The authors would like to acknowledge funding support from the University of South Florida, Department of Surgery (Mack Wu, MD, PhD).

注：以下に示すすべての動物の手順のための承認は南フロリダの動物管理委員会（IACUC）の大学から入手し、動物福祉法および実験動物の管理と使用に関する指針のすべての要件を遵守した。 シリコーンシートと手術器具の作製<ol><li>非強化0.01厚さ、医療グレードのシリコーンシートとオートクレーブを用いて滅菌のプレカットストリップ（X 3.0センチメートル10.5センチメートル）。 </li><li>清潔（手術中に無菌のフィールドを作成するハサミ、ピンセットやカーテンやタオル）、適切な手術器具を滅菌する。 </li></ol> 2.実験動物<ol><li>商業ブリーダーから入手した250〜350グラムの範囲で体重成人男性または雌ラットを使用してください。老齢ラットが利用される場合、それらは良好な生存、術後性を確保するために≥350gであるべきである。すべての実験の開始前には、少なくともためにすべての動物を順化食物および水を自由に摂取 12時間の明暗サイクルの標準的な条件下で7日間。 </li></ol> 3.麻酔、術前鎮痛と手術の準備<ol><li>皮膚の準備と手術中にO 2と1％-2％に（ノーズコーンの使用を介して）誘導室を介して3％-4％のイソフルランを使用して全身麻酔を誘導し、維持する。呼吸の速度と深さの観察、インターデジタルピンチまたは眼瞼瞬目反射によって麻酔の深さを監視します。 注：この時点では、獣医軟膏は、動物が麻酔下にある間に乾燥を防ぐために、目に配置することができる。 </li><li>リモート無菌の手術領域からの位置では、腹臥位にラットを置き、上下約11センチメートル首の付け根からバリカンで背を剃る。恒久的なマーカーでステンシル、X 10.5センチフラップ3.0センチメートルのアウトライン（ 図1Aを参照）。 </li><li>きれいな、指定されたsurgicaにラットを移動承認された加熱パッド及び滅菌外科用ドレープまたはタオルを搭載したL·エリア。疼痛管理のための最初の外科的切開に5mg / kgのケトプロフェンを皮下注射する前に。必要に応じて、追加の流体（生理食塩水）は、皮下（5 CCまで）が挙げられる。 </li><li>無菌のフィールドを作成する無菌ドレープを適用した後、0.2％クロルヘキシジンと70％イソプロピルアルコール及び第二と第一拭き取りによってさらに肌を準備します。 10％ポビドンヨード（ベタジン）を使用することもできる。 NOTE：抗生物質（15 mg / kgをアンピシリン）を皮下投与することができるが、良好な無菌技術が使用される場合、それは要求されない。 </li></ol> 4.切除創傷とBipedicledフラップの作成<ol><li>無菌の、使い捨ての6ミリメートル生検パンチツールを使用して、指定されたフラップ面積（ 図1B）の中央に2円形「虚血性&quot;傷を作る。傷の深さはダウン（しないを介して）に皮筋層のCarnoのの基本的な筋膜であるべきSUS筋肉（ 図1B挿入図）。 </li><li>使用して鉗子は、パンチ生検によって作成された創傷の輪郭の途中で皮膚を持ち上げた後、（皮筋層の筋肉を含む）は、組織の円形部分を切除する（湾曲した先端を有する）アイリスハサミを使用しています。その結果、傷のベースとして筋膜で巻か全層になります。 注：切除した組織（創傷プラグ）スナップ液体窒素中で凍結させるか、または制御、正常な皮膚のような後の処理のために10％緩衝ホルマリンO / Nに固定することができる。 </li><li>長さ10.5センチ、離れて3.0センチメートルである前描線（ 図1C）に沿って、虚血性傷の両側に滅菌メスで切開を行うことによってbipedicledフラップを作成します。