This work was supported by grant 13IRG14560023 from the American Heart Association and NIH grants AI078972 and HL108054 to CJO. Support for AOB was from the NIH National Center for Advancing Translational Sciences NIH ULTR001120, and F31 A110417701.

注：所有小鼠实验进行了审查，并在得克萨斯健康科学中心大学圣安东尼奥（协议＃13032-34-01C）批准的机构动物护理和使用委员会。动物保健和遵守公法89-544（动物福利法）及其修正案，公共卫生服务的指导方针，以及指南实验动物的护理和使用（健康与人类服务部）的实验方案。 1.感染<ol><li>获得10-12周龄岁之间的男性和女性的BALB / c小鼠。 注：S.血清型肺炎 4个应变TIGR4是已测序的16剧毒临床分离。 </li><li>生长TIGR4中的Todd-Hewitt肉汤，在37℃在5％CO 2，直到达到一个光学密度（OD）0.5 620（对应于1.0×10 8菌落形成单位[CFU] / ml）中。颗粒通过离心肺炎球菌培养，在3 10分钟，500×g离心并除去使用真空线上清液。暂停和稀的细菌，用无菌磷酸盐缓冲盐水（PBS）以1.0×10 4 CFU / ml的终浓度。 </li><li>使用感应室，麻醉小鼠在氧气2.5％异氟烷气化。轻轻捏脚趾用生硬的镊子确认麻醉状态。 <ol><li>由它的浮渣保持了麻醉的小鼠和直立用一只手，注入每只小鼠腹膜内（IP）用100μl的S的使用注射器用27-30号针（对应于1.0×10 3 CFU一个挑战剂量） 肺炎悬浮液。将鼠标放回笼子。小鼠通常唤醒注射后30-40秒。 </li></ol></li><li>允许感染进行至少24（早期）或30（高级）小时。 <ol><li>安乐死小鼠CO 2窒息。执行颈椎脱位，确保鼠标已经死亡。安乐死是通过去除进一步证实在1.6步的心脏。 </li></ol></li><li>消毒用酒精棉签尾部操起一个2-3毫米的部分。收集2微升的血液和连续稀释血液的10倍PBS含肝素钠1 U / ml的五倍。在胰蛋白大豆血琼脂平板上平板系列稀释并在5％CO 2，从菌落第二天外推计算菌血症水平孵育过夜，在37℃。 </li><li>固定鼠标在上一个外科平台仰卧位置。喷雾胸部用70％乙醇和拍干。用手术剪刀和镊子打开胸腔，取出肋骨，和横切隔膜暴露心脏和肺部。用剪刀剪连接到心脏的血管，轻轻摘除心脏小心，避免用钳子青紫。 </li><li>冲洗PBS的心脏，然后放入组织标本收集磁带，这样冠状切片将组织切片中获得。对于石蜡包埋，解放军CE在10％缓冲的甲醛溶液和第二天卡匣发送用于石蜡包埋。 <ol><li>可替代地使用cryomold已放置在干冰上内OCT化合物闪光冻结。 </li></ol></li></ol> 2.可视化心脏病变及病变范围内的肺炎球菌<ol><li>减少石蜡包埋的心脏部分，使4室的心脏在每个组织切片可见。切载玻片在5微米厚。 </li><li> Deparaffinize和染色的幻灯片使用标准方法17苏木和伊红（H＆E）。 </li><li>视图，H＆E染色使用光学显微镜在低（100X，200X）和高（油浸：400X，1,000X）心脏切片的放大倍数。