3.受信者の操作<ol><li>上記、ドナー操作ごとに受信者を麻酔。乾燥状態から、動物の目を保護するために獣医軟膏を使用してください。ペダルピンチによって頻繁に麻酔を確認してください。 </li><li>ドナー操作ごとに動物を置き、この時間はバイタルサインと麻酔深度を監視すること自由に右前肢を残します。 </li><li>剣状突起からペニスに腹部を剃ります。 （その後のIVアクセスのための）左鼠径部の小さな領域を剃ります。適用します作用面に薬剤を脱毛、約5分間待機し、ガーゼで髪を削除します。 </li><li>準備ポビドンヨードまたはchlorexidineベースのソリューションで手術部位。 0.5％リドカイン皮下 - 0.1と切開部位に潜入。 </li><li> Tracheotomize上記指示されたように、受信者を換気。代わりに、ノーズコーン麻酔下での受信者を維持します。 </li><li>上述したように、大腿静脈にカテーテルを導入します。 IVチューブを通して未分画ヘパリンの500 IU、その後3でチューブをフラッシュ - - 300を注入し、生理食塩水5mlの。 </li><li>剣状突起から陰茎に22ブレードメスで腹部正中切開を行うことで、開腹術を行います。自己保持リトラクターを使用して腹壁を撤回。次に、上方およびラットの左側に腸を撤回。暖かい、生理食塩水に浸したガーゼでラップ。 </li><li>急激にその上の再を通して解剖によってIVCと腹部大動脈を露出させますTRO-腹膜脂肪。 </li><li>湾曲した血管のクランプが使用可能であり、準備ができています。 </li><li>円周方向に近位および遠位IVCと大動脈を解放し、4-0シルクネクタイとそれらを取り囲みます。これらのサイトとの間の空間の3センチメートル - 約2が必要です。 </li><li> IVCと大動脈の両方の十分な部分がクランプのジョーの上に露出していることを確認して、湾曲した血管のクランプを慎重に適用されます。 </li><li>生理食塩水を満たした1ml注射器に接続された25 G針でIVCの前壁の切開を行います。ドナーIVCのオリフィスの長さに合わせて、ポッツはさみで切開を拡張します。 </li><li>その容器から心肺移植片を取り外し、それは心停止から切断。静脈吻合を実行するために最適な位置に受信者の腹部に移植片を配置します。 注：腹部年代に向けて頂点ポインティングが下腹部を左に心肺移植片は、最終的には、少し斜めに配向することになります。 </li><li>かかとを固定とつま先は、9-0ナイロン縫合糸との吻合部の端部。縫合糸の長腕に取り付けられた針、およびそれ以降に接続するための縫合糸の短い方の端を残して、各端部に安全な結び目を作ります。心筋保護の用量を投与します。 </li><li>ファッションを実行するには、反対側の短い縫合糸アームに縫合線とネクタイの完全半分。心筋保護の用量を投与します。 </li><li>縫合線の他の半分を完了し、それを結びます。静脈吻合が完了しました。心筋保護の用量を投与</li><li>オリエントは、腹部のに向かって心の頂点ポインティングと心肺移植片は、下腹部を残しました。静脈吻合がよじれやねじれがないことを確認してください。 </li><li>テンションフリーと非ねじれた構成でIVCと大動脈吻合の両方を維持し、受信者の大動脈に到達するために必要とされるドナー大動脈の長さを評価します。 </li><li>アイリスハサミで（左鎖骨下動脈の遠位）下行大動脈を分割します。 </li><li> incisioを作りますnは生理食塩水を満たした1ml注射器に接続された25 G針でレシピエント大動脈の前壁で。ドナー大動脈オリフィスの長さに合わせて、ポッツはさみで切開を拡張します。 </li><li> 9-0ナイロン縫合糸を使用して、大動脈受信者にドナー大動脈のつま先（優れた側面）をアンカー。そして、大動脈の内側面に針を渡すとファッションの実行中に、吻合部の2分の1を完了します。 </li><li>大動脈吻合のかかとでは、縫合線（ステッチの方向の遷移）の向きを逆にし、ファッションを実行するには吻合部の横方向の半分を完了します。 </li><li>ゆっくりSVCにIVカテーテルを除去し、外科手術用クリップでSVCを連結。 </li><li>未クリップさ左総頸動脈を配置することにより脱空気大動脈。空気が自由に避難できるようになります直立姿勢で、オープン動脈を保持します。 </li><li>簡単に言えば、D（湾曲した血管クランプのジョーを開き、頸動脈が出血することができます電子空気）2 - 3秒。血管クランプを再適用します。 </li><li>頸動脈に外科用クリップを適用します。湾曲した血管のクランプを外します。 </li><li>縫合線での出血を確認してください。存在する場合、（出血の重症度に応じて）9-0ナイロン縫合糸の長さの外科ガーゼや修復に穏やかな圧縮を適用します。 </li><li>ドナー心臓は、数分以内に暴行を再開する必要があります。 </li><li>受信者のバイタルサイン、麻酔の深さに細心の注意を払い、必要に応じてボリュームの蘇生を管理します。 </li><li>実験のエンドポイントに応じて、動物の腹部を閉じて、それが麻酔から回復させ、または心血管評価のために動物を準備するのどちらか。 注：移植性評価の例としては、負荷に依存し、負荷に依存しない血行動態の in vivoでの対策、 インビトロランゲンドルフ関数の対策と作業心臓モード、（生存手術で）心エコーやMRIの調査に含まれています。 </li><li>に急性端末実験の終わりに、動物を放血によって安楽死させます。 </li></ol>

