この作品は1R03DK096151によって部分的にサポートされていました。

全てのマウスは、ジャクソン研究所（バーハーバー、メイン州）から購入し、NIHガイドラインとコロラド州デンバーIACUCの大学の承認を得て応じコロラド州デンバー、バーバラ・デイビス・センター動物施設の大学で病原体を含まない条件下で飼育しました。 1.ドナー腎臓収穫<ol><li> 、すべての楽器を滅菌手順を通して滅菌手袋を着用し、無菌領域を維持します。手術用顕微鏡を用いて、すべての手術を行ってください。 </li><li>外科ペントバルビタール麻酔（60mgの/ kgのIP）のドナーからのマウスドナーの腎臓を削除します。つま先ピンチによって麻酔の深さを確認し、呼吸数を観察します。 </li><li> 4ウェイの制約により、マウスを固定化後、ファークリップ。ポビドンヨードで皮膚を準備し、無菌の方法で、マウスを羽織ります。 </li><li> 2cmの正中線垂直腹部切開を行い、腹腔を入力してください。上方腸を撤回し、それを外部化胸の上。手順を通して無菌湿ったガーゼに包まれた腸を保管してください。 </li><li>大きな腹部の血管を識別し、それらを動員、任意の腰椎枝を特定し、焼灼または10/0ナイロン縫合糸でそれらを連結のいずれか。 </li><li>左腎血管へのすぐ遠位約2ミリメートル以上の鈍的切開によって離れて下大静脈（IVC）と腹部大動脈（AA）をいじめます。 10/0ナイロンで結紮し、腎血管から小動脈および静脈枝を分けます。 </li><li>今これらの構造の鈍的切開によって慎重に腎動脈から腎静脈を分離。これは、受信者IVCに端側吻合を形成するために使用される腎静脈の近位端にカレルパッチの正確な作成を可能にします。 </li><li>特定し、ライゲーションし、左副腎血管を分割します。これは、副腎大動脈へのアクセスを許可し、6-0絹縫合糸を結紮の準備のために大動脈の周りに配置されますが、この時点で結ばれていません。 </LI&gt; <li>周囲フェーシャから腎臓、血管および尿管を動員。右と連結するために腎臓を回す/任意の後方分岐を分けます。その後、左に腎臓を返します。 </li><li>今尿管に注意を向けます。腎門部を乱すことなく、尿管、血管を維持するために世話をして、周囲のfaciaから尿管を解放します。精管のレベルで尿管を分割します。腎臓は現在回復するための準備ができました。 </li><li>ゆっくりと、それによってドナーをheparinizing遠位IVCへのヘパリン300単位を注射します。副腎AAの周りに配置され、6-0絹縫合糸を下に結ぶとヘパリン生理食塩水（100 U / ml）を0.8ミリリットル遠位腹大動脈を介して、左腎臓を灌流。 </li><li>灌流は静脈カレルパッチを作成して停止した後はすぐに腎臓に逆流を排除します。 AAと下の腎動脈を明らかにし、腎臓に向かって腎静脈を撤回。 </li><li>腎動脈に隣接する大動脈かかととするの作成を分割図1に示すように、電子カフ。ドナーから腎臓、血管および尿管を外します。 </li><li>移植時まで事前に準備された受信者（0分冷 ​​虚血時間）に直接腎臓を削除するか、事前に決められた時間のための最適なソリューションで4 O℃で保存しました。 exsanguanationと頸椎脱臼によってドナーを安楽死させました。ドナー腎臓を回復するための合計時間は約です。 15〜20分。 </li></ol> 2.腎臓インプラント技術<ol><li>ペントバルビタールでレシピエントマウスを麻酔（60mgの/ kgのIP初期投与量、25mgの/ kgのIP補足用量必要な場合）。つま先ピンチによって麻酔の深さを確認し、呼吸数を観察します。 4ウェイの制約により、マウスを固定化し、眼に眼軟膏を適用し、その後の毛皮をクリップ。ポビドンヨードで皮膚を準備し、無菌の方法で、マウスを羽織ります。 </li><li> 2cmの正中線垂直腹部切開を行い、腹腔を入力してください。腸を撤回上方と胸へ外部化。手順を通して無菌湿ったガーゼに包まれた腸を保管してください。 </li><li>このとき、右腎摘出を行います。 6-0絹縫合糸で腎動脈と静脈を結紮。縫合糸に腎遠位を切り出します。 6-0絹糸で尿管を結紮した後、腎臓の近位に分割します。 </li><li>腎血管下の腹部大動脈と下大静脈（IVC）を分離します。吻合部位に比べて劣っその後大動脈の周りに4-0綿の絆を置き、IVCが優れました。 