この作品は、NIH R01 HL093183、HL088434、HL071763、HL080498、HL083156、そしてP20HL100396（RJH）で、Caerusネットワーク（RJH）を通じてLeducq財団によってサポートされています。 DLは、ドイツ研究財団によってサポートされていました。我々はこのプロジェクトでは彼女の技術支援のためにローレンLeonardsonに感謝します。

1。心筋梗塞の作成<ol><li>アニマルバイオテクノロジー産業の女性ヨークシャーブタは（20キロ）（ドイルズタウン、PA）Telazol（tiletamine / zolazepam）（8.0 mg / kg体重）およびブプレノルフィン（0.6 mg）を使用してプリ薬用です。 </li><li>動物は挿管し、100％酸素で換気されています。全身麻酔は、プロシージャ全体でプロポフォール（5-8mg/kg/hr）で維持されます。 </li><li>大腿サイトはポビドンヨードに続く70％イソプロピルアルコールで用意されています。経皮的穿刺は、動脈へのアクセスとシースの配置のために用意されています。シース挿入後、ヘパリンの3000から5000 IUは、250〜300秒の活性化凝固時間を維持するためにIVを投与する。 </li><li>アミオダロン（75 mg）のボーラス投与は筋肉内に投与される。アミオダロン（75 mg）を、アトロピン（2mg）を、および酢酸カリウム（20 MEQ）で1000 ml生理食塩水バッグの静脈内注入が開始され、プロシージャ全体で150ミリリットル/時の速度を使用して維持されます。 </li><li> 7神父ホッケースティックのカテーテルは、（コーディス、マイアミ、フロリダ州）左冠状動脈に高度であり、冠動脈造影が行われます。 0.014インチのガイドワイヤー（アボット、パーク、イリノイ州）は冠状動脈（LAD）を左前に進んでいる。 8 mmの長、4.0 mmのVOYAGERオーバーザワイヤーバルーン（アボット、パーク、イリノイ州）が進んでおり、位置付け近位LADで、冠動脈バルーンは3気圧に膨張される。 LADの閉塞が血管造影によって確認されます。 </li><li>連続的な心電図と血行動態モニタリングで、閉塞は90分間維持されます。悪性心室性不整脈は、時折投与される場合200 Jから300 Jまで直流ショックの即時配信をで、最初の40分以内に発生する可能性があります。通常、不整脈の二つのピークの時間枠、1つの2〜10分、バルーン拡張後に25〜35分の間に他のがあります。フレームが示された時間内に心室細動の入力の95％以上の発生率があるので、入念に監視し、即座に除細動は、成功した心筋梗塞の創造への鍵です。迅速かつ適切な除細動が行われている場合、生存率は非常に高いです。血圧が80 mmHgの下に低下した場合、容積負荷は、急速な静脈内生理食塩水注入で実現されます。 </li><li>バルーンが収縮され、すべてのデバイスが削除されます。 </li><li>シースは、麻酔が源泉徴収され、大腿動脈から削除され、そして動物は、リカバリ中に監視されます。アミオダロン、アトロピンと酢酸カリウムの点滴と静脈内生理食塩水を50ml /時間で減少し、一晩与えられます。 NitroglycerlineとFuroscemideの筋肉内注射が投与される。その後、動物を回復されており、慎重な心電図モニタリングは少なくとも3時間続ける。 </li><li>アスピリン（200mg）をPOは5日間が与えられます。 </li></ol>心筋梗塞の作成手順の実行中および終了後は一般的な合併症は、冠動脈解離、穿刺部位の血腫、不整脈、および心不全です。技術を洗練し、徹底的な監視は、これらの合併症を予防して管理するために必要です。 2。ウイルスのインジェクション<ol><li>上記の心筋梗塞の作成steps1 - 3で示されているように動物が用意されています。ヘパリンの同一投与量は、ウイルスの注入中に血栓症を防ぐために与えられるべきである。 </li><li> 5Frホッケースティックのカテーテル（コーディス、マイアミ、FL）は左冠状動脈に進められます。血管造影後、0.014インチのガイドワイヤーの二人は、LADおよびカテーテルの位置を修正するために、左冠動脈回旋枝（LCX）への1つへの1つ進んでいます。 </li><li>動脈血（-20 ml）をカテーテルから引き出される。注入時の虚血を防ぐために、ハーフ希釈した血液は、ソリューションに使用されます。ウイルスは、10 mlの血液と混合し、生理食塩水で20mlに希釈される。残りの10 mlを、20 mlに希釈し、カテーテルの内腔内残存ウイルスを提供するために使用されます。 </li><li>静脈注射用ニトログリセリン（20μgを/分）は耳静脈を介して開始されます。 </li><li>カテーテルの先端は、近位左主管に配置されます。カテーテルを介して圧力は、カテーテルが楔されていないことを保証するために監視されます。 </li><li>ウイルス混合血液（15ml）を、左冠状動脈に注入ポンプを使用して、15分にわたってカテーテルを介して注入される。 15分後、注射器は、フラッシュに交換され、そして注入は10分間継続しています。 </li><li>ワイヤとカテーテルが右冠動脈に左冠状動脈と同じ方法で固定されている。 </li><li>ウイルスと混合血液の残りの5mlをフラッシュの10分に続いて5分間右冠動脈に注入されます。 </li></ol> 3。代表的なケース図1は、心筋梗塞の作成後1ヶ月で左室造影を示しています。左心室壁運動が有意の駆出率（EF）と低下している31％であった。図2は、1ヶ月での左心室の断面を示しています。ベースから前壁の頂部に大きな傷が見られます。このモデルでは、心筋梗塞後1ヶ月での平均EFは36％であり、心筋梗塞のない同じサイズの動物のそれらの平均はそれぞれ、68％、42ミリリットルであるのに対し、平均的な拡張末期容積は、83ミリリットルです。図3は、左心室の境界ゾーンの領域におけるβ- galの発現アデノ随伴ウイルス（rAAV1 CMVラック- Zの2 × 10 12 DNアーゼresitant粒子）の注入後1ヶ月を示しています。増強された発現は、ウイルスの注入領域で発見された。 <img src="/files/ftp_upload/2778/2778fig1.jpg" alt="図1" /> 図1。駆出率は心筋梗塞の作成 ​​後1ヶ月左室造影から測定。左心室は92.5ミリリットルに瞳孔が開いていると前壁の動きが大幅に低下します。 <img src="/files/ftp_upload/2778/2778fig2.jpg" alt="図2" /> 瘢痕領域の輪郭を描くためにトリフェニルテトラゾリウムクロライドで染色した心筋梗塞の作成 ​​後1ヶ月での左心室の2断面、 図 。ベースから前壁の頂部に大きな傷が見られます。 <img src="/files/ftp_upload/2778/2778fig3.jpg" alt="図3" /> 図3。アデノ随伴ウイルス（rAAV1 CMVラック- Zの2 × 10 12 DNアーゼ性粒子）により誘導されるβ- gal発現。増強された発現は梗塞及び正常組織、冠動脈内のウイルス注入後1ヶ月の間の国境地帯の領域で発見されています。右下の写真は、生理食塩水は、同じプロトコルを使用して冠状動脈に注入したネガティブコントロールを示しています。