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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+simple+new+model+of+physiologically+working+heterotopic+rat+heart+transplantation+provides+hemodynamic+performance+equivalent+to+that+of+an+orthotopic+heart.">A simple new model of physiologically working heterotopic rat heart transplantation provides hemodynamic performance equivalent to that of an orthotopic heart.</a> J Heart Lung Transplant. 18 (10), 927-936 (1999).</li><li>Figueiredo, J. -. L., Nahrendorf, M., Sosnovik, D. E., Weissleder, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=MRI+of+a+novel+murine+working+heart+transplant+model.">MRI of a novel murine working heart transplant model.</a> Circ Heart Fail. 2 (3), 272-274 (2009).</li><li>Wen, P., Wang, X., 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+simple+technique+for+a+new+working+heterotopic+heart+transplantation+model+in+rats.">A simple technique for a new working heterotopic heart transplantation model in rats.</a> Transplant Proc. 45 (6), 2522-2526 (2013).</li><li>Didi&#233;, M., Biermann, D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preservation+of+left+ventricular+function+and+morphology+in+volume-loaded+versus+volume-unloaded+heterotopic+heart+transplants.">Preservation of left ventricular function and morphology in volume-loaded versus volume-unloaded heterotopic heart transplants.</a> A.Am J Physiol Heart Circ Physiol. 305 (4), H533-H541 (2013).</li><li>Gali&#241;anes, M., Zhai, X., Hearse, D. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effect+of+load+on+atrophy,+myosin+isoform+shifts+and+contractile+function:+studies+in+a+novel+rat+heart+transplant+preparation.">The effect of load on atrophy, myosin isoform shifts and contractile function: studies in a novel rat heart transplant preparation.</a> J Mol Cell Cardiol. 27 (1), 407-417 (1995).</li><li>Yokoyama, H., Ohmi, M., Murata, S., Nakame, T., Tabayashi, K., Mohri, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Proposal+of+a+working+left+heart+model+with+a+heterotopic+transplantation+technique+in+rats.">Proposal of a working left heart model with a heterotopic transplantation technique in rats.</a> J Heart Lung Transplant. 14 (4), 706-712 (1995).</li><li>Maruyama, T., Swartz, M. T., McBride, L. R., Pennington, D. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Working+heart+model+of+heterotopic+heart-lung+transplantation+in+rats.">Working heart model of heterotopic heart-lung transplantation in rats.</a> J Thorac Cardiovasc Surg. 107 (1), 210-215 (1994).</li><li>Pacher, P., Nagayama, T., Mukhopadhyay, P., B&#225;tkai, S., Kass, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Measurement+of+cardiac+function+using+pressure-volume+conductance+catheter+technique+in+mice+and+rats.">Measurement of cardiac function using pressure-volume conductance catheter technique in mice and rats.</a> Nat Protoc. 3 (9), 1422-1434 (2008).</li><li>Jespersen, B., Knupp, L., Northcott, C. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Femoral+arterial+and+venous+catheterization+for+blood+sampling,+drug+administration+and+conscious+blood+pressure+and+heart+rate+measurements.">Femoral arterial and venous catheterization for blood sampling, drug administration and conscious blood pressure and heart rate measurements.</a> J Vis Exp. (59), 3496-3410 (2012).</li><li>Habertheuer, A., Kocher, 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=Innovative,+simplified+orthotopic+lung+transplantation+in+rats).">Innovative, simplified orthotopic lung transplantation in rats).</a> J Surg Res. 185 (1), 419-425 (2013).</li></ol>

著者らは、開示することは何もありません。

ここに記載の技術の成功は、いくつかの要因に基づいて予測されます。それらの中のキーは、両方のドナーとレシピエント動物の安定性を確保し、安全かつ最小限の血液損失に関連付けられている細心の手術技術を採用し、均一なグラフト冷却との完全な心停止停止を確保し、総虚血時間を最小限に抑え、かつ適切に移植片を脱気されます。上記認めたように、技術者の技術的な複雑さは、?...