10-0ナイロン縫合糸でフィールド内の任意の腰椎血管を識別し、連結。 </li><li>コットンネクタイ、優れ続く最初の劣る結び目。このように、いくつかの血液が大動脈切開を作るより簡単に大動脈内に保持されています。 </li><li>大動脈の内腔を入力するように30G針を用いて大動脈切開を形成します。約2ミリメートルの長さを微マイクロハサミで切開を拡張します。 </li><li>ドナー大動脈ヒールアンドトウカフの端部から側部に到る吻合を作ります以下の方法で受信者の大動脈。ドナー大動脈および受信者の大動脈とネクタイの切開部の下角に10-0ナイロン縫合糸滞在ステッチを配置します。 </li><li>ドナー大動脈と腹部大動脈とネクタイの切開部の優れたコーナーの最初の反対の第2の10-0ナイロンを配置します。大動脈の側壁に劣るよりも優れてから実行している縫合線を作成し、以前に配置された滞在ステッチに対して結びます。そして、実行中のファッションやタイで内側壁を縫合。 </li><li>以下の方法で受信者IVCへのドナー腎静脈の端部から側部に到る吻合を行います。 30G針でIVCを穿刺し、細かいマイクロハサミで切開を拡張します。 10-0ナイロンでIVCの切開部の下位隅にドナー腎静脈を接続します。腎静脈と下大静脈とタイの間で実行されている縫合線を作成します。 注：これは、完全な厚さは、血管adventit含む縫合針の通過することも極めて重要ですIAと内膜が達成されます。エッジの裏返しにもシールと吻合の治癒を助ける内膜ツー内膜の接触があることを保証します。止血凝固剤が漏れを減少させるために有用であることができるが、我々は、外科医が代わりに良い技術に依存していることをお勧めします。 </li><li>吻合は「クリーン」であることを確認してください。それはステッチを配置する際に、対向壁がキャッチされていないこと、です。これは失敗した移植片および後肢麻痺に極端な例になりますことを流す重要な収縮を引き起こします。 注：別の極めて重要な要因は、吻合縫合線のテンションも最適であることを保証することです。緩すぎると不可逆的な漏れ、きつすぎるとなります流れるように狭窄があるでしょう。動脈側の場合は混雑した腎臓はなり静脈側の場合、これが、移植片の不十分な灌流になります。 </li><li>静脈の流れを再確立遠位4-0綿のネクタイをリリース。静脈の止血たら私たちの吻合は、近位4-0綿のネクタイを緩め、止血のために動脈吻合を観察徐々に観察されています。 </li><li>両方の吻合が安全と見なされているいったんマウスから綿の関係を削除します。ピアース二つの穴を作成し、20G針で膀胱。 </li><li>貫​​通孔を湾曲した鉗子の先端を通過し、2 10-0ナイロン縫合糸で膀胱壁に固定されて尿管の近位端とピンセットで膀胱を介してドナー尿管を引っ張ります。尿管が膀胱内に撤回し、2 10-0ナイロン縫合糸で穴を閉鎖することを可能にする第二の孔から突出した尿管の余分な長さをトリミング。 </li><li>腹部に腸を返します。実行中のファッションで5-0シルク縫合糸を使用して2つの層で腹壁を閉じます。 </li><li>決算時に輸液蘇生などの腹部に滅菌、温かい生理食塩水1.0mlのボーラスを管理し、通常の生理食塩水を皮下術後の0.8ミリリットルを注入。他のsupportiありませんVEの対策は手術中に必要とされます。 </li><li>温暖化の毛布の上に動物を回復します。総インプラント時間は約です。 35〜45分。このような手順の開始時と72時間後のオペアンプごとに6〜12時間でブプレノルフィン、0.05ミリグラム/キログラム、SC、0.1〜0.2ミリリットルとして鎮痛剤を投与します。 </li></ol> 3.対側腎摘出術<ol><li>プロトコル要件に応じて、数日間は、インプラント手順の後に、イソフルラン麻酔下で対側腎摘出（誘導のための5％吸入イソフルラン、メンテナンスのための1.5から2.0パーセント）を行います。 </li><li>腹腔を入力して、優しく、動物の右に腸を撤回。周囲の筋膜から左腎臓と鈍い解剖を公開します。 </li><li> 6-0絹縫合糸で腎動脈、静脈および尿管を結紮した後、縫合糸の上に腎臓を切除し、腎臓を削除します。総手術時間は約です。 10分。 </li></ol> 4.グラフト評価<ol><li>メジャー血清creatiアルカリ性ピクリン酸法（ヤッフェ反応）を使用して9。 </li></ol>