表征心中microlesions在心肌的存在。在这里，获得使用蔡司Axioskop 2显微镜配备了100X 1.3数值孔径计划，NEOFLUAR客观H＆E影像。 注：在早期心脏升esions由它们的差是有色的外观和在某些情况下的免疫细胞（单核细胞和粒细胞）什么似乎是单核细胞的存在区别。在先进的病变，尤其是在看到30小时，免疫细胞缺失和大空泡状病变在他们的地方观察。要注意的是，虽然在心肌病变发展的早期阶段的免疫细胞流入可发生不似乎是一个要求是很重要的。 </li></ol> 3.免疫荧光显微镜的范围内Microlesions肺炎球菌<ol><li>部分心脏中cryomolds用切片机以厚度为5微米，代替部分上带正电荷的载玻片上。让心脏冰冻切片解冻和空气干燥。定为10分钟，在10％的中性缓冲的福尔马林部分（pH为6.8-7.2，在25℃）。 </li><li>在PBS中5分钟洗载玻片（3×），透化在0.2％的Triton X-100的PBS中15分钟，然后在PBS中再次洗涤。 </li><li>块组织secti<html>对载玻片用1小时在PBS中的10％山羊血清。 </li><li>漂洗载玻片用PBS，盖孵育心脏切片用兔抗血清型4肺炎球菌血清在1：在37℃下在PBS 1000，用10％山羊血清2小时。 注：对于阴性对照调查人员应使用天真兔抗血清</li><li>之后用PBS洗涤，盖孵育，用10％山羊血清的PBS中的部分用山羊抗兔FITC缀合的抗体以1：在37℃2000 30分钟。 </li><li>按照制造商的指示，用DAPI（4&#39;，6-二脒基-2-苯基吲哚）以5mg / ml的真核细胞核可视化。清洗和安装组织切片与FluorSave。 </li><li>使用共聚焦显微镜系统在低和高放大倍数取得的荧光图像。调节的数值孔径相应的，以获得最佳的效果。这里，使用共聚焦系统，具有60X 1.42数值孔径物镜获取荧光图像。 </li></ol>化粪池小鼠“&gt; 4。抗生素救援<ol><li>小鼠感染IP用1.0×10 3 CFU TIGR4的开始如先前在步骤1中所示。 </li><li>在30小时后的感染开始，施用在100μl盐水药用级氨苄青霉素IP（80毫克/公斤体重），每12小时36小时。如在用于石蜡包埋和组织，切片步骤1.6中所述收集心在期望的时间点和处理。 </li></ol> 5.胶原沉积检测<ol><li>使用二甲苯和分级醇洗涤与使用标准方法17去离子水，deparaffinize和水合物的组织切片。治疗用含水0.2％磷酸部分为5分钟，并用去离子水冲洗5分钟。 </li><li>染色切片用0.1％天狼星红在苦味酸2小时，在室温下进行。 </li><li>在0.01N盐酸3分钟洗部分和漂洗用70％乙醇1分钟。 </li><li> Dehydra使用相同的TE成功的部分，但是为了反洗到5.1概括的，脱水增加心脏部分，分级乙醇浓度再二甲苯。 </li><li>摩组织切片与微观评估的安装解决方案。 </li><li>以确定胶原沉积的区域中，检查染色组织切片的深读染色使用光学显微镜在低和高放大倍数的区域。这是红色心脏的心室中的任何区域是胶原蛋白已存入的区域。如果没有这个红染色领域是健康组织的网站缺乏胶原沉积。此处，获取使用装有20X 0.5数值孔径物镜的显微镜中苦天狼红染色的图像。 注：该庭污渍红健康组织。 </li></ol>