The first rodent model of heterotopic abdominal heart transplantation (HAHT) was described by Abbott and colleagues in 19641. This technique, and subsequent modifications have been widely applied to characterize transplant graft function and immunologic status. The majority of HAHT techniques described involve a non-volume loaded heart2,3. Models of HAHT involving volume-loaded ventricles have been described, but they are frequently limited in one or more respects.
Heterotopic abdominal heart-lung transplantation (HAHLT) with a volume-loaded left ventricle (LV) has been described previously. Chen and colleagues4, and subsequently Ibrahim and colleagues5 described HAHLT with a single aorto-aortic (donor ascending to recipient abdominal aorta) anastomosis. The only volume load presented to the ventricle in this circulation is the coronary venous return. Asfour and colleagues described a HAHT technique in which the lung circuit was eliminated by anastomosing donor pulmonary artery (PA) to donor left atrium (LA)6. In this circulation, venous inflow to right ventricle (RV) occurs via a donor SVC to recipient IVC anastomosis, and the subsequent LV load is ejected into the aorto-aortic anastomosis. Cardiac function was partially assessed in vivo, and also in vitro using a Langendorff rig. Figueiredo and colleagues described a HAHLT model similar to our own7, but in mice. Venous inflow to the RV occurs via donor SVC to recipient IVC anastomosis. Blood subsequently passes through the single lung circulation and LV load is ejected into the aorto-aortic anastomosis. Cardiac function in their study was assessed by magnetic resonance imaging (MRI). Wen and colleagues described a unique HAHT technique in which the LV is loaded by means of a recipient aorta to donor LA anastomosis8. The LV, therefore, fills at systemic pressures. Cardiac function, and whether LV stroke volume is ejected antegradely in their model was not assessed.
Many of the techniques referenced above involve non-physiologic LV loading conditions, including the techniques whose partial LV load is represented only by coronary venous return. On the other hand, many techniques do approach physiologic LV loading. The majority of these techniques, as with the technique of Asfour and colleagues, omit the pulmonary circulation and utilize a donor PA to donor LA anastomosis6,9. The circulation described by Galinanes and colleagues10 employs a direct recipient cava to donor LA anastomosis, omitting the pulmonary circulation and the right heart. Yokoyama and colleagues achieve the same effect by ligating the donor PA and creating an interatrial communication in the donor heart (omitting donor lung and right heart circulations)11. The circulation of Maruyama and colleagues12 involves an anastomosis between donor left PA and recipient Ao, which permits LV filling via the pulmonary circulation as a conduit, but effectively excludes the right heart.
In cases where near physiologic loading conditions were met, we advance the technique of HAHLT in 2 major respects. First, to our knowledge, the exact configuration we report has not been described in rats. It is possibly the most versatile circulation for investigators wishing to study the physiology, structure, and immunology of the transplanted heart-lung graft. Second, we describe how the function of the transplant graft can be directly characterized in vivo. For this application, pressure-volume conductance catheters can be introduced directly into the LV apex of the transplant graft, which allows for complete cardiac functional characterization. 
The technique described here can be applied to both acute and chronic studies of transplant graft function, while the functional assessment may be performed either in vivo or in vitro. We present a model in which the loading conditions can be near physiologic, however the degree of ventricular loading may be manipulated both acutely and chronically by diverting venous return towards or away from the graft. Afterload conditions can also be manipulated. Because the lung and its airway are retained in this transplant configuration, investigators can re-ventilate the donor lung acutely. Uniquely, lung re-ventilation changes the composition of blood perfusing the transplant coronary arteries. Under non-ventilated conditions, blood ejected from the donor aorta is deoxygenated, and mixes with oxygenated blood in the recipient aorta. Under acutely ventilated conditions, ejected blood becomes oxygenated. Thus, transplant graft function can be compared under ventilated and non-ventilated conditions, and also under variably loaded conditions.
The protocol below describes important modifications to previously described HAHLT donor and recipient operations. It also describes an optimal technique for protecting the transplant graft throughout the period of ischemia (time between donor explant and recipient implant). Advantages of this technique include physiologic conditions potentially approaching that of an orthotopically transplanted graft, and a wide range of investigative applications. An important limitation is its technical complexity. With adequate mentoring and practice, the advantages of this technique will likely outweigh the challenges in adopting it.