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The authors have nothing to disclose.

この移植技術をマスターすることは困難であるが、一度それは非常に強力な研究ツールで達成しました。患者外科医/研究者は小さな動物モデルでそれ以上に、任意の外科的手技を習得するための鍵である、細部および技術の整合性に注意することによって報われる。マウスの腎臓移植​​をマスタリングの技術的な問題は、多くのひだであり、それは他の小動物の移植モデルでの経験は、こ...

1973年以来、マウスにおける腎移植モデルは、貴重な研究ツールとなっているが、技術的な問題は、その広範な使用を妨げてきました。年間でいくつかの論文は、この手順の改善/改良を詳細に公開されています。主に血管新生した固形臓器移植のモデルとして、この手順は、また、1973年6でラッセルの研究所で考案された異心臓移植モデルにおそらく秒です。どちらのモデルも、同種異系拒絶反応、遅延移植片機能の開発に検索するには自分自身を貸しますそして、虚血再灌流障害。 腎臓移植に報告するための最も一般的な問題の一つは、我々はまた、我々の研究室で経験した動脈血栓症4,5,7の比較的高い発生率です。そこで我々は、血栓形成の文献レビューを行い、場合によっては、この技術的な問題の原因を見つけるために着手したとも考案します可能な解決策。血栓症の最も可能性の高い原因は、血液がドナー腎動脈の上、次にドナー腎大動脈に、受信者の大動脈から取り多少曲がりくねった道です。この経路は、血小板活性化および血栓形成を引き起こし得る、腎動脈の乱れを引き起こします。最近の観察と関連文献8-14の検索に基づいて、我々は0％に血栓症を減少させた新しい技術を思い付きました。 ここに記載の技術は、施設は、血流を改善し、動脈ヒールアンドトウカフの形成以前に報告の手法によって異なると著しく血栓形成を減少させます。カフ未満45 O大動脈（ 図1Aおよび1B）の長手方向軸に対してある角度で、腎動脈口の面を横切る赤外線の腎動脈を分割して形成されています。この長さは約2ミリメートルカフをもたらします。静脈カレルパッチをrを横断することによって形成されますIVCにエナール静脈ことにより、カフの直径を増加させます。赤外線腎ドナー腹部大動脈ヒールアンドトウカフは、レシピエントの腹部大動脈およびドナー腎静脈/ IVCパッチに吻合端側である受信者の腹部下大静脈（IVC）に吻合端側であります。尿管は、その後に導入され、Hanら3に記載されているように、膀胱に固定されます。 この研究のためにのみ温虚血時間（ すなわち 、ノー冷たい虚血）で処理されていない移植が比較されます。この場合、暖かい虚血は、レシピエント（ステップ以下2.11）、ドナーの腎臓を通る血流の停止（以下ステップ1.11）、および移植片の再潅流からの時間を指します。冷虚血は、腎臓を灌流されていないインプラント手順の開始まで冷蔵で維持される時間を指します。