<ol>
	<li>Perez de Prado, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Closed-chest+experimental+porcine+model+of+acute+myocardial+infarction-reperfusion.">Closed-chest experimental porcine model of acute myocardial infarction-reperfusion.</a> J Pharmacol Toxicol Methods. 60, 301-306 (2009).</li><li>Angeli, F. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Left+ventricular+remodeling+after+myocardial+infarction:+characterization+of+a+swine+model+on+beta-blocker+therapy.">Left ventricular remodeling after myocardial infarction: characterization of a swine model on beta-blocker therapy.</a> Comp Med. 59, 272-279 (2009).</li><li>Urdaneta, F., Lobato, E. B., Kirby, D. S. &. a. m. p. ;. a. m. p., Sidi, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Treating+myocardial+stunning+randomly,+with+either+propofol+or+isoflurane+following+transient+coronary+occlusion+and+reperfusion+in+pigs.">Treating myocardial stunning randomly, with either propofol or isoflurane following transient coronary occlusion and reperfusion in pigs.</a> Ann Card Anaesth. 12, 113-121 (2009).</li><li>Jaski, B. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Calcium+upregulation+by+percutaneous+administration+of+gene+therapy+in+cardiac+disease+(CUPID+Trial),+a+first-in-human+phase+1/2+clinical+trial.">Calcium upregulation by percutaneous administration of gene therapy in cardiac disease (CUPID Trial), a first-in-human phase 1/2 clinical trial.</a> J Card Fail. 15, 171-181 (2009).</li></ol>