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>Material/Equipment</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>Celsior Cardioplegic Soution</td>
			<td>Genzyme</td>
			<td></td>
			<td>The solution is kept on ice throughout the procedure. We prepare our own solution, with slight modifications.</td>
		</tr>
		<tr>
			<td>Rodent Ventilator</td>
			<td>Harvard Apparatus</td>
			<td>Model 683</td>
			<td></td>
		</tr>
		<tr>
			<td>Vital Sign Monitor</td>
			<td>Nonin</td>
			<td>Model 9847V</td>
			<td>Displays SpO2 and heart rate.</td>
		</tr>
		<tr>
			<td>IV Cannulae</td>
			<td>Jelco</td>
			<td>3063</td>
			<td>24-26G x 3/4&#34; cannulae.</td>
		</tr>
		<tr>
			<td>IV Tubing</td>
			<td>CareFusion</td>
			<td>MP9259-C</td>
			<td>Short-length connector tubing (18cm).</td>
		</tr>
		<tr>
			<td>Surgical Clips</td>
			<td>Teleflex Medical</td>
			<td>001204</td>
			<td>Horizon titanium ligating clips.</td>
		</tr>
		<tr>
			<td>Sutures</td>
			<td>Ethicon, Sharpoint</td>
			<td>LA54G, AK-0107</td>
			<td>3-0 silk reel, and 9-0 prolene suture (single-armed, DR5 needle).</td>
		</tr>
		<tr>
			<td>Surgical Instruments</td>
			<td>Not Applicable</td>
			<td>Not Applicable</td>
			<td>The instruments used are generic, and can be purchased from any surgical supply company.</td>
		</tr>
	</tbody>
</table>

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.

上記のHAHLT技術は、高度な技術であり、詳細には細心の注意が必要です。 表1のハイライト失敗手順対成功に関連付けられており、技術的な問題のトラブルシューティングを行うためのガイドとして使用することができる重要な要因のいくつか。 受信者の大動脈がアンクランプされた後、グラフト冠状動脈は酸素を豊富に含んだ血液でいっぱいに見られるべ?...

Watch this Scientific Journal Video about Rat Heterotopic Abdominal Heart/Single-lung Transplantation in a Volume-loaded Configuration at JoVE.com

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

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.

ラット異腹部ボリュームにロードされた構成でのハート/シングル肺移植

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

rat-heterotopic-abdominal-heartsingle-lung-transplantation-volume

Research

JoVE Journal

Medicine

8.3K ビュー.  UBC James Hogg Research Center, St. Paul's Hospital. 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.

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

Orthotopic Liver Transplantation in Rats

Clinical progress in the field of liver transplantation has been largely supported by animal models1,2. Since the publication of the first orthotopic rat liver transplantation in 1979 by Kamada et al.3, this model has remained the gold standard despite various proposed alternative techniques4. Nevertheless, its broader use is limited by its steep learning curve5.
In this video paper, we show a simple and easy-to-establish revision of Kamada's two-cuff technique. The suprahepatic vena cava anastomosis is performed manually with a running suture, and the vena porta and infrahepatic vena cava anastomoses are performed utilizing a quick-linker cuff system6. Manufacturing the quick-linker kit is shown in a separate video paper.

We present an easy-to-establish revision of the classical two-cuff technique for orthotopic liver transplantation in rat.

ラット同所性肝移植

Clinical progress in the field of liver transplantation has been largely supported by animal models1,2. Since the publication of the first orthotopic rat ...

医学

Heterotopic Auxiliary Rat Liver Transplantation With Flow-regulated Portal Vein Arterialization in Acute Hepatic Failure