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The authors declare no conflict of interest in this study.

在这份报告中，高度重复的方法诱导S.肺炎介导的小鼠心脏microlesions IPD期间和技术的可视化展示。小鼠感染剂量的细菌，优于通关免疫系统的门槛，导致高档菌血症，​​类似这人类败血症期间发生，并导致最终的细菌移位到心肌。对于未知的原因，S.肺炎的心脏是能够通过宿主的免疫反应来复制偏向虎山行。这是正在进行的调查的主题，本出版物可以促进其他研究者在他们的努力?...

成人社区获得性肺炎（CAP）住院进行了心脏不良事件，有助于死亡率1-4文件化的风险。在最近的一项研究科拉莱斯-Medina 等人 ，发现心脏并发症要关联有和/或负责肺炎相关死亡3的27％。 肺炎链球菌 （肺炎球菌），CAP和败血症5的最常见原因，已直接与多达19％的人承认的成人患者6心脏不良事件相关。与肺炎相关的不良心脏事件包括为了降低频率6的新的或恶化充血性心脏衰竭，心律失常和心肌梗死。 在最近的一个公共科学图书馆病原布朗等人的文章。，肺炎链球菌 ，发现通过心脏血管内皮，进入myocardiu是能够易位米，并形成独立非化脓性病变显微镜（microlesions）侵袭性肺炎球菌疾病（IPD）7时充满细菌侧脑室。观察心脏样本中心脏microlesion形成的证据来自非人类灵长类动物和个人谁曾死于肺炎球菌感染。同样，实验感染小鼠重复开发的心脏microlesions。在小鼠中，microlesion大小和数量呈正的持续时间和菌血症，心肌肌钙蛋白血清水平的严重程度，以及异常心脏电相关。细菌易位入心脏，发现发生通过负责跨越血-脑屏障和肺炎球菌性脑膜炎的发展， 也就是 ，胆碱结合蛋白A介导的血管内皮细胞的浸润在层粘连蛋白受体和血小板的肺炎球菌的易位相同的机制-activating因子受体依赖性8。 Microles离子的形成还需要肺炎球菌成孔毒素肺炎球菌被发现杀死心肌7。 肺炎球菌心脏microlesions是从由其他革兰氏阳性菌，包括金黄色葡萄球菌所引起的化脓性软组织和心脏脓肿鲜明。这些特点是由细菌中性粒细胞和纤维蛋白沉积9,10包围的单一病灶。 Microlesions由S.形成肺炎在尺寸，在整个受影响的心脏分布更小，且缺乏免疫细胞浸润。在感染的早期阶段，microlesions引起由S.肺炎表现为受损或发炎的组织让人想起心肌病的病理征象领域。一些单核细胞可能在此期间观察到，但是他们的存在是短暂的，病变迅速成为坏死在外观，充满了细菌，同时继续GR流的大小，直到动物或抗微生物干预的死亡。重要的是，内抗生素干预后3天，大量的免疫细胞浸润被观察到前者病变部位，这是伴随着强劲的胶原沉积。类似心脏重塑已报道发生对心脏功能11-15持久后果沿着以下梗死。因此，microlesions是心脏相关死亡率的恢复期个体的发病率增加谁生存了疾病发作期间的IPD和可能发生的不良心脏事件的潜在的解释。 这里，指令被设置在IPD与心肌损伤形成和心肌microlesions可视化的实验小鼠模型中，在感染的早期和晚期阶段。已保存的抗菌干预动物的协议检测胶原沉积的证明。本文的目的是至facilitate在这一重要和新颖的肺炎病理其他研究者的研究。

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	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:308px;">Name of Reagent/ Equipment</td>
			<td style="width:175px;">Company</td>
			<td style="width:119px;">Catalog Number</td>
			<td style="width:167px;">Comments/Description</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:308px;">Todd-Hewitt broth&#160;</td>
			<td style="width:175px;">Neogen</td>
			<td>7161A</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:308px;">O.C.T Compound&#160;</td>
			<td style="width:175px;">Tissue-Tek (Sakura Finetek)&#160;</td>
			<td style="width:119px;">4583</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Triton X-100&#160;</td>
			<td style="width:175px;">Fisher Scientific (Acros)</td>
			<td>9002-93-1</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:308px;">Normal Goat Serum</td>
			<td style="width:175px;">Abcam</td>
			<td style="width:119px;">ab7481</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">anti-serotype 4 pneumococcus antiserum&#160;</td>
			<td>Statens Serum Institut</td>
			<td style="width:119px;">16747</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;">goat anti-rabbit FITC conjugated antibody&#160;</td>
			<td style="width:175px;">Jackson ImmunoResearch</td>
			<td style="width:119px;">111-096-144</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:308px;">DAPI</td>
			<td style="width:175px;">Invitrogen</td>
			<td style="width:119px;">D1306</td>
			<td></td>
		</tr>
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			<td height="21" style="height:21px;width:308px;">Fluorsave</td>
			<td style="width:175px;">Millipore</td>
			<td>345789</td>
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			<td height="21" style="height:21px;width:308px;">Permount</td>
			<td>Fisher Scientific</td>
			<td>S70104</td>
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			<td>A9393</td>
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		<tr height="21">
			<td height="21" style="height:21px;">phosphomolybdic acid 0.2%&#160;</td>
			<td>Electron Microscopy Sciences</td>
			<td>26357-01</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:308px;">0.1% Sirius Red in picric acid&#160;</td>
			<td>Electron Microscopy Sciences</td>
			<td>26357-02</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">0.01 N hydrochloric acid&#160;</td>
			<td>Electron Microscopy Sciences</td>
			<td>26357-03</td>
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			<td height="21" style="height:21px;width:308px;">Tissue Tack Microscope Slides</td>
			<td style="width:175px;">Polysciences, Inc</td>
			<td style="width:119px;">24216</td>
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		<tr height="21">
			<td height="21" style="height:21px;width:308px;">Paralube Vet Ointment</td>
			<td style="width:175px;">Dechra</td>
			<td>12920060</td>
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		<tr height="21">
			<td height="21" style="height:21px;width:308px;">Hematoxylin and Eosin Staining Kit</td>
			<td style="width:175px;">unspeciified&#160;</td>
			<td style="width:119px;"></td>
			<td>Standard</td>
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visualization of streptococcus pneumoniae within cardiac microlesions and subsequent cardiac remodeling