すべての著者は原稿に関連して利害の対立としてもたらす可能性は商業的関連を持っていません。

心不全の様々な原因の中で、虚血は、これまでで最も頻繁に病因にすることです。この深刻な病気を克服するために、様々な治療アプローチが開発されている。これらの有望な治療法が診療所に適用することができるされる前に治療法の慎重な評価が必要です。したがって、患者の臨床状態を表す虚血性心不全モデルの確立は、心不全治療薬の開発にとって非常に重要です。私たちの豚心筋?...

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		<div>
		<table border="1">
		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>Telazol</td>
<td>&nbsp;</td>
<td>Fort Dodge Animal Health</td>
<td>0856-9050-49</td>
<td>&nbsp;</td>
<tr>
<td>Propofol</td>
<td>&nbsp;</td>
<td>APP Pharmaceuticals</td>
<td>63323-270-20</td>
<td>&nbsp;</td>
<tr>
<td>Heparin</td>
<td>&nbsp;</td>
<td>APP Pharmaceuticals</td>
<td>63323-540-11</td>
<td>&nbsp;</td>
<tr>
<td>Amiodarone</td>
<td>&nbsp;</td>
<td>APP Pharmaceuticals</td>
<td>63323-616-03</td>
<td>&nbsp;</td>
<tr>
<td>Atropin Sulfate</td>
<td>&nbsp;</td>
<td>Med-Pharmex</td>
<td>54925-063-10</td>
<td>&nbsp;</td>
<tr>
<td>Potassium acetate</td>
<td>&nbsp;</td>
<td>American Regent, Inc.</td>
<td>2053-25</td>
<td>&nbsp;</td>
<tr>
<td>Nitroglycerine</td>
<td>&nbsp;</td>
<td>American Regent, Inc.</td>
<td>4810-25 </td>
<td>&nbsp;</td>
<tr>
<td>Salix (Furosemide)</td>
<td>&nbsp;</td>
<td>Intervet Inc.</td>
<td>710461</td>
<td>&nbsp;</td>
<tr>
<td>5 Fr Hockey stick catheter</td>
<td>&nbsp;</td>
<td>Cordis</td>
<td>556-278-0L</td>
<td>&nbsp;</td>
<tr>
<td>7 Fr Hockey stick catheter</td>
<td>&nbsp;</td>
<td>Cordis</td>
<td>778-278-00</td>
<td>&nbsp;</td>
<tr>
<td>Coronary wire</td>
<td>&nbsp;</td>
<td>Abbott Laboratories</td>
<td>1044588</td>
<td>&nbsp;</td>
<tr>
<td>Coronary balloon</td>
<td>&nbsp;</td>
<td>Abbott Laboratories</td>
<td>1009447-08</td>
<td>&nbsp;</td>
<tr>
<td>8Fr sheath</td>
<td>&nbsp;</td>
<td>TERUMO</td>
<td>RSS801</td>
<td>&nbsp;</td>		</tbody>
		</table>
		</div>