切開の深さは、ダウン傍脊椎筋にすべきである。アイリスハサミを使用して、傍脊椎筋から皮筋層の筋膜を分離、6mmのパンチ（Fの「ベース」としてそのまま筋膜を維持するように注意してigure 1D）。 </li><li> 1無菌プレカットシリコーンシートを取り、皮筋層の筋膜とシートが座屈しかフォールドしないことを確実にする傍脊椎筋（ 図1E）の間にそれを配置します。ブラック、非吸収性縫合糸（サイズ4.0）を使用して、少なくとも8で皮膚へのシリコーンシートを固定することによって、両方の切開を閉じるフラップ（ 図1Fおよび1G）の長さに沿って、それぞれの側にステッチを中断。 </li><li>無菌の、使い捨ての生検パンチツールを使用して、（皮筋層の筋肉の前筋膜まで）2、内部統制の「非虚血性」の傷フラップ（ 図1G）のいずれかの側の虚血性創傷に横1cmに作成します。 </li><li>傷下記の定規を置き、（ 図3Aを参照）に巻か測定目的のために、デジタル写真を撮る。このとき、血流（灌流）は、レーザードップラーを使用してモニターすることができ、または他の操作（局所薬の配置）を行う。</li><li>頭蓋と尾の両方に承認液体接着剤は、創傷及び（無菌）しっとりとした清潔な創傷環境を維持するためにドレッシング透明フィルムに適用されます。追加のドレッシングは、最も尾側の縫合糸を除去した動物を防ぐために、フラップの尾側端部に配置することができる。 </li></ol> 5.術後の手順<ol><li>フィーダー擦れから手術部位を防止するように浅いフィーダーを装備した（単独で収容）のケージに動物を配置します。彼らは胸骨横臥を維持し、意図的な動きを示すのに十分な意識を取り戻すまで、動物は、放置または他の動物の会社に返却すべきではありません。暖房マットは回復中に最大2日間、ケージの半分の下に配置する必要があります。 </li><li>手術後の痛みを管理するには、最大48時間後に手術のために次の朝、一日あたり1倍の動物に皮下にケトプロフェン（5mg / kgの）を管理。動物はまたのために毎日監視する必要があります痛み、体重減少または手術部位感染の長期化の兆候。 </li></ol> 6.その後の創傷測定と包帯交換<ol><li>ステップ3.1のように、O 2と誘導室を介して3％-4％のイソフルランを使用して、頻繁に全身麻酔下で虚血性および非虚血性の傷を測定し、維持（ノーズコーンを介した）1％-2％である。 </li><li>皮膚から接着剤を引っ張らないよう優しくドレッシングを削除します。追加のデジタル写真は、創傷の測定のために採取され、この時点で、局所治療は再適用、レーザードップラー画像化（LDI）または研究者のニーズに合わせて行う他の操作。 </li><li>接着剤ときれいなドレッシングを適用し、動物がステップ5.1のように回復することができます。 </li></ol> 7.傷収集と安楽死<ol><li>収穫虚血性および非虚血性の傷（日に調査員が適当と認める）動物はINDを経由して3％-4％のイソフルランを使用して全身麻酔下にありながら、はじめ室とステップ3.1のようにO 2と1％-2％に（ノーズコーンを介して）維持した。 </li><li>メスを用いて、傷の周りのいくつかの健康な組織が含まれるように傷の周りに正方形の切除を行う。キャップチューブをスナップし、将来分子解析のために液体窒素中で凍結（-80℃で保存）をスナップインまたは組織学的処理のために、室温で10％緩衝ホルマリンO / Nでインキュベート1.5ミリリットルに切除を置きます。 NOTE：創傷切除はまた、分析のためのサンプルを提供するために半分に切断することができる。 </li><li>創傷組織を除去した後、CO 2吸入の承認された方法を用いて動物を安楽死させる。 </li></ol>