期间菌血症肺炎链球菌可在血管内皮转运进入心肌和形成离散菌填充微观病变（microlesions）是显着的，由于缺乏浸润免疫细胞。由于其释放心脏毒性的产品，S.肺炎 microlesions内被认为有助于向心脏衰竭是在成人中雷酸侵袭性肺炎球菌疾病中经常观察到。在此证明一个协议，用于实验的小鼠感染，导致30小时内可重复的心脏microlesion形成。设在microlesion鉴定苏木精 - 伊红染色心脏切片和早期和晚期microlesions之间的形态区别的指令被突出显示。设置在一个协议，用于验证S的指令肺炎使用抗肺炎球菌荚膜多糖和microlesions内免疫荧光显微镜。最后，提供了一种协议，用于抗生素干预拯救感染小鼠和用于检测和瘢痕形成在恢复期小鼠的心脏评估。一起，这些协议将促进肺炎球菌心脏侵袭，心肌细胞死亡，心脏重塑底层的暴露至S.的结果的分子机制的研究肺炎 ，以及在心脏的肺炎球菌的免疫应答。

Microlesions由H＆E可视化心脏microlesions观察到的H＆E染色小鼠与IPD下面的IP挑战，TIGR4心脏部分心室。 图1说明了增加24和​​30小时后感染（HPI），与这些病变的大小更多的苏木染色的（ 即 ，紫蓝色）肺炎病灶内可见。 Microlesions的特点是一个液泡样形态，更密集的曙红（红色）染色紧邻病灶部位的心肌细胞中的，病灶内双球菌，和一般缺乏免疫细胞。?...

Watch this Scientific Journal Video about Visualization of Streptococcus pneumoniae within Cardiac Microlesions and Subsequent Cardiac Remodeling at JoVE.com

Visualization of Streptococcus pneumoniae within Cardiac Microlesions and Subsequent Cardiac Remodeling

Streptococcus pneumoniae forms discrete non-purulent microscopic lesions in the heart. Outlined is the protocol for a murine model of cardiac microlesion formation. Instruction is provided on microlesion visualization using microscopy, discrimination between early and late microlesions, and methods to detect cardiac remodeling in hearts of convalescent animals.

可视化<em&gt;肺炎链球菌</em&gt;在心脏Microlesions和随后的心脏重构

During bacteremia Streptococcus pneumoniae can translocate across the vascular endothelium into the myocardium and form discrete bacteria-filled ...

visualization-streptococcus-pneumoniae-within-cardiac-microlesions

Research

JoVE Journal

Medicine

Visualization of Streptococcus pneumoniae within Cardiac Microlesions and Subsequent Cardiac Remodeling

8.9K 次观看.  The University of Texas Health Science Center at San Antonio. Streptococcus pneumoniae forms discrete non-purulent microscopic lesions in the heart. Outlined is the protocol for a murine model of cardiac microlesion formation. Instruction is provided on microlesion visualization using microscopy, discrimination between early and late microlesions, and methods to detect cardiac remodeling in hearts of convalescent animals. 

