Name: a novel murine model of arteriovenous fistula failure: the surgical procedure in detail
Uploaded: 2016-02-03T13:00:00.000+00:00
Duration: 10 min 10 s
Description: Watch this Scientific Journal Video about A Novel Murine Model of Arteriovenous Fistula Failure: The Surgical Procedure in Detail at JoVE.com

This study was supported by a grant from the Dutch Kidney Foundation (KJPB 08.0003). 
Carolien Rothuizen is acknowledged for her contribution to the study. Hoang Pham is acknowledged for his assistance with the pathology work-up.

すべての実験は、ライデン大学医療センターの動物福祉に関する委員会によって承認されました。 1.動物の準備と麻酔<ol><li>三から四パーセントイソフルランで満たされた連続麻酔導入チャンバー内のマウス（1-3ヶ月齢）を麻酔。 </li><li>首と電気かみそりを使用して、左上の脚の内側部分の腹側を剃ると毛を除去するために、テープ片を使用します。 </li><li>両眼に眼軟膏を適用します。 </li><li>鼻換気マスクが固定されている加熱毛布の上に動物を配置します。粘着テープを使用して、鼻マスクにヘッドを固定。 <ol><li>以下の流量で酸素富化空気のセットを使用して1.5-2.0％の濃度でイソフルランを送達：空気0.3リットル/分、100％の酸素0.2リットル/分。麻酔の深さを評価し、必要に応じてイソフルランの濃度を調整するために足の指の間の皮膚をつまみます。 </li></ol></li><li>に動物を固定仰臥位℃で約37に設定されているすべての手足に粘着テープで加熱ブランケット。 </li><li> 0.1ミリグラムの用量で塩化ナトリウム（NaCl）で0.9％に溶解したブプレノルフィンを注入/動物の脇腹に無菌の0.9％NaClを皮下の0.5ミリリットルと一緒kgです。 </li><li>準備されたエリアにクロルヘキシジンチンキに0.5％を適用します。 </li></ol> 2.皮膚切開<ol><li>外科医に向かって頭を顕微鏡下で動物を置きます。 </li><li>マイクロはさみを使用して、腹側頸部の正中線に約1.5センチ縦切開を行います。 </li><li>鉗子で唾液腺をつかみ、それが部分的に傷の外になるまで頭側右の唾液腺を移動させます。 </li></ol> 3.静脈を解剖し、準備<ol><li>識別し、露骨に拡散することにより、その血管周囲組織から完全に右外頸静脈の背内側枝を分析だけ長辺と血管の方向です。 </li><li>解剖静脈の周りにループ（10.0縫合糸）を配置し、それをロックすることなく、1つの結び目を適用します。 </li></ol> 4.胸鎖乳突筋の削除<ol><li>鉗子を使用して、ぶっきらぼうにその境界線に沿って拡散することによりその周囲から右胸鎖乳突筋を分析。孤立した筋肉の下でオープンピンセットを置き、それが6.0縫合糸で近位および遠位連結。その後、焼灼器を使用して筋肉を切除。 </li></ol> 5.共通の頸動脈を解剖し、準備<ol><li>鉗子を使用して、右総頸動脈を特定し、解剖。動脈の操作を補助するために、動脈の周りに6.0縫合糸を配置します。 </li><li>遠位および近位に可能な限り血管クランプを適用します。 </li><li>特殊なマイクロシザーを用いて約1ミリメートルの動脈の真ん中に縦切開を行います。 </li><li>リンスヘパリン溶液[100 IU / mlの]と動脈血管が血液の透明になるまで。 </li><li>必要に応じて幅を血管クランプ[1.1ミリメートル]を一致させるために切開の長さを測定し、調整します。 </li></ol>静脈の6.ライゲーション<ol><li>近位にすでに準備静脈の血管クランプを適用し、静脈の周りに縫合糸を配置します。 </li><li>以前に配置した10.0縫合糸を使用して、できるだけ遠位静脈を縫合糸上に配置され、ライゲートされた止血を使用して静脈に優しい尾トラクションを適用します。 </li><li>ライゲーションにちょうど近位にはさみを使用して静脈をカットします。 </li><li>血管鉗子を使用して、静かに静脈の内腔を開き、ヘパリン溶液[100 IU / mlの]でそれをすすいでください。 </li></ol>吻合パート1の7の作成<ol><li> 6 O &#39;で縫合糸に続く12時の位置に2平方ノットを使用して結節縫合（10.0）と動脈に静脈を接続しますクロック位置。それは自身の軸の周りに回転させることなく動脈に静脈を接続してください。外科的露出を改善するために動物を回転させます。 </li><li>静脈の周りに縫合糸を配置し、止血を使用して穏やかな横方向のトラクションを適用します。 </li><li>同じ結節縫合を使用して、吻合の可視腹側で約3-4追加の縫合糸を配置することによって、吻合を完了します。 </li></ol> 8.ヘパリン投与<ol><li>加熱ブランケット180度回転し、左上肢に顕微鏡の焦点を合わせます。 </li><li>上肢の内側部に縦方向に実行され、皮膚を通して見ることができる血管構造を探して、大腿静脈/動脈/神経束を特定します。直接、長手方法で大腿静脈上記の約1cmのマイクロハサミで切開を行います。 </li><li>慎重に大腿静脈の血管周囲の組織を解剖し、線量Oでヘパリンを注入大腿静脈で0.2 IU /グラムの体重F。 </li></ol>吻合パート2の9創出<ol><li>首のエリアに戻り、静脈の周囲に配置された縫合糸を除去します。 </li><li>その後、静脈の上にして、再度頸動脈の下、頸動脈の下を通る縫合糸（6.0）を配置します。 （ 図1L） </li><li>静脈血管クランプを取り外します。 </li><li>次に、同時にねじることによって時計回りに頸動脈の軸に沿って180°動静脈瘻を完了半分の両方の血管クランプをねじると、縫合糸の端部に内側トラクションを適用します。 </li><li>ステップ7.2で説明されているのと同じやり方で吻合を完了します。 </li></ol> 10.血管クランプの取り外し<ol><li>縫合糸を外し、血管が180°反時計回りにクランプ回転させます。 </li><li>近位血管クランプ続い遠位血管クランプを取り外します。 </li> &lt;李&gt;は優しく、血管鉗子で静脈流出路を閉塞することにより、吻合の開存性を評価します。特許吻合の場合は、事前に閉塞静脈セクションでは、脈動的に拡大していきます。鉗子を使用して、元の解剖学的位置に右の唾液腺を再配置します。 </li></ol> 11.皮膚閉鎖と術後ケア<ol><li>中断のない縫合糸（6.0）と上肢の皮膚を閉じます。 </li><li>中断のない縫合糸（6.0）と首の皮膚を閉じます。 </li><li>加熱ブランケットから動物を削除し、0.5ミリリットルのNaClを0.9％を皮下に注入します。 </li><li>加熱ランプで加熱して暗くケージに動物を置き、それが完全に回復しましょう​​。動物が完全に回復しない場合には、それが原因で手術領域の出血に血行動態ショックを受けないことを確認してください。このような腫れ首や血液の漏れなどの兆候を探してください。 </li><li> buprの最初の注射後約6時間後enorphine、さらに72時間、十分な鎮痛を提供するために、製造業者によって推奨される用量におけるブプレノルフィン皮下の徐放性製剤の単回投与を注入。 </li></ol> 12.組織の収穫<ol><li>メデトミジン（0.5ミリグラム/キログラム）とフェンタニル（0.​​05ミリグラム/ kg）を腹腔内投与し、麻酔薬の混合物を含有ミダゾラム（5mg / kg）で動物を麻酔。 </li><li>非加熱シリコンマットにその足や頬を通して針を挿入することにより、仰臥位で動物を固定。 </li><li>マイクロはさみで首に傷跡の上に約1.5〜2.0センチメートルの切開を行います。 </li><li> AVFを分析し、簡単に識別proximal-、遠位動脈と静脈流出路の周りに縫合糸（6.0）を配置。 </li><li>プロトコルのセクション10.3に記載されているようにAVFの開通性を評価します。 </li><li>内側開腹のuを実行することにより、腹膜腔を開きますマイクロシザーを歌います。 <ol><li>鎖骨中央線に尾側開始ハサミを使用して、左右の腹側胸郭を切断。マイクロはさみを使用して、腹側胸郭の中央部に取り付けられ、その後、ピンセットを用いて頭蓋このセクションを変位されるダイヤフラムを横断。 </li><li>下大静脈を見つけ、マイクロはさみでそれを横断します。 </li></ol></li><li>左心室に針23ゲージ針を挿入し、血管内の流体がほぼ透明になるまで約100 mmHgでの圧力で、PBSで灌流。 </li><li>針を除去することなく、今は約100 mmHgの圧力で10分間、4％ホルマリンで灌流。 </li><li>マイクロハサミで動脈と静脈の両方を横断することにより、AVFを切除し、4％ホルマリン溶液O / Nでそれを沈めます。 </li></ol> 13.組織の埋め込みとセクショニング<ol><li>製造者のprotocoに従ってパラフィンために組織を処理リットル。適用されるプロトコルは、以下のとおりであります。 <ol><li> RTで1時間、エタノール70％に浸します。 2回繰り返します。 </li><li> RTで1時間、エタノール96％に浸します。 2回繰り返します。 </li><li> RTで1時間エタノール99.5％に浸します。一回繰り返します。 </li><li> RTで1.5時間、エタノール99.5％に浸します。 </li><li> RTで1時間キシレン中に100％を浸します。一回繰り返します。 </li><li> 62℃で1時間、パラフィンに浸し。 2回繰り返します。 </li></ol></li><li>静脈流出路は、包埋カセットに垂直に配向されるように、パラフィンにAVFを埋め込みます。 </li><li>ミクロトームを用いて、それぞれが5ミクロンの厚さで30のセクションからなる12連続切片を作成します。そして、各シリアルセクションの、吻合部に​​近い領域で始まる整然と12までの位置1から始まる単一のマウントガラス上の1つのセクションを配置します。 </li></ol>

