这一工作是由美国陆军格兰特（W81XWH-08-1-0701）和卡尔基金会医院的院士。

程序符合IACUC协议号13302完全兼容，并已通过动物资源的划分。 1.仪器准备<ol><li>高压灭菌之前用来在外科手术，以保证无菌的所有非一次性的仪器。 </li><li>消毒与用途之间玻璃珠灭菌多次使用在会话仪器。 </li></ol> 2.水凝胶制剂<ol><li>解冻板干细胞至少24小时之前创建的水凝胶结构的。 </li><li>制备贴剂前一天已在全部细节在以前的出版物8,18所述的外科手术用立体光刻设备（或类似的仪器）。 </li><li>为了创建该水凝胶，首先生成一个计算机辅助设计（CAD）模型将所需的修补和出口设计stereolit​​hographic格式（STL）。 </li><li>创建由dissolvin预凝胶溶液克聚（乙二醇）二甲基丙烯酸酯（PEGDMA）在无菌的1×PBS，得到20％w / v的溶液。同时，溶解的光引发剂1- [4-（2-羟基 - 乙氧基） - 苯基] -2-羟基-2-甲基-1-丙烷-1-酮在DMSO中。添加光引发剂到PEGDMA溶液在最终瓦特/ V的0.5％。直接交联之前，加入100 ml细胞所需的密度。注:细胞密度在2.0×10 5 -2.0×10 6个细胞/ ml，推荐。 </li><li>吸取在该SLA平台中心的预凝胶溶液成菜并运行该SLA软件与以前上传的STL格式设计。 </li><li>除去从装置的补丁和孵化的水凝胶的O / N的Dulbecco改良的Eagle培养基（DMEM），补充有5％胎牛血清（FBS），100U / ml青霉素，和100毫克/毫升链霉素，以允许细胞以调节到贴片内的环境。 </li></ol> 3.动物制备及口服intubati上<ol><li>评估之前，过程中的每个鼠标，以保证动物健康状况良好，因为从他们的美容习惯和相互作用决定与他们的笼子队友。注意:女8-10周龄的C57BL / 6J小鼠用于这个过程，但是，任何小鼠年龄或类型都可以使用。 </li><li>将动物进入麻醉室，并暴露于5％异氟烷以1升/分钟O 2的支持。监测麻醉通过脚趾捏反射的水平。 </li><li>一旦动物被正确麻醉，称重，并把他们放在插管立场。 </li><li>引向动物的胸腔的光源，并使用镊子缩回喉面和露出声带。小心地将声带之间的20G的静脉留置针，并顺利插入气管。通过胸腔的机械运动检测正确插入一旦导管被连接到小动物呼吸机。 </li><li>调整通风设置为th动物根据制造商的指导方针ë重量。 </li><li>一旦插管，放置在加热垫上仰卧位的动物，以防止体温过低。确保该加热垫不超过40℃。如果必要的话，将动物和加热垫之间的屏障，以尽量减少烧伤的可能性。 </li><li>通过使用一个脱毛霜除去毛发从手术部位。由优碘和75％乙醇的3交替磨砂消毒的部位，以确保适当的消毒。放置一个悬垂性，以暴露刚好手术部位，并放置兽医软膏对眼睛，以防止在操作过程中干。 </li></ol> 4.手术过程<ol><li>开胸是在动物的左侧进行的，因此，使一个皮肤切口约1cm到胸骨的左侧，在运行胸骨的长度。 </li><li>使用镊子的肌肉层与底层胸腔分开在两个步骤。观察Deline的阿婷线表示该动物的胸大肌肌肉。