视频: Visualization of Streptococcus pneumoniae within Cardiac Microlesions and Subsequent Cardiac Remodeling

Cardiac Stress Test Induced by Dobutamine and Monitored by Cardiac Catheterization in Mice

Dobutamine is a &beta;-adrenergic agonist with an affinity higher for receptor expressed in the heart (&beta;1) than for receptors expressed in the arteries (&beta;2). When systemically administered, it increases cardiac demand. Thus, dobutamine unmasks abnormal rhythm or ischemic areas potentially at risk of infarction. 
Monitoring of heart function during a cardiac stress test can be performed by either ecocardiography or cardiac catheterization. The latter is an invasive but more accurate and informative technique that the former.
Cardiac stress test induced by dobutamine and monitored by cardiac catheterization accomplished as described here allows, in a single experiment, the measurement of the following hemodynamic parameters: heart rate (HR), systolic pressure, diastolic pressure, end-diastolic pressure, maximal positive pressure development (dP/dtmax) and maximal negative pressure development (dP/dtmin), at baseline conditions and under increasing doses of dobutamine.
As expected, in normal mice we observed a dobutamine dose-related increase in HR, dP/dtmax and dP/dtmin. Moreover, at the highest dose tested (12 ng/g/min) the cardiac decompensation of high fat diet-induced obese mice was unmasked.

We describe the protocol to perform a cardiac stress test induced by dobutamine and monitored by cardiac catheterization in normal mice. Also we show its application to unmask subclinical cardiac disease in high fat diet-induced obese mice.

多巴酚丁胺诱导的心脏压力测试和监控通过在小鼠心脏导管插入术

Dobutamine is a &beta;-adrenergic agonist with an affinity higher for receptor expressed in the heart (&beta;1) than for receptors expressed in the ...

科研

医学

Direct Pressure Monitoring Accurately Predicts Pulmonary Vein Occlusion During Cryoballoon Ablation

Cryoballoon ablation (CBA) is an established therapy for atrial fibrillation (AF). Pulmonary vein (PV) occlusion is essential for achieving antral contact and PV isolation and is typically assessed by contrast injection. We present a novel method of direct pressure monitoring for assessment of PV occlusion.
Transcatheter pressure is monitored during balloon advancement to the PV antrum. Pressure is recorded via a single pressure transducer connected to the inner lumen of the cryoballoon. Pressure curve characteristics are used to assess occlusion in conjunction with fluoroscopic or intracardiac echocardiography (ICE) guidance. PV occlusion is confirmed when loss of typical left atrial (LA) pressure waveform is observed with recordings of PA pressure characteristics (no A wave and rapid V wave upstroke). Complete pulmonary vein occlusion as assessed with this technique has been confirmed with concurrent contrast utilization during the initial testing of the technique and has been shown to be highly accurate and readily reproducible.
We evaluated the efficacy of this novel technique in 35 patients. A total of 128 veins were assessed for occlusion with the cryoballoon utilizing the pressure monitoring technique; occlusive pressure was demonstrated in 113 veins with resultant successful pulmonary vein isolation in 111 veins (98.2%). Occlusion was confirmed with subsequent contrast injection during the initial ten procedures, after which contrast utilization was rapidly reduced or eliminated given the highly accurate identification of occlusive pressure waveform with limited initial training.
Verification of PV occlusive pressure during CBA is a novel approach to assessing effective PV occlusion and it accurately predicts electrical isolation. Utilization of this method results in significant decrease in fluoroscopy time and volume of contrast.

Effective pulmonary vein isolation utilizing a cryoballoon depends on complete pulmonary vein occlusion. The point of occlusion can be effectively predicted by direct analysis of pulmonary vein pressure waveform analysis during balloon inflation using a simple and reproducible technique.