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The authors have nothing to disclose.

<html>ラットでの創傷治癒は、しばしば感染した傷や動物間の変動5率の高さを癒すする能力に議論の対象となっている。その開発中のモデルの元の目標の一つは、この変動を減少しております。特定の配置に傷の数を減少させるフラップの幅に変更が、シリコンシートのと導入（一貫した頭尾場所にフラップを中心）が、この目標を達成した。収縮により創傷治癒が、ヒトにおけるよ?...

効果的な薬剤開発やその他の創傷治癒治療薬は人間の治療を1に動物モデルでの調査結果を翻訳における既知の問題にもかかわらず、in vivoモデルで適切な必要とする。以下は、病理学的なメカニズムは、創傷治癒のさらなる理解を調査するために、虚血皮膚創傷治癒のラットモデルを使用するための詳細なプロトコールを説明する。ラット種は、しばしば、その広い利用可能性、大きさに起因して採用し、切開し、切除創傷、イメージング及び組織採取2適した皮膚領域を提供するのに十分な大きさであるように従順な性質は、創傷治癒研究に使用される。しかしそれはラットのように緩い皮の動物と呼ばれていると、ラットおよびヒトの皮膚は解剖学的に異なっている慎重に考慮に入れるべきである。この明確な特徴は、ラットの皮膚の閉鎖ワットに大きく貢献するというよりも、上皮形成、創傷収縮が可能にounds 2。さらに、ラットにおける皮下皮筋層の筋の存在は、収縮およびコラーゲン形成3,4の両方によって治癒に寄与する。これらの非常に重要な解剖学的な違いは、ラット虚血皮膚創傷モデルの開発において考慮され、特定の改変は、創傷収縮を減少させ、皮筋層の筋5の影響を低減するために実施された。 糖尿病性足部潰瘍、静脈性下腿潰瘍、および褥瘡では、治癒が遅延され、これらの創傷は、慢性と見なされる。創傷は、創傷修復6の次の段階に進むの傷を防止する過剰な炎症によって特徴付けられる。慢性創傷の開発における主な要因の1つは、炎症をクリアすることができないことに寄与して局所的な組織虚血（血流低下）5である。このモデルは2003-4で開発され、（検証された時間で、）、創傷床、正常な創傷治癒中にキーステージと、このモデル5を開発するための動機で血管新生の誘導を試験するために十分な組織を提供することができる標準化された動物モデルは存在しなかった。つまり、ここで紹介するモデルは、チェンらによって修正された形で使用する元々シュワルツら 7、その後で記述虚血性創傷モデルの変形である、と述べた。8 修正された虚血性創傷モデルでは、変更はなく、上皮化よりも収縮によって治癒につながるラットの上記の解剖​​学的特性を回避するために行われています（1）2つの全層切除創傷はbipedicled背面皮膚フラップと内に作成されている皮筋層カルノーサス筋肉は単に筋肉の筋膜の上に解剖することにより、創傷床から削除されます。 （2）フラップ自体は、血液供給がランダムであり、傷をtの中間点に位置することを確実にする、より狭い寸法を有している彼はフラップ虚血である。 （3）Aシリコーンシートは、下にある組織からのフラップの再付着および再灌流を防止フラップの下に挿入されている。創傷収縮は、アンカーやシリコーンシート5にフラップを縫合することによって（排除しない）制限されています。 モデルは、最近、老齢ラット11対若年における虚血性創傷治癒に9,10治癒 、虚血性創傷における高圧酸素効果に至るまでの研究で使用されており、長時間の組織虚血の信頼できるモデルであることが証明された。 bipedicledフラップの寸法も豚12とマウスを含むと他の種、（3.0〜3.5センチメートルワイド10.5センチ）とF344ラット（2センチ幅11によってセンチ）のSprague Dawley含む異なるラット系統に適応されている13,14。このビデオでは、虚血性皮膚創傷モデルのデモンストレーションでF344近交系ラット系統を利用しています。 以下に、すべての動物の手順のための承認を得るのだった南フロリダの動物管理委員会（IACUC）の大学編と動物福祉法および実験動物の管理と使用に関する指針のすべての要件を順守する。

<table border="0" cellpadding="0" cellspacing="0" width="605">
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		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Name of Reagent/ Equipment</td>
			<td style="width:103px;">Company</td>
			<td style="width:119px;">Catalog Number</td>
			<td style="width:167px;">Comments/Description</td>
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			<td style="width:103px;">Technical Products Inc.</td>
			<td style="width:119px;">500-3</td>
			<td>nonreinforced, 0.01 inches in thickness</td>
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			<td style="width:119px;">#503671</td>
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			<td height="21" style="height:21px;width:216px;">Ethilon Nylon Sutures</td>
			<td style="width:103px;">Ethicon</td>
			<td style="width:119px;">1964G</td>
			<td>black, size 4.0, PC-3 16mm needles (3/8 circle)</td>
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			<td height="41" style="height:41px;width:216px;">Laser Doppler Imager</td>
			<td style="width:103px;">Moor Instruments</td>
			<td style="width:119px;">moorLDI2-IR</td>
			<td>Standard blood flow imager: http://us.moor.co.uk/product/moorldi2-laser-doppler-imager/8</td>
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			<td height="21" style="height:21px;width:216px;">ImageJ</td>
			<td style="width:103px;">NIH</td>
			<td style="width:119px;">free download</td>
			<td>http://rsb.info.nih.gov/ij/</td>
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			<td height="21" style="height:21px;width:216px;">Mastisol</td>
			<td style="width:103px;">Henry Schein</td>
			<td style="width:119px;">Cat # 7289210</td>
			<td>Fisher Scientific NC9774929</td>
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			<td height="41" style="height:41px;width:216px;">Tegaderm</td>
			<td style="width:103px;">Medical Specialties&#160;</td>
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demonstration of the rat ischemic skin wound model

