我们感谢赵温迪，她在严格审查的支持。这项工作是支持博士伦玻璃体视网膜团契（HM），美国国家眼科研究所资助EY014104（JWM），防盲研究基金会（DGV），狮子会眼科研究基金（DGV），以及Yeatts家庭（JWM的慷慨捐赠和DGV）。

在所有动物实验均按照研究协会在视觉与眼科声明在眼科和视觉研究中使用动物进行的，而准则和条例规定由马萨诸塞州眼耳医院动物保健委员会。 1。预处理<ol><li>麻醉8周龄小鼠腹腔注射60毫克的混合物/公斤氯胺酮和6毫克/公斤甲苯噻嗪。 </li><li>剪下胡须。这样做是因为他们具有可视化和器械放置造成干扰。 </li><li>瞳孔扩张，用5％去氧肾上腺素和0.5％托吡卡胺。 </li><li>切纤毛。这也是做是因为他们具有可视化和仪器放置干扰。 </li><li>应用局部麻醉（0.5％盐酸丙美卡因滴眼液）。 </li></ol> 2。手术在显微镜下<ol><li>设置鼠标在横向POS银行足球比赛用鼻子向外科医生。脚趾捏进行确认手术麻醉，无菌手套前，手术开始穿上。 </li><li>切开颞结膜在后缘和从巩膜分离结膜。避免穿透巩膜。经过这一步，把握结膜角膜缘用钳子来控制眼球。 </li><li>使用30号针头的斜面的顶端自密封巩膜切口朝上。使巩膜隧道穿过巩膜和穿透巩膜进入脉络膜（ 图1）。避免穿透视网膜。如果发生脉络膜出血，并配备了经过巩膜伤口，执行便秘的外科长矛，直到出血停止。这巩膜伤口会自我封闭与眼压。 </li><li>穿破角膜与30号针头，以降低眼压。使用并行针虹膜避免Ë期间伤及虹膜和晶状体n请进入前房。眼内压会导致伤口自密封。 </li><li>插入33号针头连接到汉密尔顿10微升注射器进入视网膜下的空间与锥朝下（朝向内眼），并轻轻地注入3.5微升的透明质酸钠来从基础RPE脱离的感觉神经视网膜。避免穿透视网膜用33号针头，因为透明质酸钠将进入玻璃体空间，但不是视网膜下腔，如果视网膜被击穿。此外，避免快速注射，因为它会创建一个口头撕裂，和透明质酸钠会泄漏进入前房。 </li><li>让水状液从角膜穿刺流出通过按压角膜周围的角膜穿刺用钳子来调节眼压。 </li><li>采用外科矛确认从巩膜伤口没有泄漏。 </li><li>以降低透明质酸钠泄漏的危险，放置于氰基丙烯酸酯手术胶水巩膜伤口。 </li><li>重新将结膜使用氰基丙烯酸酯胶手术原来的位置。这将进一步降低透明质酸钠泄漏的风险。 </li><li>用玻璃盖检查眼底，并确认创建一个大疱性路无视网膜下出血。 </li></ol> 3。治疗后<ol><li>适用杆菌肽抗生素软膏的眼睛，以减少感染的风险。 </li><li>继续加热垫的鼠标，以防止麻醉（氯胺酮和甲苯噻嗪的混合物）造成低体温随后低血压。 </li><li>把老鼠回动物笼，他们从麻醉醒来后。监控每日的老鼠并发症。如果发生任何并发症出现，安乐死如下所述的小鼠。 </li></ol> 4。牺牲<ol><li>腹腔注射100毫克后颈椎脱位安乐死小鼠/公斤戊巴比妥钠和enuclea忒眼睛在每个实验的适当的时间点（未示出）。避免眼球摘除期间应用对眼睛的压力，因为透明质酸钠可能会泄漏出来。大疱性类的RD应至少14天是持久的。 </li></ol>