<ol>
	<li>Rotmans, J. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Animal+Models+for+Studying+Pathophysiology+of+Hemodialysis+Access.">Animal Models for Studying Pathophysiology of Hemodialysis Access.</a> The Open Urology &amp; Nephrology Journal. 7, 14-21 (2014).</li><li>Kwei, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Early+adaptive+responses+of+the+vascular+wall+during+venous+arterialization+in+mice.">Early adaptive responses of the vascular wall during venous arterialization in mice.</a> Am.J.Pathol. 164 (1), 81-89 (2004).</li><li>Castier, 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=Characterization+of+neointima+lesions+associated+with+arteriovenous+fistulas+in+a+mouse+model.">Characterization of neointima lesions associated with arteriovenous fistulas in a mouse model.</a> Kidney Int. 70 (2), 315-320 (2006).</li><li>Krishnamoorthy, M. K., 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=Hemodynamic+wall+shear+stress+profiles+influence+the+magnitude+and+pattern+of+stenosis+in+a+pig+AV+fistula.">Hemodynamic wall shear stress profiles influence the magnitude and pattern of stenosis in a pig AV fistula.</a> Kidney Int. 74 (11), 1410-1419 (2008).</li><li>Ene-Iordache, B., Cattaneo, L., Dubini, G., Remuzzi, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+anastomosis+angle+on+the+localization+of+disturbed+flow+in+'side-to-end'+fistulae+for+haemodialysis+access.">Effect of anastomosis angle on the localization of disturbed flow in 'side-to-end' fistulae for haemodialysis access.</a> Nephrol. Dial. Transplant. 28 (4), 997-1005 (2013).</li><li>Wong, C. 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=Vascular+remodeling+and+intimal+hyperplasia+in+a+novel+murine+model+of+arteriovenous+fistula+failure.">Vascular remodeling and intimal hyperplasia in a novel murine model of arteriovenous fistula failure.</a> J.Vasc.Surg. 59 (1), 192-201 (2014).</li><li>Rekhter, M., Nicholls, S., Ferguson, M., Gordon, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cell+proliferation+in+human+arteriovenous+fistulas+used+for+hemodialysis.">Cell proliferation in human arteriovenous fistulas used for hemodialysis.</a> Arterioscler. Thromb. 13 (4), 609-617 (1993).</li><li>Falk, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Maintenance+and+salvage+of+arteriovenous+fistulas.">Maintenance and salvage of arteriovenous fistulas.</a> J. Vasc. Interv. Radiol. 17 (5), 807-813 (2006).</li><li>Tordoir, J. 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=Prospective+evaluation+of+failure+modes+in+autogenous+radiocephalic+wrist+access+for+haemodialysis.">Prospective evaluation of failure modes in autogenous radiocephalic wrist access for haemodialysis.</a> Nephrol. Dial. Transplant. 18 (2), 378-383 (2003).</li><li>Dixon, B. S., Novak, L., Fangman, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hemodialysis+vascular+access+survival:+upper-arm+native+arteriovenous+fistula.">Hemodialysis vascular access survival: upper-arm native arteriovenous fistula.</a> Am. J. Kidney Dis. 39 (1), 92-101 (2002).</li><li>Yang, B., Shergill, U., Fu, A. A., Knudsen, B., Misra, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+mouse+arteriovenous+fistula+model.">The mouse arteriovenous fistula model.</a> J. Vasc. Interv. Radiol. 20 (7), 946-950 (2009).</li><li>Kang, 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=Regional+and+systemic+hemodynamic+responses+following+the+creation+of+a+murine+arteriovenous+fistula.">Regional and systemic hemodynamic responses following the creation of a murine arteriovenous fistula.</a> Am. J. Physiol Renal Physiol. 301 (4), F845-F851 (2011).</li><li>Wong, C. 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=Elastin+is+a+key+regulator+of+outward+remodeling+in+arteriovenous+fistulas.">Elastin is a key regulator of outward remodeling in arteriovenous fistulas.</a> Eur. J. Vasc. Endovasc. Surg. 49 (4), 480-486 (2015).</li><li>Kennedy, 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=Does+renal+failure+cause+an+atherosclerotic+milieu+in+patients+with+end-stage+renal+disease.">Does renal failure cause an atherosclerotic milieu in patients with end-stage renal disease.</a> Am. J. Med. 110 (3), 198-204 (2001).</li><li>Cheung, A. K., 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=Atherosclerotic+cardiovascular+disease+risks+in+chronic+hemodialysis+patients.">Atherosclerotic cardiovascular disease risks in chronic hemodialysis patients.</a> Kidney Int. 58 (1), 353-362 (2000).</li><li>Lee, T., 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=Severe+venous+neointimal+hyperplasia+prior+to+dialysis+access+surgery.">Severe venous neointimal hyperplasia prior to dialysis access surgery.</a> Nephrol. Dial. Transplant. 26 (7), 2264-2270 (2011).</li><li>Kokubo, T., 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=CKD+accelerates+development+of+neointimal+hyperplasia+in+arteriovenous+fistulas.">CKD accelerates development of neointimal hyperplasia in arteriovenous fistulas.</a> J. Am. Soc. Nephrol. 20 (6), 1236-1245 (2009).</li></ol>

著者らは、開示することは何もありません。

AVFは、血液透析治療でアキレス腱であると考えられます。残念ながら、AVFはまだ失敗8-10の高い数に苦しんでいます。根底にあるメカニズムに関する広範な研究にもかかわらず、正確な病態生理は不明のまま。 AVFの故障のための多数のマウスモデルは、既に文献2,3,11,12に記載されています。しかし、これらのモデルはいずれも、ほとんどの臨床状況で使用されている動脈側吻...