ヒトでの慢性創傷の傾向は、そのような原因不動に糖尿病や障害のある心血管機能、および単調な圧力などの老化、疾患状態に伴って増加する。動物モデルは、慢性創傷の複雑さの理解を促進する目的で、これらの条件を模倣しようと試みて開発されてきた。本明細書に記載されたモデルは、虚血になる傷が生じる血流量の長期の減少を可能にするラット虚血皮膚弁モデルであり、慢性創傷の表現型に似ている（縮小血管新生、炎症および遅延創傷閉鎖の増加）。それは、対照としてフラップに横方向中央に2虚血性創傷及び2非虚血性の傷付きbipedicled背側フラップで構成されています。この虚血皮膚弁モデルに対する新規追加は、障壁として機能するフラップの下のシリコンシートと血管再生を防止し、創傷が治癒するように収縮を低減するスプリントの配置である。にもかかわらず人間に比べて、それらは全く異なる解剖学的および生理学的な違いのための研究を、創傷治癒のためにラットを用いての議論（ すなわち、皮筋層の筋肉、短い寿命、毛包の数が増加し、感染した傷を癒すする能力の有無）このモデルで採用の変更は、それ以前に開発された虚血性皮膚弁モデルへの貴重な代替手段にする。

効率的に行う場合は、ラットの虚血性創傷治癒モデルプロトコルは、動物当たり約20分を取る必要があります。 図1Gに示すようにモデルをドレッシングの適用に先立ちが表示されます。それはbipedicledフラップと傷がその中の虚血性であることを確認することが重要になります。傷のレベルで皮下酸素張力（PscO 2）は 2虚血性傷の間に皮下組織にポーラログラフ電極を配置?...

Watch this Scientific Journal Video about Demonstration of the Rat Ischemic Skin Wound Model at JoVE.com

Demonstration of the Rat Ischemic Skin Wound Model

そのサイズ、可用性、むしろ従順行動によるラットは、長年の研究モデルとして利用されている。このプロトコルの目的は、慢性創傷の病態生理学に貴重な洞察を提供するために、虚血皮膚創傷治癒モデルとしてラットを利用することである。

ラット虚血皮膚創傷モデルの実証

The propensity for chronic wounds in humans increases with ageing, disease conditions such as diabetes and impaired cardiovascular function, and ...

demonstration-of-the-rat-ischemic-skin-wound-model

Research

JoVE Journal

Medicine

21.1K ビュー.  University of South Florida. そのサイズ、可用性、むしろ従順行動によるラットは、長年の研究モデルとして利用されている。このプロトコルの目的は、慢性創傷の病態生理学に貴重な洞察を提供するために、虚血皮膚創傷治癒モデルとしてラットを利用することである。 

ビデオ: Demonstration of the Rat Ischemic Skin Wound Model

Accurate and Simple Measurement of the Pro-inflammatory Cytokine IL-1&beta; using a Whole Blood Stimulation Assay

Inflammatory processes resulting from the secretion of soluble mediators by immune cells, lead to various manifestations in skin, joints and other tissues as well as altered cytokine homeostasis. The innate immune system plays a crucial role in recognizing pathogens and other endogenous danger stimuli. One of the major cytokines released by innate immune cells is Interleukin (IL)-1. Therefore, we utilize a whole blood stimulation assay in order to measure the secretion of inflammatory cytokines and specifically of the pro-inflammatory cytokine IL-1&beta; 1, 2, 3.
Patients with genetic dysfunctions of the innate immune system causing autoinflammatory syndromes show an exaggerated release of mature IL-1&beta; upon stimulation with LPS alone. In order to evaluate the innate immune component of patients who present with inflammatory-associated pathologies, we use a specific immunoassay to detect cellular immune responses to pathogen-associated molecular patterns (PAMPs), such as the gram-negative bacterial endotoxin, lipopolysaccharide (LPS). These PAMPs are recognized by pathogen recognition receptors (PRRs), which are found on the cells of the innate immune system 4, 5, 6, 7. A primary signal, LPS, in conjunction with a secondary signal, ATP, is necessary for the activation of the inflammasome, a multiprotein complex that processes pro-IL-1&beta; to its mature, bioactive form 4, 5, 6, 8, 9, 10.
The whole blood assay requires minimal sample manipulation to assess cytokine production when compared to other methods that require labor intensive isolation and culturing of specific cell populations. This method differs from other whole blood stimulation assays; rather than diluting samples with a ratio of RPMI media, we perform a white blood cell count directly from diluted whole blood and therefore, stimulate a known number of white blood cells in culture 2. The results of this particular whole blood assay demonstrate a novel technique useful in elucidating patient cohorts presenting with autoinflammatory pathophysiologies.