解除这种肌肉微微，它从底层腹外斜肌分离，向内缩回。然后，释放从在相同的方式的底层胸腔腹外斜肌肌肉，和缩回横向，提供第二，第三和第四肋的视野开阔图。 </li><li>执行第三和第四肋之间的开胸。轻轻抬起第四肋并使用cauterizer打开第三和第四肋骨之间的胸腔内。拉钩将进一步打开腔暴露心脏。 </li><li>使用镊子破裂心脏的心包薄。 </li><li>结扎左冠状动脉用8-0尼龙单丝缝合。放置缝线从心脏的顶点约4mm，左心房的底部前端的正下方。由心肌的热烫和苏图之后增加左心房的大小决定适当缝合放置再被追平。 </li></ol> 5.补丁安置和胶<ol><li>保持贴剂在37℃和5％CO 2的条件下直至使用。 </li><li>轻轻抬起用平铲端补丁，轻轻把它放在心脏的表面上，在梗塞的部位。 注意:为防止滑落左心室，这个补丁可以代替轻轻接触保持使用无论是压舌板或钳子尖举行。 </li><li> 1比纤维蛋白原凝血酶:通过混合4准备胶水。通过重复的移液混合溶液，直至其开始变稠，通常在1分钟内制备开始后。 </li><li>一旦溶液粘度到达期望的水平，迅速转移约10毫升至贴片表面。注:纤维蛋白胶的凝固时间迅速，对胶水的有效转移提供机会的一个小窗口。 </li></ol> 6.缝合<ol><l我&gt;用6-0尼龙单丝缝合线进行关闭所有缝线。 </li><li>用三到四个独立的间断缝合关闭肋层。 </li><li>肋间层的全封闭之前，插入PE-10套管插入切口撤离胸腔闭合后完成并重新建立正确的胸腔内的压力。 </li><li>关闭胸肌用三到四个独立的间断缝合。 </li><li>密封皮肤层用连续缝合。 </li><li>以下全封闭，附加1.0毫升注射器到所述套管的端部，并用它来撤离胸腔。 </li><li>应用组织粘合剂的切口部位加强缝合部位。 </li></ol> 7.手术后治疗<ol><li>给小鼠注射丁丙诺啡（0.05-1.0毫克/千克）和卡洛芬（2.2毫克/千克）经皮下之前立即恢复冠状动脉结扎和至少20分钟以下的。管理这两种药物在手术后6-8小时，然后给每日两次，三天来控制疼痛和痛苦。 </li><li>连续监测动物直至意识和每隔一小时的前4个小时以下手术。进行第二次检查手术（2-4小时后）的夜间管理止痛的第二剂量。 </li><li>第二天继手术后每天监测动物，直到动物出现稳定。 </li></ol>心功能及组织学分析8 <ol><li>对小鼠后4周的过程来确定继心肌梗塞损伤的程度进行echocardiographs。 </li><li>麻醉小鼠4-5％的异氟醚麻醉室吸入诱导麻醉范围内，然后2％经面罩异氟醚维持麻醉。 </li><li>清除成像区域用脱毛霜，并用一个小动物超声获得跳动的心脏的二维M型超声心动图扫描和记录功能参数ASSOciated它。 </li><li>安乐死的小鼠在二氧化碳室中，然后收集来自动物和地方的心在24小时的10％的福尔马林溶液中。 24小时后，将心转移到70％的乙醇，并保持在4℃直至组织处理。 </li><li>过程中的心，将它们嵌入石蜡块。 </li><li>切5毫米部分通过心脏的心室区域，开始在顶点，并在庭结束。 </li><li>染色切片与两个Masson三色和苏木精和使用标准协议伊红染色切片。 </li></ol>