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马萨诸塞州眼耳医院拥有三项美国专利涉及使用维替泊芬的所有权权益。此外，马萨诸塞州眼耳医院在针对视网膜脉络膜新生血管的黄斑变性等疾病的治疗的选择性破坏某些专利申请的所有权权益。马萨诸塞州眼耳医院接受特许权使用费为这些专利和专利申请的结果，米勒博士接受按照马萨诸塞州眼耳医院的体制专利政策和程序，其中包括专利权共享的规定相同份额。笔者琼W·米勒博士，是过去的董事会成员和爱尔康公司的前股东，目前担任顾问的爱尔康，imagen画质生物科技有限公司; Isis制药，Kalvista制药，Regeneron公司制药公司和ONL治疗，有限责任公司。德米特里G. Vavvas博士担任作为顾问，罗氏，基因泰克，卡拉药品。

几种方法在啮齿动物的眼睛建立了RD模式已报告3-15，22。其中大多数是利用视网膜下注射透明质酸钠，因为它是在人类中的眼内手术中常用的粘性材料，并且它不与任何已知的眼毒性1-15相关联。透明质酸钠，而不是生理盐水或磷酸盐缓冲盐水（PBS）中，增大了道的持续时间。 该方法为视网膜下注射透明质酸钠使用以下两种方法之一:transvitreal方法3-6或巩膜途径7-15。这两种方法都具有观察眼底的执行。在transvitreal方法中，视网膜下喷射器导入玻璃体腔，视网膜切开一个设置在周边视网膜创建的，而透明质酸钠注射到视网膜下腔。在该方法中，2视网膜裂孔制成，这增加了视网膜的风险升出血可能进入视网膜下腔。此外，还有透镜损伤的创建视网膜切开时的危险。有用于经巩膜方法几种改性的方法。在大多数这些方法7-12，降低眼内压与前房穿刺，一个30号针头连接到填充有透明质酸钠的注射器直接通过结膜，巩膜，脉络膜，视网膜色素上皮和插入到视网膜下腔后。透明质酸钠，然后注入到视网膜下腔。使用这种方法经巩膜视网膜撕裂和透镜损伤的风险小于使用transvitreal方法。然而，通过一个30号针头制成在巩膜的孔大，尤其是对小鼠的眼睛，和透明质酸钠注射到视网膜下腔容易漏出的眼睛透过巩膜创口。这导致较低的，更少持久RD和多个可变光感受器细胞死亡。此外，如果脉络膜出血发生在巩膜穿孔工序中，出血会蔓延到视网膜下的空间，因为在眼内压力已降低视网膜下注射透明质酸之前。 几种因素可以影响RD的作用，对视网膜脱落，包括视网膜下出血，高度和RD 16-21的持久性。与RD 16，17的高度增加，并视细胞损伤光感受器细胞死亡的增加可能是更广泛的，因为氧气和从具有较高RD相比浅RD脉络膜必需的营养物质的还原扩散。视网膜下出血也是感光细胞毒性18-21;这种毒性在视网膜脱离的可能机制包括缺氧和通过视网膜下出血代谢受阻的扩散阻挡层，并直接神经毒性诱导的血液成分（如铁）。透镜损伤，已报道对视网膜的保护作用NAL神经节细胞23，也可能诱导RD后会影响感光细胞死亡。此外，如果该条目伤口部位不密封，透明质酸钠可以摘除术中泄漏出与眼睛操纵。这可能会导致RD浅的误分类，进而可能影响结果的解释。 我们修改了经巩膜方法为视网膜下注射透明质酸钠，以提高该RDS的再现性和减少视网膜下出血率。此协议的关键步骤是创建使用30号针头，防止透明质酸钠泄漏注射后自密封巩膜切口。不同于以往的方式，执行不带观察眼底的这个协议，所以更加重视对巩膜伤口。胶应用还可以防止透明质酸钠泄漏出的眼睛。根据我们的经验，视网膜下出血与此协议瓦特率如显著小于与其它协议。如果在巩膜切口步骤发生脉络膜出血，它将退出眼睛穿过巩膜伤口，因为这一步是降低眼内压之前执行。如果脉络膜出血降低眼压和视网膜神经上皮层脱离后确实发生，血液会剖析到视网膜下腔。我们发现这发生在约5％的情况下在相反的约10-20％的其他技术。这些动物应该被排除在分析之外。 这种技术也可以用于视网膜下注射载体介导的基因转移到靶照相感光体或RPE细胞24,25。因为典型的载体（PBS，生理盐水），这些注射是显著比透明质酸钠粘度较低，标准技术由更多泄漏的困扰。本文中所描述的技术中，通过降低这种风险，使得载体转移实验更具有可重复性和可靠。

A correction was made to <a href="http://www.jove.com/video/50660/">Retinal Detachment Model in Rodents by Subretinal Injection of Sodium Hyaluronate</a>. The footnotes were removed from the abstract due to redundancy in the introduction.