In acute hepatic failure auxiliary liver transplantation is an interesting alternative approach. The aim is to provide a temporary support until the failing native liver has regenerated.1-3 The APOLT-method, the orthotopic implantation of auxiliary segments- averts most of the technical problems. However this method necessitates extensive resections of both the native liver and the graft.4 In 1998, Erhard developed the heterotopic auxiliary liver transplantation (HALT) utilizing portal vein arterialization (PVA) (Figure 1). This technique showed promising initial clinical results.5-6 We developed a HALT-technique with flow-regulated PVA in the rat to examine the influence of flow-regulated PVA on graft morphology and function (Figure 2).
A liver graft reduced to 30 % of its original size, was heterotopically implanted in the right renal region of the recipient after explantation of the right kidney.&#160; The infra-hepatic caval vein of the graft was anastomosed with the infrahepatic caval vein of the recipient. The arterialization of the donor&#8217;s portal vein was carried out via the recipient&#8217;s right renal artery with the stent technique. The blood-flow regulation of the arterialized portal vein was achieved with the use of a stent with an internal diameter of 0.3 mm. The celiac trunk of the graft was end-to-side anastomosed with the recipient&#8217;s aorta and the bile duct was implanted into the duodenum. A subtotal resection of the native liver was performed to induce acute hepatic failure. 7
In this manner 112 transplantations were performed. The perioperative survival rate was 90% and the 6-week survival rate was 80%. Six weeks after operation, the native liver regenerated, showing an increase in weight from 2.3&#177;0.8 g to 9.8&#177;1 g. At this time, the graft&#8217;s weight decreased from 3.3&#177;0.8 g to 2.3&#177;0.8 g.
We were able to obtain promising long-term results in terms of graft morphology and function. HALT with flow-regulated PVA reliably bridges acute hepatic failure until the native liver regenerates.

Auxiliary liver transplantation provides a temporary support in acute hepatic failure, until regeneration of the failing liver. The heterotopic auxiliary liver transplantation (HALT) with portal vein arterialization (PVA) renders sufficient liver function. We developed an analogous technique in the rat, to examine the influence of the portal vein arterialization on the morphology and function of the graft.

急性肝障害における流れ調節性門脈動脈血と異補助ラット肝移植

In acute hepatic failure auxiliary liver transplantation is an interesting alternative approach. The aim is to provide a temporary support until the ...

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.

ネズミ異心臓移植技術

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.

異心臓移植のマウスモデルを用いた肺動脈弁の移植

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

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.

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.

マウスにおける異子宮頸心臓移植

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

Blood Circuit Reconstruction in an Abdominal Mouse Heart Transplantation Model

The surgical technique of heterotopic abdominal heart transplantation in mice is a standard model for research in transplantation immunology. Here, the established technique for a modified blood circuit reconstruction in a heterotopic abdominal heart transplantation model is presented. This method uses the intrathoracic inferior vena cava (IIVC) instead of the pulmonary artery of the donor heart for the anastomosis to the inferior vena cava of the recipient. It is facilitating and improving success rates for abdominal heart transplantation in mice.

A novel technique for blood circuit reconstruction in a heterotopic abdominal mouse heart transplantation model is demonstrated.

腹部マウス心臓移植モデルにおける血液回路再建

The surgical technique of heterotopic abdominal heart transplantation in mice is a standard model for research in transplantation immunology. Here, the ...

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.

げっ歯類におけるヘテロトピック心臓弁移植のための簡略モデル

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.

心臓同種移植への治療薬の送達のためのブタヘテロトピック心臓移植プロトコル

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.

マウス頸部異所性心臓移植モデルのための修正カフ法

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

A Heterotopic Mouse Model for Studying Laryngeal Transplantation

Laryngeal heterotopic transplantation, although a technically challenging procedure, offers more scientific analysis and cost benefits compared to other animal models. Although first described by Shipchandler et al. in 2009, this technique is not widely used, possibly due to the difficulties in learning the microsurgical technique and time required to master it. This paper describes the surgical steps in detail, as well as potential pitfalls to avoid, in order to encourage effective use of this technique.
In this model, the bilateral carotid arteries of the donor larynx are anastomosed to the recipient carotid artery and external jugular vein, allowing for blood flow through the graft. Blood flow can be confirmed intraoperatively by the visualization of blood filling in the graft bilateral carotid arteries, reddening of the thyroid glands of the graft, and bleeding from micro vessels in the graft. The crucial elements for success include delicate preservation of the graft vessels, making the correct size arteriotomy and venotomy, and using the appropriate number of sutures on the arterial-arterial and arterial-venous anastomoses to secure vessels without leakage and prevent occlusion. 
Anyone can become proficient in this model with sufficient training and perform the procedure in approximately 3 h. If performed successfully, this model allows for immunologic studies to be performed with ease and at low cost.

The aim of this manuscript is to describe the microsurgical steps required to perform a heterotopic laryngeal transplant in mice. The advantages of this mouse model compared to other animal models of laryngeal transplantation are its cost-effectiveness and the availability of immunologic assays and data.

喉頭移植研究のための異所性マウスモデル

Laryngeal heterotopic transplantation, although a technically challenging procedure, offers more scientific analysis and cost benefits compared to other ...