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The authors declare that they have no competing financial interests.

用这种方法与鼠心肌梗死模型中，我们已经开发出一种系统，其最小化到与其它鼠心肌梗死技术相关的非心肌区域的损害。这些领域包括引起气管损伤，肌肉层的切割，并且肋的破损，以暴露胸腔。我们认为，这些变化改善整体手术结果由于采取保持尽可能多的主要结构的，包括肋和肌肉层，完整的外科手术过程中的照料。 应当指出...

心肌梗死（MI）被定义为血液的中断造成的主要冠状动脉的闭塞心脏的区域。从一个MI造成的损害是由于活心脏组织的重塑成非功能性疤痕组织，从而降低心脏的，或者更具体地说，左心室，击败适当的能力。这导致在血液的体积可以递送到身体与每次心跳，被称为每搏输出量，以及血液泵出心脏的与每次心跳的百分比，被称为射血分数6下降。这些以及其它缩水功能，增加了对心脏的其余部分的应变，以保持足够的功能。通常情况下，它会导致第二心脏发作，这种现象出现在个体7的约10％这增加了应变能变得如此严重。 虽然医疗实践已经发展到治疗心肌梗死的直接后果，没有任何的技术已经发展到停止，慢，或逆转组织重塑的负面影响。干然而细胞疗法已成为一个可能的途径为这样的处理，尽管它们有希望潜力，干细胞还没有被证明是成功的临床设置。一个理论为他们的缺点是无法保证的有益细胞保持在梗死足够长，以产生有利的结果5的部位。它已经显示，细胞的不超过24％被简单地注入到梗死部位存活和保持在损伤部位后1天交货2。一种可能的前景为解决这一问题的细胞保留的是开发出封装任一细胞或治疗剂，其可以被输送到受损部位的生物相容水凝胶系统。所选择的这个协议的水凝胶是聚（乙二醇）二甲基丙烯酸酯，由于其先前在电池封装公关使用ocedures然而，能够包封的任何水凝胶可以用于8。贴片直接向损伤部位的递送保证细胞与组织接触过在延长的时间周期，增加的时间长度，细胞可以提供有利的因素对底层心肌。 瓶颈的修补方法是秉承补丁到心脏表面的难度。许多团体通过各种技术，最普遍的是一个简单的缝合打结构建到心脏表面9,10已经克服了这一点。这已被证明是成功的数目，其中所述构建体是由硬的材料制成的情况下，但试图在水凝胶系统时，由于高的水浓度和接插构建体的细腻性失败。为了克服这个问题，我们已利用模仿血块形成的化学纤维蛋白胶外部粘合剂系统。纤维蛋白胶已用于许多医疗手术，INC泸硬脑膜眼泪，支气管瘘，和角膜移植，突出产品的生物相容性用作伤口的密封剂11-13。此外，血纤维蛋白已经被用于各种心脏目的，包括手术治疗左心室破裂和冠状动脉旁路手术，然而，其作为粘附胶用于心脏修补不常用14-17。的凝血酶和纤维蛋白原的结果在生物相容性胶的简单制剂，可以直接放置在一个外部心脏贴片的外侧，提供了一个可行的粘附系统，以确保补丁到心脏的相互作用。

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	<colgroup>
		<col />
		<col />
		<col />
		<col />
	</colgroup>
	<tbody>
		<tr>
			<td>Harvard Model 687 Mouse Ventilator</td>
			<td>Harvard Apparatus</td>
			<td>55-0001</td>
			<td></td>
		</tr>
		<tr>
			<td>Inintech Biosciences LLC Dry Glass Bead Sterilizer</td>
			<td>Fisher Scientific</td>
			<td>NC9531961</td>
			<td></td>
		</tr>
		<tr>
			<td>Leica MZ6 surgical microscope</td>
			<td>Leica</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Cautery Kit</td>
			<td>Gemini</td>
			<td>GEM 5917</td>
			<td></td>
		</tr>
		<tr>
			<td>Delicate Forceps - 0.4mm Tips Angled</td>
			<td>Fine Science Tools</td>
			<td>11063-07</td>
			<td></td>
		</tr>
		<tr>
			<td>Agricola Retractor - 3.5cm Spread</td>
			<td>Fine Science Tools</td>
			<td>17005-04</td>
			<td></td>
		</tr>
		<tr>
			<td>Spring Scissors - 2.5mm Blades Straight</td>
			<td>Fine Science Tools</td>
			<td>15000-08</td>
			<td></td>
		</tr>
		<tr>
			<td>Castroviejo Needle Holder - w/Lock Tungsten Carbide 14cm</td>
			<td>Fine Science Tools</td>
			<td>12565-14</td>
			<td></td>
		</tr>
		<tr>
			<td>Iris Scissors - Delicate Straight 10.5 cm</td>
			<td>Fine Science Tools</td>
			<td>14060-10</td>
			<td></td>
		</tr>
		<tr>
			<td>8-0 monofilament suture</td>
			<td>Ethicon</td>
			<td>8730P</td>
			<td></td>
		</tr>
		<tr>
			<td>6-0 Silk suture</td>
			<td>Ethicon</td>
			<td>639G</td>
			<td></td>
		</tr>
		<tr>
			<td>Thrombin</td>
			<td>Sigma</td>
			<td>T7009</td>
			<td></td>
		</tr>
		<tr>
			<td>Fibrinogen</td>
			<td>Sigma</td>
			<td>F3879</td>
			<td></td>
		</tr>
		<tr>
			<td>Vetbond Tissue Adhesive</td>
			<td>3M</td>
			<td>1469SB</td>
			<td></td>
		</tr>
	</tbody>
</table>