<table><tbody><tr><td>Instrument</td><td>Roboz #</td><td>Fine Science Tools #</td><td>Arosurgical #</td></tr><tr><td>Straight micro-dissecting forcep #5</td><td>RS-5015</td><td>11295-51</td><td></td></tr><tr><td>Curved micro-dissecting forcep #7</td><td>RS-5047</td><td>11297-00</td><td></td></tr><tr><td>Curved serrated forcep</td><td>RS-5137</td><td>11052-10</td><td></td></tr><tr><td>Vannas micro-dissecting scissors, short</td><td>RS-5610</td><td></td><td>09.140.08</td></tr><tr><td>Micro-dissecting scissors, straight, sharp, long</td><td></td><td>11.602.11</td><td></td></tr><tr><td>Micro spring handle needle holder</td><td></td><td></td><td>11.549.15</td></tr><tr><td>Straight mosquito forcep</td><td></td><td>91308-12</td><td></td></tr><tr><td>Micro-dissecting scissors, straight, blunt</td><td>RS-5962</td><td>14078-10</td><td></td></tr><tr><td>Micro-dissecting scissors, curved, blunt</td><td>RS-5981</td><td>14079-10</td><td></td></tr><tr><td>Micro retractor</td><td>RS-6540</td><td></td><td></td></tr><tr><td>Instrument tray, 10&amp;rdquo; x 6 &amp;frac12;&amp;rdquo; x &amp;frac34;&amp;rdquo;</td><td>RT-1350S</td><td></td><td></td></tr><tr><td>Silk suture, 5/0, 22.5m spool</td><td></td><td>18020-50</td><td></td></tr><tr><td>Suture</td><td></td><td></td><td></td></tr><tr><td>10/0 nylon</td><td></td><td></td><td>T4A10Q07</td></tr><tr><td>5/0 silk</td><td></td><td></td><td>E19A05N</td></tr><tr><td>Gloves</td><td></td><td>Drapes</td><td></td></tr><tr><td>Biogel from Medex Supply</td><td></td><td>Precept, #64-9012-9</td><td></td></tr><tr><td>Syringes</td><td></td><td>Cotton applicators</td><td></td></tr><tr><td>B-D 1cc insulin, #329424</td><td></td><td>Fisher-brand, #23-400-100</td><td></td></tr><tr><td>Povidone-Iodine swabs</td><td></td><td></td><td></td></tr><tr><td>PDI, #B40600</td><td></td><td></td><td></td></tr><tr><td>4/0 Cotton ties</td><td></td><td></td><td></td></tr><tr><td>Domestic cotton autoclaved with instruments</td><td></td><td></td><td></td></tr></tbody></table>

murine kidney transplant technique

The first mouse kidney transplant technique was published in 19731 by the Russell laboratory. 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 using the donor aorta to form the arterial anastomosis instead of the renal artery was developed and reported in 1993 by Kalina and Mottram 2 with a further advancement coming from the same laboratory in 1999 3. While one can become proficient in this model, a search of the literature reveals that many labs still experience a high proportion of graft loss due to arterial thrombosis. We describe here a technique that was devised in our laboratory that vastly reduces the arterial thrombus reported by others 4,5. This is achieved by forming a heel-and-toe cuff of the donor infra-renal aorta that facilitates a larger anastomosis and straighter blood flow into the kidney.

この手術法は、単純な移植片生存/拒絶の研究、または非常に複雑な実験プロトコルのいずれかを可能にします。以下の図では、この改良された動脈吻合の技術を使用することの利点を実証します。この技術を用いて、我々は著しく生産性を増加させる0％と35％の動脈血栓症の発生率を減少させました。我々は維持し、同じ0％血栓症の結果で1年以上のためにこの技術を使用している。 ?...

Watch this Scientific Journal Video about Murine Kidney Transplant Technique at JoVE.com

Murine Kidney Transplant Technique

The goal of this manuscript is to describe the steps required to perform a kidney transplant in a mouse, paying particular attention to the details of the arterial anastomosis.