Erratum: Retinal Detachment Model in Rodents by Subretinal Injection of Sodium Hyaluronate

当光感受器是从底层视网膜色素上皮分离感光细胞死亡和随后的视觉衰退发生。感光体的物理分离被认为是在各种视网膜疾病，包括年龄相关性黄斑变性（AMD），中心性浆液性脉络膜视网膜病变，糖尿病性视网膜病和早产儿视网膜病，以及孔源性（ 即所引起的视网膜休息）视网膜脱离（ RD）。视网膜下注射透明质酸钠是一种被广泛接受的模型来建立，导致光感受器细胞死亡，提供洞察光感受器变性1-15的病理生理的路。 感光细胞损伤视网膜下注射透明质酸钠，先于2001年7引入，有一个合理的时间过程的优势（几天到几周）。但是，它可以具有从动物感光细胞的损失灵魂的显著变异升由于影响RD后的感光细胞死亡两个主要因素:1）高度和RD的持续时间，和视网膜下出血的2）的发生。有一个陡峭的技术学习曲线，有助于这两个因素的方法。感光细胞的变性的增加与RD的高度，如视网膜色素上皮之间的距离（RPE）层和光感受器增加16-17。根据这些报告，我们以前的实验中表现出更多的光感受器的死亡大疱的RD比浅的RD。还已经报道，视网膜下出血为感光细胞毒性和影响感光细胞死亡18-21。同样，我们观察到更多的光感受器死亡的RD与视网膜下出血的RD比无视网膜下出血。因此，技术，以减少可变性必须专注于实现的RD一致的高度，同时避免视网膜下出血。 我们修改后的方法ducing RD能在眼睛视网膜下出血很少发生的同一位置使重现性大疱性和持续性研发。我们使用的时间的方法，因为它更容易实现更宽的手术视野与其他位点相比，进行手术。后的结膜切口，自密封巩膜切口是用30号针头进行。巩膜隧道创建，其次是巩膜渗透到脉络膜。如果在此步骤中脉络膜出血，出血将退出眼穿过巩膜创口和出血可以通过便秘停止用外科矛。然后，从角膜进行前房穿刺降低眼压。这是一个重要的步骤，因为视网膜下注射单独将导致与所得视网膜动脉阻塞和视网膜内层缺血眼压增高。 A 33号针头连接到汉密尔顿10微升注射器，然后插入到视网膜下腔，和3.5μ，L透明质酸钠注射轻轻分离从底层的视网膜色素上皮视网膜神经上皮层。相反，且根据眼底观察进行诱导道的其他方法，在直接观察下进行该工艺。由于巩膜创口是自密封，透明质酸钠不会在注射后会泄漏出来。最后，胶被置于巩膜的伤口，和结膜被重新附到原来的位置。这些持续时间的步骤也降低透明质酸钠泄漏的风险。 3.5微升透明质酸钠注射液在8周龄的小鼠眼睛造成重现性的RDS（视网膜的50％）。在创建自密封的伤口是在我们的改性过程中最重要的步骤，因为它可以防止注入透明质酸钠泄漏出的眼睛，从而使可重复的大泡性和持久性的RD。

<table border="1">
<tbody>
 <tr>
 <td>Name of Reagent/Material</td>
 <td>Company</td>
 <td>Catalog Number</td>
 <td>Comments</td>
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 <tr>
 <td>Ketaject</td>
 <td>Phoenix</td>
 <td>2010025</td>
 <td></td>
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 <td>AnaSed</td>
 <td>LLOYD</td>
 <td>4004821</td>
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 <td>5% Phenylephrine / 0.5% Tropicamide</td>
 <td>Massachusetts Eye and Ear Pharmacy</td>
 <td></td>
 <td></td>
 </tr>
 <tr>
 <td>0.5% Proparacaine Hydrochloride Ophthalmic Solution</td>
 <td>AKORN</td>
 <td>17478-263-12</td>
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 <td>Microscope</td>
 <td>Leica</td>
 <td>MG90</td>
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 <td>30G1/2 PrecisionGlide Needle</td>
 <td>BD</td>
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 <td>Beaver-Visitec</td>
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 <td>10 Microliter Syringe</td>
 <td>Hamilton</td>
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 <td>33 gauge, 0.5 inch needle</td>
 <td>Hamilton</td>
 <td>7803-05</td>
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 <td>18x18mm Cover Glass</td>
 <td>Fisher Scientific</td>
 <td>18-548A</td>
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 </tbody>
</table>

retinal detachment model in rodents by subretinal injection of sodium hyaluronate