a hydrogel construct and fibrin-based glue approach to deliver therapeutics in a murine myocardial infarction model.

The murine MI model is widely recognized in the field of cardiovascular disease, and has consistently been used as a first step to test the efficacy of treatments in vivo1. The traditional, established protocol has been further fine-tuned to minimize the damage to the animal. Notably, the pectoral muscle layers are teased away rather than simply cut, and the thoracotomy is approached intercostally as opposed to breaking the ribs in a sternotomy, preserving the integrity of the ribcage. With these changes, the overall stress on the animal is decreased. 
Stem cell therapies aimed to alleviate the damage caused by MIs have shown promise over the years for their pro-angiogenic and anti-apoptotic benefits. Current approaches of delivering cells to the heart surface typically involve the injection of the cells either near the damaged site, within a coronary artery, or into the peripheral blood stream2-4. While the cells have proven to home to the damaged myocardium, functionality is limited by their poor engraftment at the site of injury, resulting in diffusion into the blood stream5. This manuscript highlights a procedure that overcomes this obstacle with the use of a cell-encapsulated hydrogel patch. The patch is fabricated prior to the surgical procedure and is placed on the injured myocardium immediately following the occlusion of the left coronary artery. To adhere the patch in place, biocompatible external fibrin glue is placed directly on top of the patch, allowing for it to dry to both the patch and the heart surface. This approach provides a novel adhesion method for the application of a delicate cell-encapsulating therapeutic construct.

在外科手术过程中，左冠状动脉的结扎可以由标热烫闭塞动脉的下游来鉴定。作为结婚前一个测试，缝合线可以简单地拧紧，以检查它是否在适当的位置。此外，由于在动脉导致在左心房的适当敲打的能力几乎瞬间降低闭塞，左心房将放大响应于血液系统中的回流。 M型超声心动图测量，作为早在2天后梗死表现出停止左室壁运动的，表明肌肉重建。从数据做出定性的计算显?...

Watch this Scientific Journal Video about A Hydrogel Construct and Fibrin-based Glue Approach to Deliver Therapeutics in a Murine Myocardial Infarction Model. at JoVE.com

A Hydrogel Construct and Fibrin-based Glue Approach to Deliver Therapeutics in a Murine Myocardial Infarction Model.

This protocol aims to alleviate the limitation of poor cell engraftment for stem cell treatment of myocardial infarctions through the use of a hydrogel system and a fibrin-based glue. With this approach, cell-to-tissue contact post-infarction can be maintained, increasing the therapeutic potential of beneficial agents at the site of injury.

水凝胶构造和纤维蛋白基胶方法来提供治疗的小鼠心肌梗死模型。

The murine MI model is widely recognized in the field of cardiovascular disease, and has consistently been used as a first step to test the efficacy of ...

a-hydrogel-construct-fibrin-based-glue-approach-to-deliver

Research

JoVE Journal

Medicine

8.6K 次观看.  University of Illinois at Urbana-Champaign. This protocol aims to alleviate the limitation of poor cell engraftment for stem cell treatment of myocardial infarctions through the use of a hydrogel system and a fibrin-based glue. With this approach, cell-to-tissue contact post-infarction can be maintained, increasing the therapeutic potential of beneficial agents at the site of injury.

视频: A Hydrogel Construct and Fibrin-based Glue Approach to Deliver Therapeutics in a Murine Myocardial Infarction Model.