gene transfer for ischemic heart failure in a preclinical model

様々な新技術は、心不全患者のために開発されている。十分に確立された前臨床評価は、その有効性と安全性を判断する必要があります。 ウイルスベクターを用いた遺伝子治療は、心臓疾患を治療するための最も有望なアプローチの一つです。キャリアの遺伝子を変えることによりさまざまな遺伝子のウイルス送達は、計り知れない治療能力を持っています。 このビデオでは、遺伝子導入に続いて心不全の作成の動物モデルの完全なプロセスは、豚モデルを使用して表示されます。最初に、心筋梗塞は冠動脈の降順近左前を閉塞することによって作成されます。慢性心不全における心臓リモデリングの結果。我々のモデルに固有の、真に肯定的な成果のための積極的な治療を必要とする重症心不全患者を反映してかなり大きな傷跡です。瘢痕組織の心筋梗塞の創造と発展を経て、ウイルスの冠動脈内注入は、同時ニトログリセリン注入で示されています。私たちの注入法は、強化された遺伝子発現でシンプルかつ効率的な遺伝子導入を提供します。冠動脈内のウイルスの配信と心筋梗塞豚モデルのこの組み合わせは、個々の遺伝子の効果をテストするだけでなく、支援の一貫性と再現性のある方法論、であることが証明さだけでなく、治療薬候補として多くの遺伝子の有効性を比較しました。

Watch this Scientific Journal Video about Gene Transfer for Ischemic Heart Failure in a Preclinical Model at JoVE.com

Gene Transfer for Ischemic Heart Failure in a Preclinical Model

虚血性心不全の治療のための遺伝子導入法は、心筋梗塞の豚モデルを使用して記述されます。私たちのシンプルで再現性のある方法は、私たちは容易に非常にシンプルかつ再現可能な方法で様々な遺伝子の転送の有効性を評価することができます。

前臨床モデルにおける虚血性心不全のための遺伝子導入

Various emerging technologies are being developed for patients with heart failure. Well-established preclinical evaluations are necessary to determine ...

gene-transfer-for-ischemic-heart-failure-in-a-preclinical-model

Research

JoVE Journal

Medicine

12.8K ビュー.  Mount Sinai School of Medicine. 虚血性心不全の治療のための遺伝子導入法は、心筋梗塞の豚モデルを使用して記述されます。私たちのシンプルで再現性のある方法は、私たちは容易に非常にシンプルかつ再現可能な方法で様々な遺伝子の転送の有効性を評価することができます。

ビデオ: Gene Transfer for Ischemic Heart Failure in a Preclinical Model

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 ...

医学

Permanent Ligation of the Left Anterior Descending Coronary Artery in Mice: A Model of Post-myocardial Infarction Remodelling and Heart Failure

Heart failure is a syndrome in which the heart fails to pump blood at a rate commensurate with cellular oxygen requirements at rest or during stress. It is characterized by fluid retention, shortness of breath, and fatigue, in particular on exertion. Heart failure is a growing public health problem, the leading cause of hospitalization, and a major cause of mortality. Ischemic heart disease is the main cause of heart failure.
Ventricular remodelling refers to changes in structure, size, and shape of the left ventricle. This architectural remodelling of the left ventricle is induced by injury (e.g., myocardial infarction), by pressure overload (e.g., systemic arterial hypertension or aortic stenosis), or by volume overload. Since ventricular remodelling affects wall stress, it has a profound impact on cardiac function and on the development of heart failure. A model of permanent ligation of the left anterior descending coronary artery in mice is used to investigate ventricular remodelling and cardiac function post-myocardial infarction. This model is fundamentally different in terms of objectives and pathophysiological relevance compared to the model of transient ligation of the left anterior descending coronary artery. In this latter model of ischemia/reperfusion injury, the initial extent of the infarct may be modulated by factors that affect myocardial salvage following reperfusion. In contrast, the infarct area at 24 hr after permanent ligation of the left anterior descending coronary artery is fixed. Cardiac function in this model will be affected by 1) the process of infarct expansion, infarct healing, and scar formation; and 2) the concomitant development of left ventricular dilatation, cardiac hypertrophy, and ventricular remodelling.
Besides the model of permanent ligation of the left anterior descending coronary artery, the technique of invasive hemodynamic measurements in mice is presented in detail.