视网膜下注射透明质酸钠是一种被广泛接受的诱导视网膜脱离（RD）的方法。然而，RD或视网膜下出血的发生的高度和持续时间可以影响在视网膜脱离感光细胞死亡。因此，有利的是，创建可再现的RD无视网膜下出血，用于评估光感受器细胞死亡。我们修改了以前报道的方法在可重现的位置与视网膜下出血很少发生大疱性创造和持续的RD。这种改性方法的关键步骤是建立一个自密封巩膜切口，注射后，可以防止透明质酸钠渗漏到视网膜下腔。使自密封巩膜切口，将创建一个巩膜隧道，随后巩膜渗透到带30号针头的脉络膜。虽然在这一步可能会出现脉络膜上腔出血，便秘有手术矛减少脉络膜下摆率orrhage。此方法允许感光死亡涉及RD如孔源性RD，早产，糖尿病视网膜病变，中心性浆液性脉络膜视网膜病变，与年龄相关的黄斑变性（AMD）的视网膜病变疾病的更可重复的，可靠的模型。

为了评估本协议作出的RD的持久性，冷冻切片上3天，7制成，和14诱导的RD后。六只眼睛被用于每个时间点。苏木精伊红（HE）染色用于可视化的部分。所有部分呈大疱性RD接近镜头（ 图2）。一只眼睛显示对7天的视网膜下出血。没有眼中流露出感染或晶状体损伤的迹象。 <img src="/files/ftp_upload/50660/50660fig1.jpg" alt="图1" fo:content-width="4.5in" fo:src="/files/ftp_upload/50660/50660fig1highres.jpg" /> 图1。创建一个自密封巩膜切口，这示意性示出了一个正常的小鼠眼睛的剖面图像。红线表示自密封巩膜切口。巩膜隧道后巩膜渗透到了30号针头脉络膜创建一个自封闭巩膜切口。这巩膜伤口将是自密封的眼内压，注射后，可以防止透明质酸钠泄漏进入视网膜下的空间，并实现持久大疱性视网膜脱离。 <img src="/files/ftp_upload/50660/50660fig2.jpg" alt="图2" fo:content-width="5in" fo:src="/files/ftp_upload/50660/50660fig2highres.jpg" /> 图2。视网膜脱离的时间过程。这照片眼底手术后立即显示无视网膜下出血的一个大泡性视网膜脱离。冰冻切片用苏木精和伊红（HE）染色显示持续性大疱性视网膜通过至少14天没有脱离视网膜下出血。

Watch this Scientific Journal Video about Retinal Detachment Model in Rodents by Subretinal Injection of Sodium Hyaluronate at JoVE.com

Retinal Detachment Model in Rodents by Subretinal Injection of Sodium Hyaluronate

创建实验性视网膜脱离与剥离的可重复性和持续的高度，没有视网膜下出血，是研究感光细胞损失的视网膜疾病的病理生理和评估潜在的治疗干预措施非常重要。在这里，我们报告中详细这样的方法。

视网膜脱离模型鼠害由透明质酸钠的视网膜下注射

Subretinal injection of sodium hyaluronate is a widely accepted method of inducing retinal detachment (RD). However, the height and duration of RD or the ...

retinal-detachment-model-rodents-subretinal-injection-sodium

Research

JoVE Journal

Medicine

20.7K 次观看.  Massachusetts Eye and Ear Infirmary, Harvard Medical School. 创建实验性视网膜脱离与剥离的可重复性和持续的高度，没有视网膜下出血，是研究感光细胞损失的视网膜疾病的病理生理和评估潜在的治疗干预措施非常重要。在这里，我们报告中详细这样的方法。 

视频: Retinal Detachment Model in Rodents by Subretinal Injection of Sodium Hyaluronate