Accurate and Simple Measurement of the Pro-inflammatory Cytokine IL-1&amp;#946; using a Whole Blood Stimulation Assay

We describe a simple immunoassay to measure the production of pro-inflammatory cytokines, such as IL-1 beta production, in patients presenting with autoinflammatory phenotypes. By activating cells in whole blood cultures with pathogen-associated molecular patterns, specifically with lipopolysaccharide, cytokine secretion can be conveniently evaluated in whole blood supernatants.

全血刺激のためのアッセイを用いて炎症性サイトカインIL -1βの正確かつ簡単に測定

Inflammatory processes resulting from the secretion of soluble mediators by immune cells, lead to various manifestations in skin, joints and other tissues ...

医学

A Simplified Technique for Producing an Ischemic Wound Model

One major obstacle in current diabetic wound research is a lack of an ischemic wound model that can be safely used in diabetic animals. Drugs that work well in non-ischemic wounds may not work in human diabetic wounds because vasculopathy is one major factor that hinders healing of these wounds. We published an article in 2007 describing a rabbit ear ischemic wound model created by a minimally invasive surgical technique. Since then, we have further simplified the procedure for easier operation. On one ear, three small skin incisions were made on the vascular pedicles, 1-2 cm from the ear base. The central artery was ligated and cut along with the nerve. The whole cranial bundle was cut and ligated, leaving only the caudal branch intact. A circumferential subcutaneous tunnel was made through the incisions, to cut subcutaneous tissues, muscles, nerves, and small vessels. The other ear was used as a non-ischemic control. Four wounds were made on the ventral side of each ear. This technique produces 4 ischemic wounds and 4 non-ischemic wounds in one animal for paired comparisons. After surgery, the ischemic ear was cool and cyanotic, and showed reduced movement and a lack of pulse in the ear artery. Skin temperature of the ischemic ear was 1-10 &deg;C lower than that on the normal ear and this difference was maintained for more than one month. Ear tissue high-energy phosphate contents were lower in the ischemic ear than the control ear. Wound healing times were longer in the ischemic ear than in the non-ischemic ear when the same treatment was used. The technique has now been used on more than 80 rabbits in which 23 were diabetic (diabetes time ranging from 2 weeks to 2 years). No single rabbit has developed any surgical complications such as bleeding, infection, or rupture in the skin incisions. The model has many advantages, such as little skin disruption, longer ischemic time, and higher success rate, when compared to many other models. It can be safely used in animals with reduced resistance, and can also be modified to meet different testing requirements.

We have developed a minimally invasive technique to create a rabbit ischemic ear wound model by dividing the central artery and nerve and the cranial neurovascular bundle. A subcutaneous tunnel then cuts all subcutaneous tissues. This procedure causes minimal skin disruption and can be safely used in diabetic animals.

虚血性創傷モデルを作製するための簡略化手法

One major obstacle in current diabetic wound research is a lack of an ischemic wound model that can be safely used in diabetic animals. Drugs that work ...