Acute Myocardial Infarction in Rats

With heart failure leading the cause of death in the USA (Hunt), biomedical research is fundamental to advance medical treatments for cardiovascular diseases. Animal models that mimic human cardiac disease, such as myocardial infarction (MI) and ischemia-reperfusion (IR) that induces heart failure as well as pressure-overload (transverse aortic constriction) that induces cardiac hypertrophy and heart failure (Goldman and Tarnavski), are useful models to study cardiovascular disease. In particular, myocardial ischemia (MI) is a leading cause for cardiovascular morbidity and mortality despite controlling certain risk factors such as arteriosclerosis and treatments via surgical intervention (Thygesen). Furthermore, an acute loss of the myocardium following myocardial ischemia (MI) results in increased loading conditions that induces ventricular remodeling of the infarcted border zone and the remote non-infarcted myocardium. Myocyte apoptosis, necrosis and the resultant increased hemodynamic load activate multiple biochemical intracellular signaling that initiates LV dilatation, hypertrophy, ventricular shape distortion, and collagen scar formation. This pathological remodeling and failure to normalize the increased wall stresses results in progressive dilatation, recruitment of the border zone myocardium into the scar, and eventually deterioration in myocardial contractile function (i.e. heart failure). The progression of LV dysfunction and heart failure in rats is similar to that observed in patients who sustain a large myocardial infarction, survive and subsequently develops heart failure (Goldman). The acute myocardial infarction (AMI) model in rats has been used to mimic human cardiovascular disease; specifically used to study cardiac signaling mechanisms associated with heart failure as well as to assess the contribution of therapeutic strategies for the treatment of heart failure. The method described in this report is the rat model of acute myocardial infarction (AMI). This model is also referred to as an acute ischemic cardiomyopathy or ischemia followed by reperfusion (IR); which is induced by an acute 30-minute period of ischemia by ligation of the left anterior descending artery (LAD) followed by reperfusion of the tissue by releasing the LAD ligation (Vasilyev and McConnell). This protocol will focus on assessment of the infarct size and the area-at-risk (AAR) by Evan's blue dye and triphenyl tetrazolium chloride (TTC) following 4-hours of reperfusion; additional comments toward the evaluation of cardiac function and remodeling by modifying the duration of reperfusion, is also presented. Overall, this AMI rat animal model is useful for studying the consequence of a myocardial infarction on cardiac pathophysiological and physiological function.

The rat model of acute myocardial infarction (AMI) is useful to study the consequence of a MI on cardiac pathophysiological and physiological function.

大鼠急性心肌梗死

With heart failure leading the cause of death in the USA (Hunt), biomedical research is fundamental to advance medical treatments for cardiovascular ...

科研

医学

Coronary Artery Ligation and Intramyocardial Injection in a Murine Model of Infarction

Mouse models are a valuable tool for studying acute injury and chronic remodeling of the myocardium in vivo. With the advent of genetic modifications to the whole organism or the myocardium and an array of biological and/or synthetic materials, there is great potential for any combination of these to assuage the extent of acute ischemic injury and impede the onset of heart failure pursuant to myocardial remodeling. 
Here we present the methods and materials used to reliably perform this microsurgery and the modifications involved for temporary (with reperfusion) or permanent coronary artery occlusion studies as well as intramyocardial injections. The effects on the heart that can be seen during the procedure and at the termination of the experiment in addition to histological evaluation will verify efficacy. 
Briefly, surgical preparation involves anesthetizing the mice, removing the fur on the chest, and then disinfecting the surgical area. Intratracheal intubation is achieved by transesophageal illumination using a fiber optic light. The tubing is then connected to a ventilator. An incision made on the chest exposes the pectoral muscles which will be cut to view the ribs. For ischemia/reperfusion studies, a 1 cm piece of PE tubing placed over the heart is used to tie the ligature to so that occlusion/reperfusion can be customized. For intramyocardial injections, a Hamilton syringe with sterile 30gauge beveled needle is used. When the myocardial manipulations are complete, the rib cage, the pectoral muscles, and the skin are closed sequentially. Line block analgesia is effected by 0.25% marcaine in sterile saline which is applied to muscle layer prior to closure of the skin. The mice are given a subcutaneous injection of saline and placed in a warming chamber until they are sternally recumbent. They are then returned to the vivarium and housed under standard conditions until the time of tissue collection. At the time of sacrifice, the mice are anesthetized, the heart is arrested in diastole with KCl or BDM, rinsed with saline, and immersed in fixative. Subsequently, routine procedures for processing, embedding, sectioning, and histological staining are performed. 
Nonsurgical intubation of a mouse and the microsurgical manipulations described make this a technically challenging model to learn and achieve reproducibility. These procedures, combined with the difficulty in performing consistent manipulations of the ligature for timed occlusion(s) and reperfusion or intramyocardial injections, can also affect the survival rate so optimization and consistency are critical.