Subretinal Injection of Gene Therapy Vectors and Stem Cells in the Perinatal Mouse Eye

The loss of sight affects approximately 3.4 million people in the United States and is expected to increase in the upcoming years.1 Recently, gene therapy and stem cell transplantations have become key therapeutic tools for treating blindness resulting from retinal degenerative diseases. Several forms of autologous transplantation for age-related macular degeneration (AMD), such as iris pigment epithelial cell transplantation, have generated encouraging results, and human clinical trials have begun for other forms of gene and stem cell therapies.2 These include RPE65 gene replacement therapy in patients with Leber's congenital amaurosis and an RPE cell transplantation using human embryonic stem (ES) cells in Stargardt's disease.3-4 Now that there are gene therapy vectors and stem cells available for treating patients with retinal diseases, it is important to verify these potential therapies in animal models before applying them in human studies. The mouse has become an important scientific model for testing the therapeutic efficacy of gene therapy vectors and stem cell transplantation in the eye.5-8 In this video article, we present a technique to inject gene therapy vectors or stem cells into the subretinal space of the mouse eye while minimizing damage to the surrounding tissue.

This surgical technique illustrates the injection of gene therapy vectors and stem cells into the subretinal space of the mouse eye.

基因治疗的载体和干细胞视网膜下注射到老鼠的眼睛

The loss of sight affects approximately 3.4 million people in the United States and is expected to increase in the upcoming years.1 Recently, gene therapy ...

科研

医学

Performing Subretinal Injections in Rodents to Deliver Retinal Pigment Epithelium Cells in Suspension

The conversion of light into electrical impulses occurs in the outer retina and is accomplished largely by rod and cone photoreceptors and retinal pigment epithelium (RPE) cells. RPE provide critical support for photoreceptors and death or dysfunction of RPE cells is characteristic of age-related macular degeneration (AMD), the leading cause of permanent vision loss in people age 55 and older. While no cure for AMD has been identified, implantation of healthy RPE in diseased eyes may prove to be an effective treatment, and large numbers of RPE cells can be readily generated from pluripotent stem cells. Several interesting questions regarding the safety and efficacy of RPE cell delivery can still be examined in animal models, and well-accepted protocols used to inject RPE have been developed. The technique described here has been used by multiple groups in various studies and involves first creating a hole in the eye with a sharp needle. Then a syringe with a blunt needle loaded with cells is inserted through the hole and passed through the vitreous until it gently touches the RPE. Using this injection method, which is relatively simple and requires minimal equipment, we achieve consistent and efficient integration of stem cell-derived RPE cells in between the host RPE that prevents significant amount of photoreceptor degeneration in animal models. While not part of the actual protocol, we also describe how to determine the extent of the trauma induced by the injection, and how to verify that the cells were injected into the subretinal space using in vivo imaging modalities. Finally, the use of this protocol is not limited to RPE cells; it may be used to inject any compound or cell into the subretinal space.

Here we present a community accepted protocol in multimedia format for subretinally injecting a bolus of RPE cells in rats and mice. This approach can be used for determining rescue potentials, safety profiles, and survival capacities of grafted RPE cells upon implantation in animal models of retinal degeneration.

视网膜下进行注射的鼠害提供视网膜色素上皮细胞悬浮

The conversion of light into electrical impulses occurs in the outer retina and is accomplished largely by rod and cone photoreceptors and retinal pigment ...

A Step by Step Protocol for Subretinal Surgery in Rabbits

Age related macular degeneration (AMD), retinitis pigmentosa, and other RPE related diseases are the most common causes for irreversible loss of vision in adults in industrially developed countries. RPE transplantation appears to be a promising therapy, as it may replace dysfunctional RPE, restore its function, and thereby vision.
Here we describe a method for transplanting a cultured RPE monolayer on a scaffold into the subretinal space (SRS) of rabbits. After vitrectomy xenotransplants were delivered into the SRS using a custom made shooter consisting of a 20-gauge metallic nozzle with a polytetrafluoroethylene (PTFE) coated plunger. The current technique evolved in over 150 rabbit surgeries over 6 years. Post-operative follow-up can be obtained using non-invasive and repetitive in vivo imaging such as spectral domain optical coherence tomography (SD-OCT) followed by perfusion-fixed histology.
The method has well-defined steps for easy learning and high success rate. Rabbits are considered a large eye animal model useful in preclinical studies for clinical translation. In this context rabbits are a cost-efficient and perhaps convenient alternative to other large eye animal models.