Heart failure is the leading cause of hospitalization and a major cause of mortality. A model of permanent ligation of the left anterior descending coronary artery in mice is applied to investigate ventricular remodelling and cardiac dysfunction post-myocardial infarction. The technique of invasive hemodynamic measurements in mice is presented.

マウスの左前下冠動脈の動脈の常設ライゲーション：心筋梗塞後のリモデリングおよび心不全のモデル

Heart failure is a syndrome in which the heart fails to pump blood at a rate commensurate with cellular oxygen requirements at rest or during stress. It ...

In Vivo Gene Transfer to the Rabbit Common Carotid Artery Endothelium

The goal of this method is to introduce a transgene into the endothelium of isolated segments of both rabbit common carotid arteries. The method achieves focal endothelial-selective transgenesis, thereby allowing an investigator to determine the biological roles of endothelial-expressed transgenes and to quantify the in vivo transcriptional activity of DNA sequences in large artery endothelial cells. The method uses surgical isolation of rabbit common carotid arteries and an arteriotomy to deliver a transgene-expressing viral vector into the arterial lumen. A short incubation period of the vector in the lumen, with subsequent aspiration of the lumen contents, is sufficient to achieve efficient and durable expression of the transgene in the endothelium, with no detectable transduction or expression outside of the isolated arterial segment. The method allows assessment of the biological activities of transgene products both in normal arteries and in models of human vascular disease, while avoiding systemic effects that could be caused either by targeting gene delivery to other sites (e.g. the liver) or by the alternative approach of delivering genetic constructs to the endothelium by germ line transgenesis. Application of the method is limited by the need for a skilled surgeon and anesthetist, a well-equipped operating room, the costs of purchasing and housing rabbits, and the need for expertise in gene-transfer vector construction and use. Results obtained with this method include: transgene-related alterations in arterial structure, cellularity, extracellular matrix, or vasomotor function; increases or reductions in arterial inflammation; alterations in vascular cell apoptosis; and progression, retardation, or regression of diseases such as intimal hyperplasia or atherosclerosis. The method also allows measurement of the ability of native and synthetic DNA regulatory sequences to alter transgene expression in endothelial cells, providing results that include: levels of transgene mRNA, levels of transgene protein, and levels of transgene enzymatic activity.

In Vivo Gene Transfer to the Rabbit Common Carotid Artery Endothelium

This method is to introduce a transgene into the endothelium of rabbit carotid arteries. Introduction of the transgene allows the assessment of the biological role of the transgene product either in normal arteries or disease models. The method is also useful for measuring activity of DNA regulatory sequences.

生体内でウサギの一般的な頚動脈内皮細胞への遺伝子導入

The goal of this method is to introduce a transgene into the endothelium of isolated segments of both rabbit common carotid arteries. The method achieves ...

Tachycardia-Induced Cardiomyopathy As a Chronic Heart Failure Model in Swine

A stable and reliable model of chronic heart failure is required for many experiments to understand hemodynamics or to test effects of new treatment methods. Here, we present such a model by tachycardia-induced cardiomyopathy, which can be produced by rapid cardiac pacing in swine.
A single pacing lead is introduced transvenously into fully anaesthetized healthy swine, to the apex of the right ventricle, and fixated. Its other end is then tunneled dorsally to the paravertebral region. There, it is connected to an in-house modified heart pacemaker unit that is then implanted in a subcutaneous pocket. 
After 4 - 8 weeks of rapid ventricular pacing at rates of 200 - 240 beats/min, physical examination revealed signs of severe heart failure - tachypnea, spontaneous sinus tachycardia, and fatigue. Echocardiography and X-ray showed dilation of all heart chambers, effusions, and severe systolic dysfunction. These findings correspond well to decompensated dilated cardiomyopathy and are also preserved after the cessation of pacing.
This model of tachycardia-induced cardiomyopathy can be used for studying the pathophysiology of progressive chronic heart failure, especially hemodynamic changes caused by new treatment modalities like mechanical circulatory supports. This methodology is easy to perform and the results are robust and reproducible.