マウスの腎臓移植​​技術

The first mouse kidney transplant technique was published in 19731 by the Russell laboratory. Although it took some years for other labs to become ...

Universidad de Cartagena UDEC

Research

JoVE Journal

Medicine

16.0K ビュー.  University of Colorado, Denver. The goal of this manuscript is to describe the steps required to perform a kidney transplant in a mouse, paying particular attention to the details of the arterial anastomosis. 

ビデオ: Murine Kidney Transplant Technique

A Method for Murine Islet Isolation and Subcapsular Kidney Transplantation

Since the early pioneering work of Ballinger and Reckard demonstrating that transplantation of islets of Langerhans into diabetic rodents could normalize their blood glucose levels, islet transplantation has been proposed to be a potential treatment for type 1 diabetes 1,2. More recently, advances in human islet transplantation have further strengthened this view 1,3. However, two major limitations prevent islet transplantation from being a widespread clinical reality: (a) the requirement for large numbers of islets per patient, which severely reduces the number of potential recipients, and (b) the need for heavy immunosuppression, which significantly affects the pediatric population of patients due to their vulnerability to long-term immunosuppression. Strategies that can overcome these limitations have the potential to enhance the therapeutic utility of islet transplantation.
Islet transplantation under the mouse kidney capsule is a widely accepted model to investigate various strategies to improve islet transplantation. This experiment requires the isolation of high quality islets and implantation of islets to the diabetic recipients. Both procedures require surgical steps that can be better demonstrated by video than by text. Here, we document the detailed steps for these procedures by both video and written protocol. We also briefly discuss different transplantation models: syngeneic, allogeneic, syngeneic autoimmune, and allogeneic autoimmune.

Transplantation of isolated islets has been proposed to be a potential treatment for type 1 diabetes. Here we describe a method to isolate islets from mouse pancreata and transplant them to the subcapsular space of the kidney.

マウス膵島の単離と被膜下腎移植のための方法

Since the early pioneering work of Ballinger and Reckard demonstrating that transplantation of islets of Langerhans into diabetic rodents could normalize ...

医学

Murine Renal Transplantation Procedure

Renal orthotopic transplantation in mice is a technically challenging procedure. Although the first kidney transplants in mice were performed by Russell et al over 30 years ago (1) and refined by Zhang et al years later (2), few people in the world have mastered this procedure. In our laboratory we have successfully performed 1200 orthotopic kidney transplantations with &gt; 90% survival rate. The key points for success include stringent control of reperfusion injury, bleeding and thrombosis, both during the procedure and post-transplantation, and use of 10-0 instead of 11-0 suture for anastomoses.
Post-operative care and treatment of the recipient is extremely important to transplant success and evaluation. All renal graft recipients receive antibiotics in the form of an injection of penicillin immediately post-transplant and sulfatrim in the drinking water continually. Overall animal health is evaluated daily and whole blood creatinine analyses are performed routinely with a portable I-STAT machine to assess graft function.

Renal transplantation in mice is a technically challenging procedure that requires careful post-operative care and treatment for success.

マウス腎移植の手順

Renal orthotopic transplantation in mice is a technically challenging procedure. Although the first kidney transplants in mice were performed by Russell ...

生物学

Isolation and Transplantation of Different Aged Murine Thymic Grafts.

The mechanisms that regulate the efficacy of thymic selection remain ill-defined. The method presented here allows in vivo analyses of the development and selection of T cells specific for self and foreign antigens. The approach entails implantation of thymic grafts derived from various aged mice into immunodeficient scid recipients. Over a relatively short period of time the recipients are fully reconstituted with T cells derived from the implanted thymus graft. Only thymocytes seeding the thymus at the time of isolation undergo selection and develop into mature T cells. As such, changes in the nature and specificity of the engrafted T cells as a function of age-dependent thymic events can be assessed. Although technical expertise is required for successful thymic transplantation, this method provides a unique strategy to study in vivo a wide range of pathologies that are due to or a result of aberrant thymic function and/or homeostasis.