機能的な血管アクセス導管は生き続けるために慢性血液透析に依存腎不全患者のために極めて重要です。動静脈瘻（AVF）の建設は、現在、血管アクセスのための好ましい選択肢です。しかし、AVF関連した合併症は、慢性血液透析を受けている患者のための罹患率の主要な原因を構成しています。広範な科学的な努力にもかかわらず、小説のどれもAVFの耐久性の大幅な改善をもたらすでしたAVFへのアクセスに関連した合併症を減らすために近づいていません。この残念​​な進歩の一部は、血液透析アクセス障害の根底にある病態生理の不完全な理解に関係します。 AVアクセス障害の病態生理を解明するために、厳密にヒト病理を模倣する動物モデルは、最も重要です。この点で、動物種だけでなく、吻合部位、必要な抗凝固療法とサージ後のフォローアップの期間ryは、アカウント1に入れなければなりません。大型動物は、新たな治療戦略を開発することを目的とした介入研究のために最も適しているが、マウスモデルは、トランスジェニックマウスの在庫状況によりAV・アクセス障害の根底にある分子メカニズムでより多くの洞察を得るための大きな可能性を持っています。また、マウスの大多数は、より大きな動物での使用に比べて低コストで、この目的のために使用することができます。 AVF障害の最初のマウスモデルはKweiとらによって 2004年に記載された。このモデルでは2、のAVFは、血管内カテーテルを使用して、エンド・ツー・エンドの方法で頸動脈と頸静脈を用いて構築しました。このモデルは、エンドツーエンドの構成及び血管内カテーテルの存在が人間のAVFのためのこのモデルの有効性を制限するが、のAVFの早期静脈適応を研究するのに有用であり得ます。改良されたAVFモデルはCastierやらによって導入された。3最後のました頸動脈は、頸静脈の側に接続されています。しかしながら、血液透析患者でのAVFは通常、動脈の側に静脈の端部をanatomizingによって構成されています。それは導管4内の血行力学的プロファイルを決定するため、AVFの正確な構成は、AVアクセスモデルの重要な特徴です。後者は、機能不全および内膜肥厚（IH）5のその後の発展を内皮細胞への重要な貢献者です。 小説のマウスモデルは、最近、ヒト6で利用されているものと同一の解剖学的構成で開発されました。このモデルでは、AVFは断続縫合で総頸動脈の側に外頸静脈の枝の先端を吻合することにより、C57BL / 6マウスに作成されます。本論文では、複雑な病態を解明することを目的とし、このマウスモデルの普及を促進するために、このモデルの顕微手順に焦点を当てます血液透析アクセス失敗の。

<table><tbody><tr><td>Dissecting microsocpe</td><td>Leica</td><td>M80</td><td></td></tr><tr><td>Forceps</td><td>Medicon</td><td>07.61.25</td><td></td></tr><tr><td>Vascular forceps</td><td>S&amp;amp;T</td><td>JFL-3D.2</td><td></td></tr><tr><td>Vascular forceps</td><td>S&amp;amp;T</td><td>D-5a.2</td><td></td></tr><tr><td>Forceps</td><td>Roboz</td><td>SS/45</td><td></td></tr><tr><td>Micro scissor 5 mm blade</td><td>Fine science tools</td><td>15000-08</td><td></td></tr><tr><td>Micro scissor 2 mm blade</td><td>Fine science tools</td><td>15000-03</td><td></td></tr><tr><td>Scissor</td><td>Medicon</td><td>05.12.21</td><td></td></tr><tr><td>Clip applier 1</td><td>S&amp;amp;T</td><td>CAF-4</td><td></td></tr><tr><td>Vascular clamp 1</td><td>S&amp;amp;T</td><td>B-1V</td><td></td></tr><tr><td>Clip applier 2</td><td>BBraun</td><td>FE572K</td><td></td></tr><tr><td>Vascular clamp 2</td><td>BBraun</td><td>FE740K</td><td></td></tr><tr><td>Hemostatic forceps</td><td>BBraun</td><td>BH110</td><td></td></tr><tr><td>10.0 sutures</td><td>BBraun</td><td>G1117041</td><td></td></tr><tr><td>6.0 sutures</td><td>BBraun</td><td>768464</td><td></td></tr><tr><td>Cauterizer</td><td>Fine science tools</td><td>18010-00</td><td></td></tr><tr><td>Needle holder</td><td>Medicon</td><td>11.82.18</td><td></td></tr><tr><td>Ocular ointment</td><td>Pharmachemie</td><td>41821101</td><td></td></tr><tr><td>Chlorhexidine tincture 0,5%</td><td>Leiden University Medical Center</td><td>NA</td><td></td></tr><tr><td>Heparin</td><td>Leo Pharma</td><td>012866-08</td><td></td></tr><tr><td>Buprenorphin</td><td>RB Pharmaceuticals&amp;nbsp;</td><td>283732</td><td></td></tr><tr><td>Isoflurane</td><td>Pharmachemie</td><td>45,112,110</td><td></td></tr><tr><td>Anesthesia mask</td><td>Maastricht university</td><td>custom made</td><td></td></tr><tr><td>Midazolam</td><td>Actavis</td><td>AAAC6877</td><td></td></tr><tr><td>Dexmedetomidine</td><td>Orion</td><td>141-267</td><td></td></tr><tr><td>Fentanyl</td><td>Bipharma</td><td>15923002</td><td></td></tr><tr><td>Continuous anaesthetic induction chamber</td><td>Vet-tech solutions</td><td>AN010R</td><td></td></tr></tbody></table>