Retinal pigment epithelium (RPE) replacement strategies and gene-based therapy are considered for several retinal degenerative conditions. For clinical translation, large eye animal models are required to study surgical techniques applicable in patients. Here we present a rabbit model for subretinal surgery geared towards RPE transplantation, which is versatile and cost-efficient.

一种用于兔视网膜下手术分步协议

Age related macular degeneration (AMD), retinitis pigmentosa, and other RPE related diseases are the most common causes for irreversible loss of vision in ...

Intrarenal Injection of Escherichia coli in a Rat Model of Pyelonephritis

Pyelonephritis is a bacterial infection of the kidney and is most commonly caused by Escherichia coli. Recurrent infections can cause significant renal inflammation and fibrosis ultimately resulting in declining kidney function. Before improved clinical management and prevention of pyelonephritis can be instituted, a reliable animal model must be established in order to study the mechanisms of progression, recurrence, and therapeutic efficacy. The transurethral infection model closely mimics human pyelonephritis but exhibits considerable variation due to its reliance on urethral reflux to transport the bacteria to the kidney. Herein, a detailed surgical protocol for performing bacterial injections into the rat renal pelvis is provided and confirmed by non-invasive Magnetic Resonance Imaging (MRI). Using this protocol, animals receive direct exposure to a desired concentration of E. coli bacteria and can fully recover from the surgical procedure with adequate post-operative care. This facilitates subsequent longitudinal MRI assessments of the experimental animal models for comparison with saline (sham) controls. Using this direct delivery approach, the severity of infection is controllable and applicable for mechanistic studies of progression as well as development of novel treatment strategies.

Intrarenal Injection of Escherichia coli in a Rat Model of Pyelonephritis

This manuscript describes a rat surgical model of pyelonephritis using direct intra-renal infection by Escherichia coli into the renal pelvis. The experimental procedure can be utilized to study the pathogenesis of pyelonephritis as well as the associated inflammation and fibrosis.

肾内注射<em&gt;大肠杆菌</em&gt;在肾性肾炎大鼠模型中

Pyelonephritis is a bacterial infection of the kidney and is most commonly caused by Escherichia coli. Recurrent infections can cause significant renal ...

Isometric Contractility Measurement of the Mouse Mesenteric Artery Using Wire Myography

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and drugs. It is widely used in many studies on the physiological functions of vascular smooth muscle, the pathogenesis of vascular diseases such as hypertension, and the development of smooth muscle relaxant drugs. The mouse is a widely used model animal with a large pool of disease models and genetically modified strains. We introduced this method to measure the isometric contraction of mouse mesenteric artery in detail. A 1.4-mm segment of mouse mesenteric resistance artery was isolated and mounted on a myograph chamber by passing two steel wires through its lumen. After equilibration and normalization steps, the vessel segment was potentiated by a high-K+ solution twice prior to the contraction assay. As an example of the application of this method in drug development, we measured the relaxant effect of a novel natural substance, neoliensinine, isolated from a Chinese herb, embryos of the lotus seed (Nelumbo nucifera Gaertn.) on mouse mesenteric arteries. The vessel segments mounted on the myograph chamber were stimulated with a high-K+ solution. When the force tension reached a stable sustained phase, cumulative doses of neoliensinine were added to the chamber. We found that neoliensinine had a dose-dependent relaxant effect on smooth muscle contraction, thus suggesting that it bears potential activity against hypertension. In addition, as the vessel segment can survive at least 4 hours after mounting and maintain contractility induced by the high-K+ solution for many times, we suggest that the wire myograph system may be used for the time-consuming process of drug screening.

The wire myograph technique is used to investigate vascular smooth muscle functions and screen new drugs. We report a detailed protocol for measuring the isometric contractility of the mouse mesenteric artery and for screening new relaxants of vascular smooth muscle.

用线 Myography 测定小鼠肠系膜动脉的等轴收缩力

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and ...