Endothelin-1 Induced Middle Cerebral Artery Occlusion Model for Ischemic Stroke with Laser Doppler Flowmetry Guidance in Rat

Stroke is the number one cause of disability and third leading cause of death in the world, costing an estimated $70 billion in the United States in 20091, 2. Several models of cerebral ischemia have been developed to mimic the human condition of stroke. It has been suggested that up to 80% of all strokes result from ischemic damage in the middle cerebral artery (MCA) area3. In the early 1990s, endothelin-1 (ET-1) 4 was used to induce ischemia by applying it directly adjacent to the surface of the MCA after craniotomy. Later, this model was modified 5 by using a stereotaxic injection of ET-1 adjacent to the MCA to produce focal cerebral ischemia. The main advantages of this model include the ability to perform the procedure quickly, the ability to control artery constriction by altering the dose of ET-1 delivered, no need to manipulate the extracranial vessels supplying blood to the brain as well as gradual reperfusion rates that more closely mimics the reperfusion in humans5-7. On the other hand, the ET-1 model has disadvantages that include the need for a craniotomy, as well as higher variability in stroke volume8. This variability can be reduced with the use of laser Doppler flowmetry (LDF) to verify cerebral ischemia during ET-1 infusion. Factors that affect stroke variability include precision of infusion and the batch of the ET-1 used6. Another important consideration is that although reperfusion is a common occurrence in human stroke, the duration of occlusion for ET-1 induced MCAO may not closely mimic that of human stroke where many patients have partial reperfusion over a period of hours to days following occlusion9, 10. This protocol will describe in detail the ET-1 induced MCAO model for ischemic stroke in rats. It will also draw attention to special considerations and potential drawbacks throughout the procedure.

Several animal models of cerebral ischemia have been developed to simulate the human condition of stroke. This protocol describes the endothelin-1 (ET-1) induced middle cerebral artery occlusion (MCAO) model for ischemic stroke in rats. In addition, important considerations, advantages, and shortcomings of this model are discussed.

ラットにおけるレーザードップラー血流計の指針と虚血性脳卒中のエンドセリン-1誘発性中大脳動脈閉塞モデル

Stroke is the number one cause of disability and third leading cause of death in the world, costing an estimated $70 billion in the United States in ...

Angiogenesis in the Ischemic Rat Lung

The adult lung is perfused by both the systemic bronchial artery and the entire venous return flowing through the pulmonary arteries. In most lung pathologies, it is the smaller systemic vasculature that responds to a need for enhanced lung perfusion and shows robust neovascularization. Pulmonary vascular ischemia induced by pulmonary artery obstruction has been shown to result in rapid systemic arterial angiogenesis in man as well as in several animal models. Although the histologic assessment of the time course of bronchial artery proliferation in rats was carefully described by Weibel 1, mechanisms responsible for this organized growth of new vessels are not clear. We provide surgical details of inducing left pulmonary artery ischemia in the rat that leads to bronchial neovascularization. Quantification of the extent of angiogenesis presents an additional challenge due to the presence of the two vascular beds within the lung. Methods to determine functional angiogenesis based on labeled microsphere injections are provided.

The lung is perfused by both the systemic bronchial artery and pulmonary arteries. In most lung pathologies, it is the smaller systemic vasculature that shows robust neovascularization. Cessation of pulmonary blood flow promotes brisk bronchial angiogenesis. We provide surgical details of inducing left pulmonary artery ischemia that promotes bronchial neovascularization.

虚血ラットの肺の血管新生

The  adult lung is perfused by both the systemic bronchial artery and the entire  venous return flowing through the pulmonary arteries. In most lung ...

Murine Model of Wound Healing

Wound healing and repair are the most complex biological processes that occur in human life. After injury, multiple biological pathways become activated. Impaired wound healing, which occurs in diabetic patients for example, can lead to severe unfavorable outcomes such as amputation. There is, therefore, an increasing impetus to develop novel agents that promote wound repair. The testing of these has been limited to large animal models such as swine, which are often impractical. Mice represent the ideal preclinical model, as they are economical and amenable to genetic manipulation, which allows for mechanistic investigation. However, wound healing in a mouse is fundamentally different to that of humans as it primarily occurs via contraction. Our murine model overcomes this by incorporating a splint around the wound. By splinting the wound, the repair process is then dependent on epithelialization, cellular proliferation and angiogenesis, which closely mirror the biological processes of human wound healing. Whilst requiring consistency and care, this murine model does not involve complicated surgical techniques and allows for the robust testing of promising agents that may, for example, promote angiogenesis or inhibit inflammation. Furthermore, each mouse acts as its own control as two wounds are prepared, enabling the application of both the test compound and the vehicle control on the same animal. In conclusion, we demonstrate a practical, easy-to-learn, and robust model of wound healing, which is comparable to that of humans.