a novel murine model of arteriovenous fistula failure: the surgical procedure in detail

The arteriovenous fistula (AVF) still suffers from a high number of failures caused by insufficient remodeling and intimal hyperplasia from which the exact pathophysiology remains unknown. In order to unravel the pathophysiology a murine model of AVF-failure was developed in which the configuration of the anastomosis resembles the preferred situation in the clinical setting. A model was described in which an AVF is created by connecting the venous end of the branch of the external jugular vein to the side of the common carotid artery using interrupted sutures. At a histological level, we observed progressive stenotic intimal lesions in the venous outflow tract that is also seen in failed human AVFs. Although this procedure can be technically challenging due to the small dimensions of the animal, we were able to achieve a surgical success rate of 97% after sufficient training. The key advantage of a murine model is the availability of transgenic animals. In view of the different proposed mechanisms that are responsible for AVF failure, disabling genes that might play a role in vascular remodeling can help us to unravel the complex pathophysiology of AVF failure.

吻合（ 図1）を作成した後、開存はまもなく血管鉗子で静脈流出路を閉塞することによって評価されるべきです。吻合は、特許である場合には、閉塞に対する血管管の近位は明らかに拍動の方法で展開する必要があります。さらに、開通性を効果的に血管造影（ 図2）として機能する近赤外蛍光透視法（NIRF）を使用して確認されます。 （図2）に示されて...

Watch this Scientific Journal Video about A Novel Murine Model of Arteriovenous Fistula Failure: The Surgical Procedure in Detail at JoVE.com

A Novel Murine Model of Arteriovenous Fistula Failure: The Surgical Procedure in Detail

Here we present a murine model of arteriovenous fistula (AVF) failure in which a clinically relevant anastomotic configuration is incorporated. This model can be used to study the pathophysiology and to test possible therapeutic interventions.

動静脈瘻の障害の新規マウスモデル：詳細の外科的手順

The arteriovenous fistula (AVF) still suffers from a high number of failures caused by insufficient remodeling and intimal hyperplasia from which the ...

a-novel-murine-model-arteriovenous-fistula-failure-surgical-procedure

Research

JoVE Journal

Medicine

13.0K ビュー.  Leiden University Medical Center. Here we present a murine model of arteriovenous fistula (AVF) failure in which a clinically relevant anastomotic configuration is incorporated. This model can be used to study the pathophysiology and to test possible therapeutic interventions.

ビデオ: A Novel Murine Model of Arteriovenous Fistula Failure: The Surgical Procedure in Detail

Microsurgical Venous Pouch Arterial-Bifurcation Aneurysms in the Rabbit Model: Technical Aspects

For ruptured human cerebral aneurysms endovascular embolization has become an equivalent alternative to aneurysm clipping.1 However, large clinical trials have shown disappointing long-term results with unacceptable high rates of aneurysm recanalization and delayed aneurysm rupture.2 To overcome these problems, animal experimental studies are crucial for the development of better endovascular devices.3-5
Several animal models in rats, rabbits, canines and swine are available.6-8 Comparisons of the different animal models showed the superiority of the rabbit model with regard to hemodynamics and comparability of the coagulation system and cost-effectiveness.9-11 
The venous pouch arterial bifurcation model in rabbits is formed by a venous pouch sutured into an artificially created true bifurcation of both common carotid arteries (CCA). The main advantage of this model are true bifurcational hemodynamics.12 The major drawbacks are the sofar high microsurgical technical demands and high morbidity and mortality rates of up to 50%.13 These limitations have resulted in less frequent use of this aneurysm model in the recent years. These shortcomings could be overcome with improved surgical procedures and modified peri- and postoperative analgetic management and anticoagulation.14-16 Our techniques reported in this paper demonstrate this optimized technique for microsurgical creation of arterial bifurcation aneurysms.

An optimized technique for the microsurgical creation of arterial bifurcation aneurysms mimicking bifurcation human cerebral aneurysms is described. A venous pouch is sutured into an artificially created true bifurcation of both common carotid arteries. Facilitated microsurgical techniques and aggressive postoperative anticoagulation and analgesia lead to minimized morbidity rates and high aneurysm patency rates.