Studying Left Ventricular Reverse Remodeling by Aortic Debanding in Rodents

To better understand the left ventricular (LV) reverse remodeling (RR), we describe a rodent model wherein, after aortic banding-induced LV remodeling, mice undergo RR upon removal of the aortic constriction. In this paper, we describe a step-by-step procedure to perform a minimally invasive surgical aortic debanding in mice. Echocardiography was subsequently used to assess the degree of cardiac hypertrophy and dysfunction during LV remodeling and RR and to determine the best timing for aortic debanding. At the end of the protocol, terminal hemodynamic evaluation of the cardiac function was conducted, and samples were collected for histological studies. We showed that debanding is associated with surgical survival rates of 70-80%. Moreover, two weeks after debanding, the significant reduction of ventricular afterload triggers the regression of ventricular hypertrophy (~20%) and fibrosis (~26%), recovery of diastolic dysfunction as assessed by the normalization of left ventricular filling and end-diastolic pressures (E/e&#39;&#160;and LVEDP). Aortic debanding is a useful experimental model to study LV RR in rodents. The extent of myocardial recovery is variable between subjects, therefore, mimicking the diversity of RR that occurs in the clinical context, such as aortic valve replacement. We conclude that the aortic banding/debanding model represents a valuable tool to unravel novel insights into the mechanisms of RR, namely the regression of cardiac hypertrophy and the recovery of diastolic dysfunction.

Here we describe a step-by-step protocol of surgical aorta debanding in the well-established mice model of aortic-constriction. This procedure not only allows studying the mechanisms underlying the left ventricular reverse remodeling and regression of hypertrophy but also to test novel therapeutic options that might accelerate myocardial recovery.

研究左心室反向改造由主动脉脱体在啮齿动物

To better understand the left ventricular (LV) reverse remodeling (RR), we describe a rodent model wherein, after aortic banding-induced LV remodeling, ...

Osteoarthritis Pain Model Induced by Intra-Articular Injection of Mono-Iodoacetate in Rats

The current animal models of osteoarthritis (OA) can be divided into spontaneous models and induced models, both of which aim to simulate the pathophysiological changes of human OA. However, as the main symptom in the late stage of OA, pain affects the patients&#39; daily life, and there are not many available models. The mono-iodoacetate (MIA)-induced model is the most widely used OA pain model, mainly used in rodents. MIA is an inhibitor of glyceraldehyde-3-phosphate dehydrogenase, which causes chondrocyte death, cartilage degeneration, osteophyte, and measurable changes in animal behavior. Besides, expression changes of matrix metalloproteinase (MMP) and pro-inflammatory cytokines (IL1 &#946; and TNF &#945;) can be detected in the MIA-induced model. Those changes are consistent with OA pathophysiological conditions in humans, indicating that MIA can induce a measurable and successful OA pain model. This study aims to describe the methodology of intra-articular injection of MIA in rats and discuss the resulting pain-related behaviors and histopathological changes.

This study describes the method of intra-articular injection of mono-iodoacetate in rats and discusses the resulting pain-related behaviors and histopathological changes, which provide references for future applications.

大鼠关节内注射单碘乙酸酯诱导的骨关节炎疼痛模型

The current animal models of osteoarthritis (OA) can be divided into spontaneous models and induced models, both of which aim to simulate the ...

Oxygen-Induced Retinopathy Model for Ischemic Retinal Diseases in Rodents

One of the commonly used models for ischemic retinopathies is the oxygen-induced retinopathy (OIR) model. Here we describe detailed protocols for the OIR model induction and its readouts in both mice and rats. Retinal neovascularization is induced in OIR by exposing rodent pups either to hyperoxia (mice) or alternating levels of hyperoxia and hypoxia (rats). The primary readouts of these models are the size of neovascular (NV) and avascular (AVA) areas in the retina. This preclinical in vivo model can be used to evaluate the efficacy of potential anti-angiogenic drugs or to address the role of specific genes in the retinal angiogenesis by using genetically manipulated animals. The model has some strain and vendor specific variation in the OIR induction which should be taken into consideration when designing the experiments.

Oxygen-induced retinopathy (OIR) can be used to model ischemic retinal diseases such as retinopathy of prematurity and proliferative diabetic retinopathy and to serve as a model for proof-of-concept studies in evaluating antiangiogenic drugs for neovascular diseases. OIR induces robust and reproducible neovascularization in the retina that can be quantified.