A murine model of cutaneous wound healing that can be used to assess therapeutic compounds in physiological and pathophysiological settings.

創傷治癒のマウスモデル

Wound healing and repair are the most complex biological processes that occur in human life. After injury, multiple biological pathways become activated. ...

Ischemic Tissue Injury in the Dorsal Skinfold Chamber of the Mouse: A Skin Flap Model to Investigate Acute Persistent Ischemia

Despite profound expertise and advanced surgical techniques, ischemia-induced complications ranging from wound breakdown to extensive tissue necrosis are still occurring, particularly in reconstructive flap surgery. Multiple experimental flap models have been developed to analyze underlying causes and mechanisms and to investigate treatment strategies to prevent ischemic complications. The limiting factor of most models is the lacking possibility to directly and repetitively visualize microvascular architecture and hemodynamics. The goal of the protocol was to present a well-established mouse model affiliating these before mentioned lacking elements. Harder et al. have developed a model of a musculocutaneous flap with a random perfusion pattern that undergoes acute persistent ischemia and results in ~50% necrosis after 10 days if kept untreated. With the aid of intravital epi-fluorescence microscopy, this chamber model allows repetitive visualization of morphology and hemodynamics in different regions of interest over time. Associated processes such as apoptosis, inflammation, microvascular leakage and angiogenesis can be investigated and correlated to immunohistochemical and molecular protein assays. To date, the model has proven feasibility and reproducibility in several published experimental studies investigating the effect of pre-, peri- and postconditioning of ischemically challenged tissue.

The window of the murine dorsal skinfold chamber presented visualizes a zone of acute persistent ischemia of a musculocutaneous flap. Intravital epi-fluorescence microscopy permits for direct and repetitive assessment of the microvasculature and quantification of hemodynamics. Morphologic and hemodynamic results can further be correlated with histological and molecular analyses.

マウスの背部皮脂商工における虚血性組織損傷：皮膚弁モデル急性永続虚血を調査する

Despite profound expertise and advanced surgical techniques, ischemia-induced complications ranging from wound breakdown to extensive tissue necrosis are ...

A Mouse Fetal Skin Model of Scarless Wound Repair

Early in utero, but not in postnatal life, cutaneous wounds undergo regeneration and heal without formation of a scar. Scarless fetal wound healing occurs across species but is age dependent. The transition from a scarless to scarring phenotype occurs in the third trimester of pregnancy in humans and around embryonic day 18 (E18) in mice. However, this varies with the size of the wound with larger defects generating a scar at an earlier gestational age. The emergence of lineage tracing and other genetic tools in the mouse has opened promising new avenues for investigation of fetal scarless wound healing. However, given the inherently high rates of morbidity and premature uterine contraction associated with fetal surgery, investigations of fetal scarless wound healing in vivo require a precise and reproducible surgical model. Here we detail a reliable model of fetal scarless wound healing in the dorsum of E16.5 (scarless) and E18.5 (scarring) mouse embryos.

During mammalian development, early gestational skin wounds heal without a scar. Here we detail a reliable and reproducible model of fetal scarless wound healing in the cutaneous dorsum of E16.5 (scarless) and E18.5 (scarring) mouse embryos.

瘢痕創傷修復のマウス胎児皮膚モデル

Early in utero, but not in postnatal life, cutaneous wounds undergo regeneration and heal without formation of a scar. Scarless fetal wound healing occurs ...

Rat Model of Photochemically-Induced Posterior Ischemic Optic Neuropathy

Posterior Ischemic optic neuropathy (PION) is a sight-devastating disease in clinical practice. However, its pathogenesis and natural history have&#160;remained poorly understood. Recently, we developed a reliable, reproducible animal model of PION and tested the treatment effect of some neurotrophic factors in this model1. The purpose of this video is to demonstrate our photochemically induced model of posterior ischemic optic neuropathy, and to evaluate its effects with retrograde labeling of retinal ganglion cells. Following surgical exposure of the posterior optic nerve, a photosensitizing dye, erythrosin B, is intravenously injected and a laser beam is focused onto the optic nerve surface. Photochemical interaction of erythrosin B and the laser during irradiation damages the vascular endothelium, prompting microvascular occlusion mediated by platelet thrombosis and edematous compression. The resulting ischemic injury yields a gradual but pronounced retinal ganglion cell dieback, owing to a loss of axonal input - a remote, injury-induced and clinically relevant outcome. Thus, this model provides a novel platform to study the pathophysiologic course of PION,&#160;and can be further optimized for testing therapeutic approaches for optic neuropathies as well as other CNS ischemic diseases.