ウサギモデルにおける顕微静脈ポーチ動脈 - 分岐瘤：技術的側面

For ruptured human cerebral aneurysms endovascular embolization has become an equivalent alternative to aneurysm clipping.1 However, large clinical trials ...

医学

Technical Aspects of the Mouse Aortocaval Fistula

Technical aspects of creating an arteriovenous fistula in the mouse are discussed. Under general anesthesia, an abdominal incision is made, and the aorta and inferior vena cava (IVC) are exposed. The proximal infrarenal aorta and the distal aorta are dissected for clamp placement and needle puncture, respectively. Special attention is paid to avoid dissection between the aorta and the IVC. After clamping the aorta, a 25 G needle is used to puncture both walls of the aorta into the IVC. The surrounding connective tissue is used for hemostatic compression. Successful creation of the AVF will show pulsatile arterial blood flow in the IVC. Further confirmation of successful AVF can be achieved by post-operative Doppler ultrasound.

The goal is to produce an arteriovenous fistula that is simple and reproducible. This method does not use sutures or glue adhesive. Therefore the samples can be used with the least amount of foreign materials for analysis.

マウスAortocaval瘻の技術的な側面

Technical aspects of creating an arteriovenous fistula in the mouse are discussed. Under general anesthesia, an abdominal incision is made, and the aorta ...

Adult Mouse Venous Hypertension Model: Common Carotid Artery to External Jugular Vein Anastomosis.

The understanding of the pathophysiology of brain arteriovenous malformations and arteriovenous fistulas has improved thanks to animal models. A rat model creating an artificial fistula between the common carotid artery (CCA) and the external jugular vein (EJV) has been widely described and proved technically feasible. This construct provokes a consistent cerebral venous hypertension (CVH), and therefore has helped studying the contribution of venous hypertension to formation, clinical symptoms, and prognosis of brain AVMs and dural AVFs. Equivalent mice models have been only scarcely described and have shown trouble with stenosis of the fistula. An established murine model would allow the study of not only pathophysiology but also potential genetic therapies for these cerebrovascular diseases.
We present a model of arteriovenous fistula that produces a durable intracranial venous hypertension in the mouse. Microsurgical anastomosis of the murine CCA and EJV can be difficult due to diminutive anatomy and frequently result in a non-patent fistula. In this step-by-step protocol we address all the important challenges encountered during this procedure. Avoiding excessive retraction of the vein during the exposure, using 11-0 sutures instead of 10-0, and making a carefully planned end-to-side anastomosis are some of the critical steps. Although this method requires advanced microsurgical skills and a longer learning curve that the equivalent in the rat, it can be consistently developed.
This novel model has been designed to integrate transgenic mouse techniques with a previously well-established experimental system that has proved useful to study brain AVMs and dural AVFs. By opening the possibility of using transgenic mice, a broader spectrum of valid models can be achieved and genetic treatments can also be tested. The experimental construct could also be further adapted to the study of other cerebrovascular diseases related with venous hypertension such as migraine, transient global amnesia, transient monocular blindness, etc.

We describe a method for creating a reliable model of cerebral venous hypertension in the adult mouse. This model has been widely described and tested in the rat. This new counterpart in the mice opens the possibility of using genetic modified animals and thereby broadens the applications of the model.

成体マウス静脈性高血圧モデル：外頸静脈吻合部の総頸動脈。

The understanding of the pathophysiology of brain arteriovenous malformations and arteriovenous fistulas has improved thanks to animal models. A rat model ...

A New Murine Model of Endovascular Aortic Aneurysm Repair

Endovascular aneurysm exclusion is a validated technique to prevent aneurysm rupture. Long-term results highlight technique limitations and new aspects of Abdominal aortic aneurysm (AAA) pathophysiology. There is no abdominal aortic aneurysm endograft exclusion model cheap and reproducible, which would allow deep investigations of AAA before and after treatment. We hereby describe how to induce, and then to exclude with a covered coronary stentgraft an abdominal aortic aneurysm in a rat. The well known elastase induced AAA model was first reported in 19901 in a rat, then described in mice2. Elastin degradation leads to dilation of the aorta with inflammatory infiltration of the abdominal wall and intra luminal thrombus, matching with human AAA. Endovascular exclusion with small covered stentgraft is then performed, excluding any interactions between circulating blood and the aneurysm thrombus. Appropriate exclusion and stentgraft patency is confirmed before euthanasia by an angiography thought the left carotid artery. Partial control of elastase diffusion makes aneurysm shape different for each animal. It is difficult to create an aneurysm, which will allow an appropriate length of aorta below the aneurysm for an easy stentgraft introduction, and with adequate proximal and distal neck to prevent endoleaks. Lots of failure can result to stentgraft introduction which sometimes lead to aorta tear with pain and troubles to stitch it, and endothelial damage with post op aorta thrombosis. Giving aspirin to rats before stentgraft implantation decreases failure rate without major hemorrhage. Clamping time activates neutrophils, endothelium and platelets, and may interfere with biological analysis.