Numerous genetic manipulations and/or intramyocardial injections of genes, proteins, cells, and/or biomaterials are superimposed upon the dimension of time in studies of acute ischemia/ reperfusion injury and chronic remodeling in mice. This video illustrates the microsurgical procedures for ischemia/reperfusion, permanent coronary artery ligation, and intramyocardial injection studies.

结扎冠状动脉内注射在小鼠模型梗死

Mouse models are a valuable tool for studying acute injury and chronic remodeling of the myocardium in vivo.  With the advent of genetic modifications to ...

A Murine Closed-chest Model of Myocardial Ischemia and Reperfusion

Surgical trauma by thoracotomy in open-chest models of coronary ligation induces an immune response which modifies different mechanisms involved in ischemia and reperfusion. Immune response includes cytokine expression and release or secretion of endogenous ligands of innate immune receptors. Activation of innate immunity can potentially modulate infarct size. We have modified an existing murine closed-chest model using hanging weights which could be useful for studying myocardial pre- and postconditioning and the role of innate immunity in myocardial ischemia and reperfusion. This model allows animals to recover from surgical trauma before onset of myocardial ischemia.
Volatile anesthetics have been intensely studied and their preconditioning effect for the ischemic heart is well known. However, this protective effect precludes its use in open chest models of coronary artery ligation. Thus, another advantage could be the use of the well controllable volatile anesthetics for instrumentation in a chronic closed-chest model, since their preconditioning effect lasts up to 72 hours. Chronic heart diseases with intermittent ischemia and multiple hit models are other possible applications of this model.
For the chronic closed-chest model, intubated and ventilated mice undergo a lateral blunt thoracotomy via the 4th intercostal space. Following identification of the left anterior descending a ligature is passed underneath the vessel and both suture ends are threaded through an occluder. Then, both suture ends are passed through the chest wall, knotted to form a loop and left in the subcutaneous tissue. After chest closure and recovery for 5 days, mice are anesthetized again, chest skin is reopened and hanging weights are hooked up to the loop under ECG control.
At the end of the ischemia/reperfusion protocol, hearts can be stained with TTC for infarct size assessment or undergo perfusion fixation to allow morphometric studies in addition to histology and immunohistochemistry.

Surgical trauma induces an inflammatory response. Cytokines and endogenous ligands are known to modulate myocardial infarct size following ischemia and reperfusion. We present a modified closed-chest model of murine ischemia and reperfusion using hanging weights to minimize effects of thoracotomy.

闭胸鼠心肌缺血再灌注模型

Surgical trauma by thoracotomy in open-chest models of coronary ligation induces an immune response which modifies different mechanisms involved in ...

Myocardial Infarction and Functional Outcome Assessment in Pigs

Introduction of newly discovered cardiovascular therapeutics into first-in-man trials depends on a strictly regulated ethical and legal roadmap. One important prerequisite is a good understanding of all safety and efficacy aspects obtained in a large animal model that validly reflect the human scenario of myocardial infarction (MI). Pigs are widely used in this regard since their cardiac size, hemodynamics, and coronary anatomy are close to that of humans. Here, we present an effective protocol for using the porcine MI model using a closed-chest coronary balloon occlusion of the left anterior descending artery (LAD), followed by reperfusion. This approach is based on 90 min of myocardial ischemia, inducing large left ventricle infarction of the anterior, septal and inferoseptal walls. Furthermore, we present protocols for various measures of outcome that provide a wide range of information on the heart, such as cardiac systolic and diastolic function, hemodynamics, coronary flow velocity, microvascular resistance, and infarct size. This protocol can be easily tailored to meet study specific requirements for the validation of novel cardioregenerative biologics at different stages (i.e. directly after the acute ischemic insult, in the subacute setting or even in the chronic MI once scar formation has been completed). This model therefore provides a useful translational tool to study MI, subsequent adverse remodeling, and the potential of novel cardioregenerative agents.