Here, we present a protocol to produce tachycardia-induced cardiomyopathy in swine. This model represents a potent way to study the hemodynamics of progressive chronic heart failure and the effects of applied treatment.

豚の慢性心不全モデルとして頻拍誘発性心筋

A stable and reliable model of chronic heart failure is required for many experiments to understand hemodynamics or to test effects of new treatment ...

Isolation of Atrial Cardiomyocytes from a Rat Model of Metabolic Syndrome-related Heart Failure with Preserved Ejection Fraction

In this article, we describe an optimized, Langendorff-based procedure for the isolation of single-cell atrial cardiomyocytes (ACMs) from a rat model of metabolic syndrome (MetS)-related heart failure with preserved ejection fraction (HFpEF). The prevalence of MetS-related HFpEF is rising, and atrial cardiomyopathies associated with atrial remodeling and atrial fibrillation are clinically highly relevant as atrial remodeling is an independent predictor of mortality. Studies with isolated single-cell cardiomyocytes are frequently used to corroborate and complement in vivo findings. Circulatory vessel rarefication and interstitial tissue fibrosis pose a potentially limiting factor for the successful single-cell isolation of ACMs from animal models of this disease.
We have addressed this issue by employing a device capable of manually regulating the intraluminal pressure of cardiac cavities during the isolation procedure, substantially increasing the yield of morphologically and functionally intact ACMs. The acquired cells can be used in a variety of different experiments, such as cell culture and functional Calcium imaging (i.e., excitation-contraction-coupling).
We provide the researcher with a step-by-step protocol, a list of optimized solutions, thorough instructions to prepare the necessary equipment, and a comprehensive troubleshooting guide. While the initial implementation of the procedure might be rather difficult, a successful adaptation will allow the reader to perform state-of-the-art ACM isolations in a rat model of MetS-related HFpEF for a broad spectrum of experiments.

Here, we describe an optimized, Langendorff-based procedure for the isolation of single-cell atrial cardiomyocytes from a rat model of metabolic syndrome-related heart failure with preserved ejection fraction. A manual regulation of intraluminal pressure of cardiac cavities is implemented to yield functionally intact myocytes suitable for excitation-contraction-coupling studies.

保存された駆出率とメタボリック症候群関連心不全ラットモデルから心房の心筋細胞の単離

In this article, we describe an optimized, Langendorff-based procedure for the isolation of single-cell atrial cardiomyocytes (ACMs) from a rat model of ...

Induction and Phenotyping of Acute Right Heart Failure in a Large Animal Model of Chronic Thromboembolic Pulmonary Hypertension

The development of acute right heart failure (ARHF) in the context of chronic pulmonary hypertension (PH) is associated with poor short-term outcomes. The morphological and functional phenotyping of the right ventricle is of particular importance in the context of hemodynamic compromise in patients with ARHF. Here, we describe a method to induce ARHF in a previously described large animal model of chronic PH, and to phenotype, dynamically, right ventricular function using the gold standard method (i.e., pressure-volume PV loops) and with a non-invasive clinically available method (i.e., echocardiography). Chronic PH is first induced in pigs by left pulmonary artery ligation and right lower lobe embolism with biological glue once a week for 5 weeks. After 16 weeks, ARHF is induced by successive volume loading using saline followed by iterative pulmonary embolism until the ratio of the systolic pulmonary pressure over systemic pressure reaches 0.9 or until the systolic systemic pressure decreases below 90 mmHg. Hemodynamics are restored with dobutamine infusion (from 2.5 &#181;g/kg/min to 7.5 &#181;g/kg/min). PV-loops and echocardiography are performed during each condition. Each condition requires around 40 minutes for induction, hemodynamic stabilization and data acquisition. Out of 9 animals, 2 died immediately after pulmonary embolism and 7 completed the protocol, which illustrates the learning curve of the model. The model induced a 3-fold increase in mean pulmonary artery pressure. The PV-loop analysis showed that ventriculo-arterial coupling was preserved after volume loading, decreased after acute pulmonary embolism and was restored with dobutamine. Echocardiographic acquisitions allowed to quantify right ventricular parameters of morphology and function with good quality. We identified right ventricular ischemic lesions in the model. The model can be used to compare different treatments or to validate non-invasive parameters of right ventricular morphology and function in the context of ARHF.