Histological and biochemical modifications after aneurysm endograft exclusion are still unclear. We describe a new model of endograft implantation on a murine aneurysm. Through thrombus analysis with and without persistant circulating blood stream interface, and new endovascular biomaterials evaluation, this model will help to better understand endovascular aneurysm exclusion pathobiology.

血管内大動脈瘤の修復の新しいマウスモデル

Endovascular aneurysm exclusion is a validated technique to prevent aneurysm rupture. Long-term results highlight technique limitations and new aspects of ...

Murine Model of Femoral Artery Wire Injury with Implantation of a Perivascular Drug Delivery Patch

Percutaneous interventions including balloon angioplasty and stenting have been used to restore blood flow in vessels with occlusive vascular disease. While these therapies lead to the rapid restoration of blood flow, these technologies remain limited by restenosis in the case of bare metal stents and angioplasty, or reduced healing and possibly enhanced risk of thrombosis in the case of drug eluting stents. A key pathophysiological mechanism in the formation of restenosis is intimal hyperplasia caused by the activation of vascular smooth muscle cells and inflammation due to arterial stretch and injury. Surgeries that induce arterial injury in genetically modified mice are useful for the mechanistic study of the vascular response to injury but are often technically challenging to perform in mouse models due to the their small size and lack of appropriate sized devices. We describe two approaches for a surgical technique that induces endothelial denudation and arterial stretch in the femoral artery of mice to produce robust neointimal hyperplasia. The first approach creates an arteriotomy in the muscular branch of the femoral artery to obtain vascular access. Following wire injury this arterial branch is ligated to close the arteriotomy. A second approach creates an arteriotomy in the main femoral artery that is later closed through localized cautery. This method allows for vascular access through a larger vessel and, consequently, provides a less technically demanding procedure that can be used in smaller mice. Following either method of arterial injury, a degradable drug delivery patch can be placed over or around the injured artery to deliver therapeutic agents.

We describe a surgical technique that produces wire injury in the femoral artery of mice to induce neointimal hyperplasia to serve as a model testing system for the perivascular delivery of therapeutic compounds for the inhibition of restenosis.

血管周囲ドラッグデリバリーパッチの移植に大腿動脈ワイヤ傷害のマウスモデル

Percutaneous interventions including balloon angioplasty and stenting have been used to restore blood flow in vessels with occlusive vascular disease. ...

Intraluminal Drug Delivery to the Mouse Arteriovenous Fistula Endothelium

Delivery of therapeutic agents to enhance arteriovenous fistula (AVF) maturation can be administered either via intraluminal or external routes. The simple murine AVF model was combined with intraluminal administration of drug solution to the venous endothelium at the same time as fistula creation. Technical aspects of this model are discussed. Under general anesthesia, an abdominal incision is made and the aorta and inferior vena cava (IVC) are exposed. The infra-renal aorta and IVC are dissected for clamping. After proximal and distal clamping, the puncture site is exposed and a 25 G needle is used to puncture both walls of the aorta and into the IVC. Immediately after the puncture, a reporter gene-expressing viral vector was infused in the IVC via the same needle, followed by 15 min of incubation. The intraluminal administration method enabled more robust viral gene delivery to the venous endothelium compared to administration by the external route. This novel method of delivery will facilitate studies that explore the role of the endothelium in AVF maturation and enable intraluminal drug delivery at the time of surgical operation.

After puncturing the aorta through the inferior vena cava (IVC) to create an aorto-caval fistula in the mouse, solution containing a drug is infused into the IVC via the same needle, followed by incubation. This method enables more robust drug delivery to the venous endothelium compared to the external route.

マウスの動静脈瘻内皮への管腔内薬物送達

Delivery of therapeutic agents to enhance arteriovenous fistula (AVF) maturation can be administered either via intraluminal or external routes. The ...

Murine Model of Central Venous Stenosis using Aortocaval Fistula with an Outflow Stenosis

Central venous stenosis is an important entity contributing to arteriovenous fistula (AVF) failure. A murine AVF model was modified to create a partial ligation of the inferior vena cava (IVC) in the outflow of the fistula, mimicking central venous stenosis. Technical aspects of this model are introduced. The aorta and IVC are exposed, following an abdominal incision. The infra-renal aorta and IVC are dissected for proximal clamping, and the distal aorta is exposed for puncture. The IVC at the midpoint between the left renal vein and the aortic bifurcation is carefully dissected to place an 8-0 suture beneath the IVC. After clamping the aorta and IVC, an AVF is created by puncturing the infra-renal aorta through both walls into the IVC with a 25 G needle, followed by ligating a 22 G intra-venous (IV) catheter and IVC together. The catheter is then removed, creating a reproducible venous stenosis without occlusion. The aorta and IVC are unclamped after confirming primary hemostasis. This novel model of central vein stenosis is easy to perform, reproducible, and will facilitate studies on AVF failure.

An aortocaval fistula was created by puncturing the murine infra-renal aorta through both walls into the inferior vena cava and was followed by creation of a stenosis in its outflow via partial ligation of the inferior vena cava. This reproducible model can be used to study central venous stenosis.

流出狭窄を伴う大軌道瘻を用した中枢静脈狭窄のマウスモデル

Central venous stenosis is an important entity contributing to arteriovenous fistula (AVF) failure. A murine AVF model was modified to create a partial ...