This report provides a detailed description of transplanting murine thymi from different aged donor mice under the kidney capsule of immunodeficient mouse recipients. The goal of this approach is to model T cell development and thymic selection events in vivo.

単離と異なる高齢マウス胸腺移植片の移植。

The mechanisms that regulate the efficacy of thymic selection remain ill-defined. The method presented here allows in vivo analyses of the development and ...

免疫・感染学

Mouse Kidney Transplantation: Models of Allograft Rejection

Rejection of the transplanted kidney in humans is still a major cause of morbidity and mortality. The mouse model of renal transplantation closely replicates both the technical and pathological processes that occur in human renal transplantation. Although mouse models of allogeneic rejection in organs other than the kidney exist, and are more technically feasible, there is evidence that different organs elicit disparate rejection modes and dynamics, for instance the time course of rejection in cardiac and renal allograft differs significantly in certain strain combinations. This model is an attractive tool for many reasons despite its technical challenges. As inbred mouse strain haplotypes are well characterized it is possible to choose donor and recipient combinations to model acute allograft rejection by transplanting across MHC class I and II loci. Conversely by transplanting between strains with similar haplotypes a chronic process can be elicited were the allograft kidney develops interstitial fibrosis and tubular atrophy. We have modified the surgical technique to reduce operating time and improve ease of surgery, however a learning curve still needs to be overcome in order to faithfully replicate the model. This study will provide key points in the surgical procedure and aid the process of establishing this technique.

Here, we present a protocol to study the immunology of rejection. The surgical model presented reports a short operating time and a concise technique. Depending on the donor-recipient strain combination, the transplanted kidney may develop acute cellular rejection or chronic allograft damage, defined by interstitial fibrosis and tubular atrophy.

マウス腎臓移植：同種移植片拒絶のモデル

Rejection of the transplanted kidney in humans is still a major cause of morbidity and mortality. The mouse model of renal transplantation closely ...

Fate Mapping of Human Embryonic Stem Cells by Teratoma Formation 

Human embryonic stem cells (hESCs) have an unlimited capacity for self-renewal, and the ability to differentiate into cells derived from all three embryonic germ layers (1). Directed differentiation of hESCs into specific cell types has generated much interest in the field of regenerative medicine (e.g., (2-5)), and methods for determining the in vivo fate of selected or manipulated hESCs are essential to this endeavor. We have adapted a highly efficient teratoma formation assay for this purpose. A small number of specifically selected hESCs is mixed with undifferentiated wild type hESCs and Phaseolus vulgaris lectin to form a cell pellet. This is grafted beneath the kidney capsule in an immunodeficient mouse. As few as 2.5 x 105 hESCs are needed to form a 16 cm3 teratoma within 8-12 weeks. The fate of the originally selected hESCs can then be determined by immunohistochemistry. This method provides a valuable tool for characterizing tissue-specific reagents for cell-based therapy.

Fate Mapping of Human Embryonic Stem Cells by Teratoma Formation

Directed differentiation of hESCs into specific cells has generated much interest in regenerative medicine. We provide a concise, step-by-step protocol for determining the in vivo fate of selected hESCs that provides a valuable tool for characterizing tissue-specific reagents for cell-based therapy.

テラトーマ形成により、ヒト胚性幹細胞の運命のマッピング

Human embryonic stem cells (hESCs) have an unlimited capacity for self-renewal, and the ability to differentiate into cells derived from all three ...