The goal of this protocol is to photochemically induce ischemic injury to the posterior optic nerve in rat. This model is critical to studies of the pathophysiology of posterior ischemic optic neuropathy, and therapeutic approaches for this and other optic neuropathies, as well as of other CNS ischemic diseases.

光化学誘起後方虚血性視神経障害のラットモデル

Posterior Ischemic optic neuropathy (PION) is a sight-devastating disease in clinical practice. However, its pathogenesis and natural history ...

A Piglet Model of Neonatal Hypoxic-Ischemic Encephalopathy

Birth asphyxia, which causes hypoxic-ischemic encephalopathy (HIE), accounts for 0.66 million deaths worldwide each year, about a quarter of the world&#8217;s 2.9 million neonatal deaths. Animal models of HIE have contributed to the understanding of the pathophysiology in HIE, and have highlighted the dynamic process that occur in brain injury due to perinatal asphyxia. Thus, animal studies have suggested a time-window for post-insult treatment strategies. Hypothermia has been tested as a treatment for HIE in pdiglet models and subsequently proven effective in clinical trials. Variations of the model have been applied in the study of adjunctive neuroprotective methods and piglet studies of xenon and melatonin have led to clinical phase I and II trials1,2. The piglet HIE model is further used for neonatal resuscitation- and hemodynamic studies as well as in investigations of cerebral hypoxia on a cellular level. However, it is a technically challenging model and variations in the protocol may result in either too mild or too severe brain injury. In this article, we demonstrate the technical procedures necessary for establishing a stable piglet model of neonatal HIE. First, the newborn piglet (&#60; 24 hr old, median weight 1500 g) is anesthetized, intubated, and monitored in a setup comparable to that found in a neonatal intensive care unit. Global hypoxia-ischemia is induced by lowering the inspiratory oxygen fraction to achieve global hypoxia, ischemia through hypotension and a flat trace amplitude integrated EEG (aEEG) indicative of cerebral hypoxia. Survival is promoted by adjusting oxygenation according to the aEEG response and blood pressure. Brain injury is quantified by histopathology and magnetic resonance imaging after 72 hr.

Hypoxic-ischemic encephalopathy following perinatal asphyxia can be studied using animal models. We demonstrate the procedures necessary for establishing a piglet model of neonatal hypoxic-ischemic encephalopathy.

新生児低酸素性虚血性脳症の子豚モデル

Birth asphyxia, which causes hypoxic-ischemic encephalopathy (HIE), accounts for 0.66 million deaths worldwide each year, about a quarter of the ...

The Rodent Model of Nonarteritic Anterior Ischemic Optic Neuropathy (rNAION)

Nonarteritic anterior ischemic optic neuropathy (NAION) is a focal ischemic lesion of the optic nerve that affects 1/700 individuals throughout their lifetime. NAION results in optic nerve edema, selective loss of the retinal ganglion cell neurons (RGCs) and atrophy of the optic nerve. A rodent model of NAION that expresses most NAION features and sequelae has been developed, which is applicable to both rats and mice. This model utilizes a focal laser application of 532 nm wavelength to illuminate a photoactive dye, Rose Bengal (RB), to cause capillary damage and leakage at the targeted anterior optic nerve (the laminar region). After rNAION induction, there is an early optic nerve ischemia, optic nerve edema, and intraneural inflammation, followed by selective RGC and axonal loss. Since the optic nerve is a CNS white matter tract, the rNAION model is applicable to mechanistic studies of selective white matter ischemia, as well as neuroprotective analyses and short and long-term mechanisms of glial and neuronal response to ischemia.

The Rodent Model of Nonarteritic Anterior Ischemic Optic Neuropathy rNAION

The following report describes how to replicate the rodent model of nonarteritic anterior ischemic optic neuropathy (rNAION), using the appropriate dye, contact lens and laser parameters. We also reveal the appropriate steps for evaluating the rNAION lesion in vivo.

Nonarteritic前部虚血性視神経症の齧歯類モデル（rNAION）

Nonarteritic anterior ischemic optic neuropathy (NAION) is a focal ischemic lesion of the optic nerve that affects 1/700 individuals throughout their ...