This protocol describes the porcine myocardial infarction (MI) model using a 90 min closed-chest coronary balloon occlusion of the left anterior descending artery (LAD), followed by reperfusion. Furthermore, the protocol for several outcome parameters, such as cardiac function, hemodynamics, microvascular resistance, and infarct size, are also presented.

心肌梗死和功能预后评估的猪

Introduction of newly discovered cardiovascular therapeutics into first-in-man trials depends on a strictly regulated ethical and legal roadmap. One ...

Primary Outcome Assessment in a Pig Model of Acute Myocardial Infarction

Mortality after acute myocardial infarction remains substantial and is associated with significant morbidity, like heart failure. Novel therapeutics are therefore required to confine cardiac damage, promote survival and reduce the disease burden of heart failure. Large animal experiments are an essential part in the translational process from experimental to clinical therapies. To optimize clinical translation, robust and representative outcome measures are mandatory. The present manuscript aims to address this need by describing the assessment of three clinically relevant outcome modalities in a pig acute myocardial infarction (AMI) model: infarct size in relation to area at risk (IS/AAR) staining, 3-dimensional transesophageal echocardiography (TEE) and admittance-based pressure-volume (PV) loops. Infarct size is the main determinant driving the transition from AMI to heart failure and can be quantified by IS/AAR staining. Echocardiography is a reliable and robust tool in the assessment of global and regional cardiac function in clinical cardiology. Here, a method for three-dimensional transesophageal echocardiography (3D-TEE) in pigs is provided. Extensive insight into cardiac performance can be obtained by admittance-based pressure-volume (PV) loops, including intrinsic parameters of myocardial function that are pre- and afterload independent. Combined with a clinically feasible experimental study protocol, these outcome measures provide researchers with essential information to determine whether novel therapeutic strategies could yield promising targets for future testing in clinical studies.

Reliable and accurate outcome assessment is the key for translation of preclinical therapies into clinical treatment. The current paper describes how to assess three clinically relevant primary outcome parameters of cardiac performance and damage in a pig acute myocardial infarction model.

急性心肌梗死的猪模型主要成果评估

Mortality after acute myocardial infarction remains substantial and is associated with significant morbidity, like heart failure. Novel therapeutics are ...

Myocardial Infarction in Neonatal Mice, A Model of Cardiac Regeneration

Myocardial infarction induced by coronary artery ligation has been used in many animal models as a tool to study the mechanisms of cardiac repair and regeneration, and to define new targets for therapeutics. For decades, models of complete heart regeneration existed in amphibians and fish, but a mammalian counterpart was not available. The recent discovery of a postnatal window during which mice possess regenerative capabilities has led to the establishment of a mammalian model of cardiac regeneration. A surgical model of mammalian cardiac regeneration in the neonatal mouse is presented herein. Briefly, postnatal day 1 (P1) mice are anesthetized by isoflurane and placed on an ice pad to induce hypothermia. After the chest is opened, and the left anterior descending coronary artery (LAD) is visualized, a suture is placed around the LAD to inflict myocardial ischemia in the left ventricle. The surgical procedure takes 10-15 min. Visualizing the coronary artery is crucial for accurate suture placement and reproducibility. Myocardial infarction and cardiac dysfunction are confirmed by triphenyl-tetrazolium chloride (TTC) staining and echocardiography, respectively. Complete regeneration 21 days post myocardial infarction is verified by histology. This protocol can be used to as a tool to elucidate mechanisms of mammalian cardiac regeneration after myocardial infarction.

This protocol describes a highly reproducible model of cardiac regeneration by surgical induction of myocardial infarction in the left ventricle of postnatal day 1 mice. The method involves induction of hypothermic anesthesia and ligation of the left anterior descending coronary artery.

心肌梗死新生小鼠，心肌再生的模型

Myocardial infarction induced by coronary artery ligation has been used in many animal models as a tool to study the mechanisms of cardiac repair and ...