直接压力监测准确预测在Cryoballoon消融肺静脉阻塞

Cryoballoon  ablation (CBA) is an established therapy for atrial fibrillation (AF).  Pulmonary vein (PV) occlusion is essential for achieving antral ...

A Model of Cardiac Remodeling Through Constriction of the Abdominal Aorta in Rats

Heart failure is one of the leading causes of death worldwide. It is a complex clinical syndromethat includes fatigue, dyspnea, exercise intolerance, and fluid retention. Changes in myocardial structure, electrical conduction, and energy metabolism develop with heart failure, leading to contractile dysfunction, increased risk of arrhythmias, and sudden death. Hypertensive heart disease is one of the key contributing factors of cardiac remodeling associated with heart failure. The most commonly-used animal model mimicking hypertensive heart disease is created via surgical interventions, such as by narrowing the aorta. Abdominal aortic constriction is a useful experimental technique to induce a pressure overload, which leads to heart failure. The surgery can be easily performed, without the need for chest opening or mechanical ventilation. Abdominal aortic constriction-induced cardiac pathology progresses gradually, making this model relevant to clinical hypertensive heart failure. Cardiac injury and remodeling can be observed 10 weeks after the surgery. The method described here provides a simple and effective approach to produce a hypertensive heart disease animal model that is suitable for studying disease mechanisms and for testing novel therapeutics.

A rat model of abdominal aortic constriction that induces cardiac hypertrophy and remodeling is described. An efficient, highly-reproducible, and minimally-invasive method is used to provide a simple yet useful platform for research in myocardial hypertrophy and dysfunction.

心脏重构通过大鼠腹主动脉缩窄的模型

Heart failure is one of the leading causes of death worldwide. It is a complex clinical syndromethat includes fatigue, dyspnea, exercise intolerance, and ...

Preparation and In Vitro Characterization of Magnetized miR-modified Endothelial Cells

To date, the available surgical and pharmacological treatments for cardiovascular diseases (CVD) are limited and often palliative. At the same time, gene and cell therapies are highly promising alternative approaches for CVD treatment. However, the broad clinical application of gene therapy is greatly limited by the lack of suitable gene delivery systems. The development of appropriate gene delivery vectors can provide a solution to current challenges in cell therapy. In particular, existing drawbacks, such as limited efficiency and low cell retention in the injured organ, could be overcome by appropriate cell engineering (i.e., genetic) prior to transplantation. The presented protocol describes the efficient and safe transient modification of endothelial cells using a polyethyleneimine superparamagnetic magnetic nanoparticle (PEI/MNP)-based delivery vector. Also, the algorithm and methods for cell characterization are defined. The successful intracellular delivery of microRNA (miR) into human umbilical vein endothelial cells (HUVECs) has been achieved without affecting cell viability, functionality, or intercellular communication. Moreover, this approach was proven to cause a strong functional effect in introduced exogenous miR. Importantly, the application of this MNP-based vector ensures cell magnetization, with accompanying possibilities of magnetic targeting and non-invasive MRI tracing. This may provide a basis for magnetically guided, genetically engineered cell therapeutics that can be monitored non-invasively with MRI.

Preparation and In Vitro Characterization of Magnetized miR-modified Endothelial Cells

This manuscript describes the efficient, non-viral delivery of miR to endothelial cells by a PEI/MNP vector and their magnetization. Thus, in addition to genetic modification, this approach allows for magnetic cell guidance and MRI detectability. The technique can be used to improve the characteristics of therapeutic cell products.

制备和<em&gt;体外</em&gt;磁化的表征的miR-改性内皮细胞

To date, the available surgical and pharmacological treatments for cardiovascular diseases (CVD) are limited and often palliative. At the same time, gene ...