We present a protocol to induce and phenotype an acute right heart failure in a large animal model with chronic pulmonary hypertension. This model can be used to test therapeutic interventions, to develop right heart metrics&#160;or to improve the understanding of acute right heart failure pathophysiology.

慢性血栓塞栓性肺高血圧症の大型動物モデルにおける急性右心不全の誘導とフェノタイピング

The development of acute right heart failure (ARHF) in the context of chronic pulmonary hypertension (PH) is associated with poor short-term outcomes. The ...

Preclinical Model of Hind Limb Ischemia in Diabetic Rabbits

Peripheral vascular disease is a widespread clinical problem that affects millions of patients worldwide. A major consequence of peripheral vascular disease is the development of ischemia. In severe cases, patients can develop critical limb ischemia in which they experience constant pain and an increased risk of limb amputation. Current therapies for peripheral ischemia include bypass surgery or percutaneous interventions such as angioplasty with stenting or atherectomy to restore blood flow. However, these treatments often fail to the continued progression of vascular disease or restenosis or are contraindicated due to the overall poor health of the patient. A promising potential approach to treat peripheral ischemia involves the induction of therapeutic neovascularization to allow the patient to develop collateral vasculature. This newly formed network alleviates peripheral ischemia by restoring perfusion to the affected area. The most frequently employed preclinical model for peripheral ischemia utilizes the creation of hind limb ischemia in healthy rabbits through femoral artery ligation. In the past, however, there has been a strong disconnect between the success of preclinical studies and the failure of clinical trials regarding treatments for peripheral ischemia. Healthy animals typically have robust vascular regeneration in response to surgically induced ischemia, in contrast to the reduced vascularity and regeneration in patients with chronic peripheral ischemia. Here, we describe an optimized animal model for peripheral ischemia in rabbits that includes hyperlipidemia and diabetes. This model has reduced collateral formation and blood pressure recovery in comparison to a model with a higher cholesterol diet. Thus, the model may provide better correlation with human patients with compromised angiogenesis from the common co-morbidities that accompany peripheral vascular disease.

We describe a surgical procedure used to induce peripheral ischemia in rabbits with hyperlipidemia and diabetes. This surgery acts as a preclinical model for conditions experienced in peripheral artery disease in patients. Angiography is also described as a means to measure the extent of introduced ischemia and recovery of perfusion.

糖尿病性ウサギにおける後肢虚血の前臨床モデル

Peripheral vascular disease is a widespread clinical problem that affects millions of patients worldwide. A major consequence of peripheral vascular ...

Establishing a Swine Model of Post-myocardial Infarction Heart Failure for Stem Cell Treatment

Although advances have been achieved in the treatment of heart failure (HF) following myocardial infarction (MI), HF following MI remains one of the major causes of mortality and morbidity around the world. Cell-based therapies for cardiac repair and improvement of left ventricular function after MI have attracted considerable attention. Accordingly, the safety and efficacy of these cell transplantations should be tested in a preclinical large animal model of HF prior to clinical use. Pigs are widely used for cardiovascular disease research due to their similarity to humans in terms of heart size and coronary anatomy. Therefore, we sought to present an effective protocol for the establishment of a porcine chronic HF model using closed-chest coronary balloon occlusion of the left circumflex artery (LCX), followed by rapid ventricular pacing induced with pacemaker implantation. Eight weeks later, the stem cells were administered by intramyocardial injection in the peri-infarct area. Then the infarct size, cell survival, and left ventricular function (including echocardiography, hemodynamic parameters, and electrophysiology) were evaluated. This study helps establish a stable preclinical large animal HF model for stem cell treatment.