罗登斯缺血性视网膜疾病的氧诱导视网膜病变模型

One of the commonly used models for ischemic retinopathies is the oxygen-induced retinopathy (OIR) model. Here we describe detailed protocols for the OIR ...

A Murine Model of Ischemic Retinal Injury Induced by Transient Bilateral Common Carotid Artery Occlusion

Diverse vascular diseases such as diabetic retinopathy, occlusion of retinal veins or arteries and ocular ischemic syndrome can lead to retinal ischemia. To investigate pathological mechanisms of retinal ischemia, relevant experimental models need to be developed. Anatomically, a main retinal blood supplying vessel is the ophthalmic artery (OpA) and OpA originates from the internal carotid artery of the common carotid artery (CCA). Thus, disruption of CCA could effectively cause retinal ischemia. Here, we established a mouse model of retinal ischemia by transient bilateral common carotid artery occlusion (tBCCAO) to tie the right CCA with 6-0 silk sutures and to occlude the left CCA transiently for 2 seconds via a clamp, and showed that tBCCAO could induce acute retinal ischemia leading to retinal dysfunction. The current method reduces reliance on surgical instruments by only using surgical needles and a clamp, shortens occlusion time to minimize unexpected animal death, which is often seen in mouse models of middle cerebral artery occlusion, and maintains reproducibility of common retinal ischemic findings. The model can be utilized to investigate the pathophysiology of ischemic retinopathies in mice and further can be used for in vivo drug screening.

Here, we describe a mouse model of retinal ischemia by transient bilateral common carotid artery occlusion using simple sutures and a clamp. This model can be useful for understanding the pathological mechanisms of retinal ischemia caused by cardiovascular abnormalities.

由瞬态双边常见胡萝卜动脉闭塞引起的缺血性视网膜损伤的 Murine 模型

Diverse vascular diseases such as diabetic retinopathy, occlusion of retinal veins or arteries and ocular ischemic syndrome can lead to retinal ischemia. ...

Retinal Pigment Epithelium Transplantation in a Non-human Primate Model for Degenerative Retinal Diseases

Retinal pigment epithelial (RPE) transplantation holds great promise for the treatment of inherited and acquired retinal degenerative diseases. These conditions include retinitis pigmentosa (RP) and advanced forms of age-related macular degeneration (AMD), such as geographic atrophy (GA). Together, these disorders represent a significant proportion of currently untreatable blindness globally. These unmet medical needs have generated heightened academic interest in developing methods of RPE replacement. Among the animal models commonly utilized for preclinical testing of therapeutics, the non-human primate (NHP) is the only animal model that has a macula. As it shares this anatomical similarity with the human eye, the NHP eye is an important and appropriate preclinical animal model for the development of advanced therapy medicinal products (ATMPs) such as RPE cell therapy.
This manuscript describes a method for the submacular transplantation of an RPE monolayer, cultured on a polyethylene terephthalate (PET) cell carrier, underneath the macula onto a surgically created RPE wound in immunosuppressed NHPs. The fovea-the central avascular portion of the macula-is the site of the greatest mechanical weakness during the transplantation. Foveal trauma will occur if the initial subretinal fluid injection generates an excessive force on the retina. Hence, slow injection under perfluorocarbon liquid (PFCL) vitreous tamponade is recommended with a dual-bore subretinal injection cannula at low intraocular pressure (IOP) settings to create a retinal bleb. 
Pretreatment with an intravitreal plasminogen injection to release parafoveal RPE-photoreceptor adhesions is also advised. These combined strategies can reduce the likelihood of foveal tears when compared to conventional techniques. The NHP is a key animal model in the preclinical phase of RPE cell therapy development. This protocol addresses the technical challenges associated with the delivery of RPE cellular therapy in the NHP eye.

The non-human primate (NHP) is an ideal model for studying human retinal cellular therapeutics due to the anatomical and genetic similarities. This manuscript describes a method for submacular transplantation of retinal pigment epithelial cells in the NHP eye and strategies to prevent intraoperative complications associated with macular manipulation.

非人灵长类动物模型中视网膜色素上皮移植治疗退行性视网膜疾病

Retinal pigment epithelial (RPE) transplantation holds great promise for the treatment of inherited and acquired retinal degenerative diseases. These ...