Murine Myocardial Infarction Model using Permanent Ligation of Left Anterior Descending Coronary Artery

Myocardial infarction (MI) and acute coronary diseases are among the most prominent causes of death in population with western lifestyle. The murine models of MI with permanent ligation of left-anterior descending (LAD) coronary artery closely mimics MI in humans. Murine models benefit from the extensive genetic engineering available nowadays. Here we propose a reproducible murine surgical model of myocardial infarction by permanent LAD coronary ligation. Our technique comprises anesthesia with ketamine/xylazine that can be rapidly reversed by administration of an antagonist, intubation without tracheotomy for mechanical-assisted ventilation, ventilation with application of extrinsic positive end-expiratory pressure (PEEP) to avoid alveolar collapse, a thoracotomy method limiting to the minimum surgical lesions made to skeletal muscles, and lung inflation without thoracentesis. This method is sparsely invasive, reproducible and reduces post-surgery mortality and complications.

Herein we describe a surgical procedure showing how to achieve permanent ligation of the left-anterior descending coronary artery in mice. This model is of high relevance to investigate the pathophysiology of myocardial infarction and the concomitant biological processes.

使用左前部降冠动脉的永久结扎的月心肌梗死模型

Myocardial infarction (MI) and acute coronary diseases are among the most prominent causes of death in population with western lifestyle. The murine ...

A Cryoinjury Model to Study Myocardial Infarction in the Mouse

The use of animal models is essential for developing new therapeutic strategies for acute coronary syndrome and its complications. In this article, we demonstrate a murine cryoinjury infarct model that generates precise infarct sizes with high reproducibility and replicability. In brief, after intubation and sternotomy of the animal, the heart is lifted from the thorax. The probe of a handheld liquid nitrogen delivery system is applied onto the myocardial wall to induce cryoinjury. Impaired ventricular function and electrical conduction can be monitored with echocardiography or optical mapping. Transmural myocardial remodeling of the infarcted area is characterized by collagen deposition and loss of cardiomyocytes. Compared to other models (e.g., LAD-ligation), this model utilizes a handheld liquid nitrogen delivery system to generate more uniform infarct sizes.

This article demonstrates a model to study cardiac remodeling after myocardial cryoinjury in mice.

研究小鼠心肌梗死的低温损伤模型

The use of animal models is essential for developing new therapeutic strategies for acute coronary syndrome and its complications. In this article, we ...

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.

建立干细胞治疗心肌梗塞心力衰竭的猪模型

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

Intramyocardial Transplantation of MSC-Loading Injectable Hydrogels after Myocardial Infarction in a Murine Model

One of the major issues facing current cardiac stem cell therapies for preventing postinfarct heart failure is the low retention and survival rates of transplanted cells within the injured myocardium, limiting their therapeutic efficacy. Recently, the use of scaffolding biomaterials has gained attention for improving and maximizing stem cell therapy. The objective of this protocol is to introduce a simple and straightforward technique to transplant bone marrow-derived mesenchymal stem cells (MSCs) using injectable hydroxyphenyl propionic acid (GH) hydrogels; the hydrogels are favorable as a cell delivery platform for cardiac tissue engineering applications due to their ability to be cross-linked in situ and high biocompatibility. We present a simple method to fabricate MSC-loading GH hydrogels (MSC/hydrogels) and evaluate their survival and proliferation in three-dimensional (3D) in vitro culture. In addition, we demonstrate a technique for intramyocardial transplantation of MSC/hydrogels in mice, describing a surgical procedure to induce myocardial infarction (MI) via left anterior descending (LAD) coronary artery ligation and subsequent MSC/hydrogels transplantation.

Stem cell-based therapy has emerged as an efficient strategy to repair injured cardiac tissues after myocardial infarction. We provide an optimal in vivo application for stem cell transplantation using gelatin hydrogels that are able to be enzymatically cross-linked.

Murine 模型心肌梗塞后 MSC 加载注射水凝胶的心内移植

One of the major issues facing current cardiac stem cell therapies for preventing postinfarct heart failure is the low retention and survival rates of ...