We sought to establish a swine model of heart failure induced by left circumflex artery blockage and rapid pacing to test the effect and safety of intramyocardial administration of stem cells for cell-based therapies.

幹細胞治療のための心筋梗塞後心不全の豚モデルの確立

Although advances have been achieved in the treatment of heart failure (HF) following myocardial infarction (MI), HF following MI remains one of the major ...

A Preclinical Model of Exertional Heat Stroke in Mice

Heat stroke is the most severe manifestation of heat-related illnesses. Classic heat stroke (CHS), also known as passive heat stroke, occurs at rest, whereas exertional heat stroke (EHS) occurs during physical activity. EHS differs from CHS in etiology, clinical presentation, and sequelae of multi-organ dysfunction. Until recently, only models of CHS have been well established. This protocol aims to provide guidelines for a refined preclinical mouse model of EHS that is free from major limiting factors such as the use of anesthesia, restraint, rectal probes, or electric shock. Male and female C57Bl/6 mice, instrumented with core temperature (Tc) telemetric probes were utilized in this model. For familiarization with the running mode, mice undergo 3 weeks of training using both voluntary and forced running wheels. Thereafter, mice run on a forced wheel inside a climatic chamber set at 37.5 &#176;C and 40%-50% relative humidity (RH) until displaying symptom limitation (e.g., loss of consciousness) at Tc of 42.1-42.5 &#176;C, although suitable results can be obtained at chamber temperatures between 34.5-39.5 &#176;C and humidity between 30%-90%. Depending on the desired severity, mice are removed from the chamber immediately for recovery in ambient temperature or remain in the heated chamber for a longer duration, inducing a more severe exposure and a higher incidence of mortality. Results are compared with sham-matched exercise controls (EXC) and/or na&#239;ve controls (NC). The model mirrors many of the pathophysiological outcomes observed in human EHS, including loss of consciousness, severe hyperthermia, multi-organ damage as well as inflammatory cytokine release, and acute phase responses of the immune system. This model is ideal for hypothesis-driven research to test preventative and therapeutic strategies that may delay the onset of EHS or reduce the multi-organ damage that characterizes this manifestation.

The protocol describes the development of a standardized, repeatable, preclinical model of exertional heat stroke (EHS) in mice free from adverse external stimuli such as electric shock. The model provides a platform for mechanistic, preventative, and therapeutic studies.

マウスにおける運動熱中症の前臨床モデル

Heat stroke is the most severe manifestation of heat-related illnesses. Classic heat stroke (CHS), also known as passive heat stroke, occurs at rest, ...

Large Animal Model for Evaluating the Efficacy of the Gene Therapy in Ischemic Heart

Coronary artery disease is one of the significant causes of mortality and morbidity worldwide. Despite the progression of current therapeutics, a considerable proportion of coronary artery disease patients remain symptomatic. Gene therapy-mediated therapeutic angiogenesis offers a novel therapeutic method for improving myocardial perfusion and relieving symptoms. Gene therapy with different angiogenic factors has been studied in few clinical trials. Due to the novelty of the method, the progress of myocardial gene therapy is a continuous path from bench to bedside. Therefore, large animal models are needed for evaluating the safety and efficacy. The more the large animal model identifies the original disease and the endpoints used in clinics, the more predictable outcomes are from clinical trials. Here, we introduce a large animal model for evaluating the efficacy of the gene therapy in the ischemic porcine heart. We use clinically relevant imaging methods such as ultrasound imaging and 15H2O-PET. For targeting the gene transfers into the desired area, electroanatomical mapping is used. The aim of this method is: (1) to mimic chronic coronary artery disease, (2) to induce therapeutic angiogenesis at hypoxic areas of the heart, and (3) to evaluate the safety and efficacy of the gene therapy by using relevant endpoints.

Myocardial gene therapy for ischemic heart disease holds great promise for future therapeutics. Here, we introduce a large animal model for evaluating the efficacy of gene therapy in the ischemic heart.