Cardiac Magnetic Resonance for the Evaluation of Suspected Cardiac Thrombus: Conventional and Emerging Techniques

We present the conventional cardiac magnetic resonance (CMR) protocol for evaluating a suspected thrombus and highlight emerging techniques.&#160;The appearance of a mass on certain magnetic resonance (MR) sequences can help differentiate a thrombus from competing diagnoses such as a tumor. T1 and T2 signal characteristics of a thrombus are related to the evolution of hemoglobin properties. A thrombus typically does not enhance following contrast administration, which also helps differentiation from a tumor. We also highlight the emerging role of T1 mapping in the evaluation of a thrombus, which can add another level of support in diagnosis. Prior to any CMR exam, patient screening and interviews are critical to ensure safety and to optimize patient comfort. Effective communication during the exam between the technologist and the patient promotes proper breath holding technique and higher quality images. Volumetric post processing and structured reporting are helpful to ensure that the radiologist answers the ordering services&#39; question and communicates these results effectively. Optimal pre-MR safety evaluation, CMR exam execution, and post exam processing and reporting allow for delivery of high quality radiological service in the evaluation of a suspected cardiac thrombus.

The goal of this article is to describe how cardiac magnetic resonance can be used for the evaluation and diagnosis of a suspected cardiac thrombus. The method presented will describe data acquisition as well as the pre-procedure and post-procedure protocol.

心脏磁共振评估疑似心脏血栓:常规和新兴技术

We present the conventional cardiac magnetic resonance (CMR) protocol for evaluating a suspected thrombus and highlight emerging techniques.&#160;The ...

Isolation and Purification of Murine Cardiac Pericytes

Pericytes, perivascular cells of microvessels and capillaries, are known to play a part in angiogenesis, vessel stabilization, and endothelial barrier integrity. However, their tissue-specific functions in the heart are not well understood. Moreover, there is currently no protocol utilizing readily accessible materials to isolate and purify pericytes of cardiac origin. Our protocol focuses on using the widely used mammalian model, the mouse, as our source of cells. Using the enzymatic digestion and mechanical dissociation of heart tissue, we obtained a crude cell mixture that was further purified by fluorescence activating cell sorting (FACS) by a plethora of markers. Because there is no single unequivocal marker for pericytes, we gated for cells that were CD31-CD34-CD45-CD140b+NG2+CD146+. Following purification, these primary cells were cultured and passaged multiple times without any changes in morphology and marker expression. With the ability to regularly obtain primary murine cardiac pericytes using our protocol, we hope to further understand the role of pericytes in cardiovascular physiology and their therapeutic potential.

We have optimized a protocol to isolate and purify murine cardiac pericytes for basic research and investigation of their biology and therapeutic potential.

鼠心围细胞的分离与纯化

Pericytes, perivascular cells of microvessels and capillaries, are known to play a part in angiogenesis, vessel stabilization, and endothelial barrier ...

An Intact Pericardium Ischemic Rodent Model

This protocol has shown that the pericardium and its contents play an essential anti-fibrotic role in the ischemic rodent model (coronary ligation to induce myocardial injury). The majority of pre-clinical myocardial infarction models require the disruption of pericardial integrity with loss of the homeostatic cellular milieu. However, recently a methodology has been developed by us to induce myocardial infarction, which minimizes pericardial damage and retains the heart&#39;s resident immune cell population. An improved cardiac functional recovery in mice with an intact pericardial space following coronary ligation has been observed. This method provides an opportunity to study inflammatory responses in the pericardial space following myocardial infarction. Further development of the labeling techniques can be combined with this model to understand the fate and function of pericardial immune cells in regulating the inflammatory mechanisms that drive remodeling in the heart, including fibrosis.

This protocol outlines the steps for inducing myocardial infarction in mice while preserving the pericardium and its contents.

完整的心包缺血性啮齿动物模型

This protocol has shown that the pericardium and its contents play an essential anti-fibrotic role in the ischemic rodent model (coronary ligation to ...

Robotized Testing of Camera Positions to Determine Ideal Configuration for Stereo 3D Visualization of Open-Heart Surgery

Stereo 3D video from surgical procedures can be highly valuable for medical education and improve clinical communication. But access to the operating room and the surgical field is restricted. It is a sterile environment, and the physical space is crowded with surgical staff and technical equipment. In this setting, unobscured capture and realistic reproduction of the surgical procedures are difficult. This paper presents a method for rapid and reliable data collection of stereoscopic 3D videos at different camera baseline distances and distances of convergence. To collect test data with minimum interference during surgery, with high precision and repeatability, the cameras were attached to each hand of a dual-arm robot. The robot was ceiling-mounted in the operating room. It was programmed to perform a timed sequence of synchronized camera movements stepping through a range of test positions with baseline distance between 50-240 mm at incremental steps of 10 mm, and at two convergence distances of 1100 mm and 1400 mm. Surgery was paused to allow 40 consecutive 5-s video samples. A total of 10 surgical scenarios were recorded.