Heterotopic Renal Autotransplantation in a Porcine Model: A Step-by-Step Protocol

Kidney transplantation is the treatment of choice for patients suffering from end-stage renal disease. It offers better life expectancy and higher quality of life when compared to dialysis. Although the last few decades have seen major improvements in patient outcomes following kidney transplantation, the increasing shortage of available organs represents a severe problem worldwide. To expand the donor pool, marginal kidney grafts recovered from extended criteria donors (ECD) or donated after circulatory death (DCD) are now accepted for transplantation. To further improve the postoperative outcome of these marginal grafts, research must focus on new therapeutic approaches such as alternative preservation techniques, immunomodulation, gene transfer, and stem cell administration.
Experimental studies in animal models are the final step before newly developed techniques can be translated into clinical practice. Porcine kidney transplantation is an excellent model of human transplantation and allows investigation of novel approaches. The major advantage of the porcine model is its anatomical and physiological similarity to the human body, which facilitates the rapid translation of new findings to clinical trials. This article offers a surgical step-by-step protocol for an autotransplantation model and highlights key factors to ensure experimental success. Adequate pre- and postoperative housing, attentive anesthesia, and consistent surgical techniques result in favorable postoperative outcomes. Resection of the contralateral native kidney provides the opportunity to assess post-transplant graft function. The placement of venous and urinary catheters and the use of metabolic cages allow further detailed evaluation. For long-term follow-up studies and investigation of alternative graft preservation techniques, autotransplantation models are superior to allotransplantation models, as they avoid the confounding bias posed by rejection and immunosuppressive medication.

Porcine models of organ transplantation provide an important platform to study mechanisms of organ preservation. This article describes a heterotopic porcine renal autotransplantation model, which allows investigating new approaches to improve the outcome of transplantation using marginal kidney grafts.

ブタモデルにおける異自家腎移植：ステップバイステップのプロトコル

Kidney transplantation is the treatment of choice for patients suffering from end-stage renal disease. It offers better life expectancy and higher quality ...

Production of Humanized Mouse via Thymic Renal Capsule Grafting, CD34+ Cells Injection, and Cytokine Delivery

Animal models provide a vital translation between in vitro and in vivo biomedical research. Humanized mouse models provide a bridge in the representation of human systems, thereby allowing for a more accurate study of pathogenesis, biomarkers, and many other scientific queries. In this method described, immune-deficient NOD-scid IL2R&#947;null (NSG) mice are implanted with autologous thymus, injected with liver-derived CD34+ cells followed by a series of injected cytokine deliveries. In contrast to other models of a similar nature, the model described here promotes an improved reconstitution of immune cells by delivering cytokines and growth factors via transgenes encoded in AAV8 or pMV101 DNA-based vectors. Moreover, it offers long-term stability with reconstituted mice having an average lifespan of 30 weeks after CD34+ injections. Through this model, we hope to provide a stable and impactful method of studying immunotherapy and human disease in a murine model, thus demonstrating the need for predictive preclinical models.

<meta charset="utf8"></head><body>胸腺腎カプセル移植、CD34+細胞注入、およびサイトカイン送達によるヒト化マウスの作製

Humanized mouse models provide a more accurate representation of the human immune microenvironment. This manuscript describes the process in which these models are created through a renal graft of human thymus, injection of human CD34+ cells, and the targeted delivery of human cytokine transgenes to promote CD34+ cell proliferation and differentiation.

胸腺腎カプセル移植、CD34+ 細胞注入、およびサイトカイン送達によるヒト化マウスの作製

Animal models provide a vital translation between in vitro and in vivo biomedical research. Humanized mouse models provide a bridge in the representation ...

A Modified Surgical Technique for Kidney Transplantation in Mice

Kidney transplantation in mice is a complicated and challenging surgery procedure. There are very few publications demonstrating the key steps of this operation. Therefore, this article introduces the technique and points out the surgical caveats associated with this operation. In addition, important modifications in comparison to the conventional procedure are demonstrated. Firstly, a patch of the abdominal aorta is cut and prepared so that the proximal bifurcations of the renal artery, including the ureteral artery are transected together with the donor kidney en bloc. This reduces the risk of a ureter necrosis and avoids the development of a urinary tract occlusion. Secondly, a new method of the vascular anastomosis is demonstrated that allows the operator to flexibly increase or decrease the size of the anastomosis after renal transplant reperfusion has already been initiated. This avoids the development of vessel strictures and intraabdominal bleeding. Thirdly, a technique that enables the anastomosis of the delicate donor ureter and the recipient bladder that does not cause a trauma is shown. Adopting this protocol can shorten the operation time and reduces the damage to the recipient's bladder, thereby significantly increasing the operation success rate for the recipient mice.