A Murine Model of Hemodialysis Access-Related Hand Dysfunction

Chronic kidney disease is a major public health problem, and the prevalence of end-stage renal disease (ESRD) requiring chronic renal replacement therapies such as hemodialysis continues to increase. Autogenous arteriovenous fistula (AVF) placement remains a primary vascular access option for ESRD patients. Unfortunately, approximately half of the hemodialysis patients experience dialysis access-related hand dysfunction (ARHD), ranging from subtle paresthesia to digital gangrene. Notably, the underlying biologic drivers responsible for ARHD are poorly understood, and no adequate animal model exists to elucidate the mechanisms and/or develop novel therapeutics for the prevention/treatment of ARHD. Herein, we describe a new mouse model in which an AVF is created between the left common iliac artery and vein, thereby facilitating the assessment of limb pathophysiology. The microsurgery includes vessel isolation, longitudinal venotomy, creation of arteriovenous anastomosis, and venous reconstruction. Sham surgeries include all the critical steps except for AVF creation. Iliac AVF placement results in clinically relevant alterations in central hemodynamics, peripheral ischemia, and impairments in hindlimb neuromotor performance. This novel preclinical AVF model provides a useful platform that recapitulates common neuromotor perturbations reported by hemodialysis patients, allowing researchers to investigate the mechanisms of ARHD pathophysiology and test potential therapeutics.

This protocol details the surgical steps of murine common iliac arteriovenous fistula creation. We developed this model to study hemodialysis access-related limb pathophysiology.

血液透析アクセス関連手機能障害のマウスモデル

Chronic kidney disease is a major public health problem, and the prevalence of end-stage renal disease (ESRD) requiring chronic renal replacement ...

A Modified Technique for Arteriovenous Fistula Construction in Rabbits

Juxta-anastomotic stenosis is a challenging problem that often causes non-maturation and decreases the patency of an arteriovenous fistula (AVF). Injury to the veins and arteries during the operation and hemodynamic changes can lead to intimal hyperplasia, leading to juxta-anastomotic stenosis. To reduce injury to the veins and arteries during the operation, this study proposes a new modified no-touch technique (MNTT) for AVF construction that can decrease the rate of juxta-anastomotic stenosis and improve the AVF patency. To unravel the hemodynamic changes and mechanisms of the MNTT, this study presented an AVF procedure using this technique. Although this procedure is technically challenging, 94.4% procedural success was achieved after adequate training. Ultimately, 13 out of 34 rabbits had a functional AVF 4 weeks after the surgery, leading to a 38.2% AVF patency rate. However, at 4 weeks, the survival rate was 86.1%. Ultrasonography showed active blood flow through AVF anastomosis. Furthermore, the spiral laminar flow was observed in the vein and artery near the anastomosis, suggesting that this technique may improve the hemodynamics of the AVF. On histological observation, significant venous intimal hyperplasia was observed at the AVF anastomosis, whereas no significant intimal hyperplasia was observed at the proximal external jugular vein (EJV) of the anastomosis. This technique will improve the understanding of the mechanisms underlying the use of MNTT for AVF construction and provide technical support for the further optimization of the surgical approach in AVF construction.

The present protocol proposes the creation of an arteriovenous fistula in rabbits using a modified no-touch technique. The technique involves the side-to-side anastomosis of the common carotid artery and external jugular vein without the dissection of the perivenous tissues or cutting off the artery.

ウサギの動静脈瘻構築のための改良技術

Juxta-anastomotic stenosis is a challenging problem that often causes non-maturation and decreases the patency of an arteriovenous fistula (AVF). Injury ...

RVボリューム過負荷マウスモデルを確立するための修正動静脈瘻手術

Establishment and Confirmation of a Postnatal Right Ventricular Volume Overload Mouse Model

Right ventricular (RV) volume overload (VO) is common in children with congenital heart disease. In view of distinct developmental stages,the RV myocardium may respond differently to VO in children compared to adults. The present study aims to establish a postnatal RV VO model in mice using a modified abdominal arteriovenous fistula. To confirm the creation of VO and the following morphological and hemodynamic changes of the RV, abdominal ultrasound, echocardiography, and histochemical staining were performed for 3 months. As a result, the procedure in postnatal mice showed an acceptable survival and fistula success rate. In VO mice, the RV cavity was enlarged with a thickened free wall, and the stroke volume was increased by about 30%-40% within 2 months after surgery. Thereafter, the RV systolic pressure increased, corresponding pulmonary valve regurgitation was observed, and small pulmonary artery remodeling appeared. In conclusion, modified arteriovenous fistula (AVF) surgery is feasible to establish the RV VO model in postnatal mice. Considering the probability of fistula closure and elevated pulmonary artery resistance, abdominal ultrasound and echocardiography must be performed to confirm the model status before application.

著者スポットライト:出生後右室容積過負荷マウスモデルの確立と確認  

This protocol presents the establishment and confirmation of a postnatal right ventricular volume overload (VO) model in mice with abdominal arteriovenous fistula (AVF), which can be applied to investigate how VO contributes to postnatal heart development.