The human depth perception of 3D stereo videos depends on the camera separation, point of convergence, distance to, and familiarity of the object. This paper presents a robotized method for rapid and reliable test data collection during live open-heart surgery to determine the ideal camera configuration.

相机位置的机器人化测试，以确定心脏直视手术立体3D可视化的理想配置

Stereo 3D video from surgical procedures can be highly valuable for medical education and improve clinical communication. But access to the operating room ...

Large-Animal Model of Donation after Circulatory Death and Normothermic Regional Perfusion for Cardiac Assessment

The increase in demand for cardiac transplantation throughout the years has fueled interest in donation after circulatory death (DCD) to expand the organ donor pool. However, the DCD process is associated with the risk of cardiac tissue injury due to the inevitable period of warm ischemia. Normothermic regional perfusion (NRP) allows for an in situ organ assessment, allowing the procurement of hearts determined to be viable. Here, we describe a clinically relevant large-animal model of DCD followed by NRP. Circulatory death is established in anesthetized pigs by stopping mechanical ventilation. After a preset warm ischemia period, an extracorporeal membrane oxygenator (ECMO) is used for a NRP period lasting at least 30 min. During this reperfusion period, the model allows the collection of various myocardial biopsies and blood samples for initial cardiac evaluation. Once NRP is weaned, biochemical, hemodynamic, and echocardiographic assessments of cardiac function and metabolism can be performed before organ procurement. This protocol closely simulates the clinical scenario previously described for DCD and NRP in heart transplantation and has the potential to facilitate studies aimed at decreasing ischemia-reperfusion injury and enhance cardiac functional preservation and recovery.

The protocol describes a large-animal (porcine) model of donation after circulatory death, followed by thoracoabdominal normothermic regional perfusion that closely simulates the clinical scenario in heart transplantation, and has the potential to facilitate therapeutic studies and strategies.

用于心脏评估的循环死亡后捐献和正常区域灌注的大型动物模型

The increase in demand for cardiac transplantation throughout the years has fueled interest in donation after circulatory death (DCD) to expand the organ ...

Analysis of Cardiac Contractile Dysfunction and Ca2+ Transients in Rodent Myocytes

Contractile dysfunction and Ca2+ transients are often analyzed at the cellular level as part of a comprehensive assessment of cardiac-induced injury and/or remodeling. One approach for assessing these functional alterations utilizes unloaded shortening and Ca2+ transient analyses in primary adult cardiac myocytes. For this approach, adult myocytes are isolated by collagenase digestion, made Ca2+ tolerant, and then adhered to laminin-coated coverslips, followed by electrical pacing in serum-free media. The general protocol utilizes adult rat cardiac myocytes but can be readily adjusted for primary myocytes from other species. Functional alterations in myocytes from injured hearts can be compared to sham myocytes and/or to in vitro therapeutic treatments. The methodology includes the essential elements needed for myocyte pacing, along with the cell chamber and platform components. The detailed protocol for this approach incorporates the steps for measuring unloaded shortening by sarcomere length detection and cellular Ca2+ transients measured with the ratiometric indicator Fura-2 AM, as well as for raw data analysis.

Analysis of Cardiac Contractile Dysfunction and Ca2+ Transients in Rodent Myocytes

A set of protocols are presented that describe the measurement of contractile function via sarcomere length detection along with calcium (Ca2+) transient measurement in isolated rat myocytes. The application of this approach for studies in animal models of heart failure is also included.

啮齿动物心肌细胞心脏收缩功能障碍和Ca2+ 瞬变分析

Contractile dysfunction and Ca2+ transients are often analyzed at the cellular level as part of a comprehensive assessment of cardiac-induced injury ...