This protocol presents a new surgical technique of mouse kidney transplantation focusing on a modified arterial anastomosis strategy. A vascular suture technique including a simple and safer ureter-bladder anastomosis method is also presented. These modifications shorten the operation time and improve the success rate of the mouse kidney transplantation procedure.

マウスの腎臓移植のための修正外科的技術

Kidney transplantation in mice is a complicated and challenging surgery procedure. There are very few publications demonstrating the key steps of this ...

Orthotopic Rat Kidney Transplantation: A Novel and Simplified Surgical Approach

Kidney transplantation offers increased survival rates and improved quality of life for patients with end-stage renal disease, as compared to any type of renal replacement therapy. Over the past few decades, the rat kidney transplantation model has been used to study the immunological phenomena of rejection and tolerance. This model has become an indispensable tool to test new immunomodulatory pharmaceuticals and regimens prior to proceeding with expensive preclinical large animal studies.
This protocol provides a detailed overview of how to reliably perform orthotopic kidney transplantation in rats. This protocol includes three distinctive steps that increase the probability of success: perfusion of the donor kidney by flushing through the portal vein and the use of a cuff system to anastomose the renal veins and ureters, thereby decreasing cold and warm ischemia times. Using this technique, we have achieved survival rates beyond 6 months with normal serum creatinine in animals with syngeneic or tolerant kidney transplants. Depending on the aim of the study, this model can be modified by pre- or posttransplant treatments to study the acute, chronic, cellular, or antibody-mediated rejection. It is a reproducible, reliable, and cost-effective animal model to study different aspects of kidney transplantation.

The purpose of this manuscript and protocol is to explain and demonstrate in detail the surgical procedure of orthotopic kidney transplantation in rats. This method is simplified to achieve the correct perfusion of the donor kidney and shorten the reperfusion time by using the venous and ureteral cuff anastomosis technique.

Orthotopic ラット腎移植: 新規かつ簡便な外科的アプローチ

Kidney transplantation offers increased survival rates and improved quality of life for patients with end-stage renal disease, as compared to any type of ...

マウスの腎臓移植技術

要約

さらに動画を探す

マウス膵島の単離と被膜下腎移植のための方法

マウス腎移植の手順

単離と異なる高齢マウス胸腺移植片の移植。

マウス腎臓移植：同種移植片拒絶のモデル

テラトーマ形成により、ヒト胚性幹細胞の運命のマッピング

ブタモデルにおける異自家腎移植：ステップバイステップのプロトコル

胸腺腎カプセル移植、CD34⁺ 細胞注入、およびサイトカイン送達によるヒト化マウスの作製

胸腺腎カプセル移植、CD34⁺ 細胞注入、およびサイトカイン送達によるヒト化マウスの作製

マウスの腎臓移植のための修正外科的技術

Orthotopic ラット腎移植: 新規かつ簡便な外科的アプローチ

マウスの腎臓移植​​技術

要約

さらに動画を探す

マウス膵島の単離と被膜下腎移植のための方法

マウス腎移植の手順

単離と異なる高齢マウス胸腺移植片の移植。

マウス腎臓移植：同種移植片拒絶のモデル

テラトーマ形成により、ヒト胚性幹細胞の運命のマッピング

ブタモデルにおける異自家腎移植：ステップバイステップのプロトコル

胸腺腎カプセル移植、CD34+ 細胞注入、およびサイトカイン送達によるヒト化マウスの作製

胸腺腎カプセル移植、CD34+ 細胞注入、およびサイトカイン送達によるヒト化マウスの作製

マウスの腎臓移植のための修正外科的技術

Orthotopic ラット腎移植: 新規かつ簡便な外科的アプローチ

マウスの腎臓移植技術

胸腺腎カプセル移植、CD34⁺ 細胞注入、およびサイトカイン送達によるヒト化マウスの作製

胸腺腎カプセル移植、CD34⁺ 細胞注入、およびサイトカイン送達によるヒト化マウスの作製