Name: a reversible silicon oil-induced ocular hypertension model in mice
Uploaded: 2019-11-15T10:00:00
Description: Watch this Scientific Journal Video about A Reversible Silicon Oil-Induced Ocular Hypertension Model in Mice at JoVE.com

这项工作由NIH授予EY024932、EY023295和EY028106至YH支持。

所有程序都已获得斯坦福大学机构动物护理和使用委员会（IACUC）的批准。
1. 通过SO宫内注射诱导眼部高血压
<ol><li>通过用移液器拉拔器拉制玻璃毛细管来生成微移液器，为宫内 SO 注射准备玻璃微移液器。在微移液器尖端切一个开口，用微磨机进一步锐化尖端，以制作 35°40° 斜面。</li>
	<li>抛光斜面的边缘，用水清洗清除所有碎屑。使用前对微管进行高压灭菌。</li>
	<li>为?...

<ol>
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在这里，我们演示了一个简单但有效的程序，通过内部注入SO诱导小鼠眼睛持续IOP升高。在显微镜下有微解剖经验的人可以很快地学习这个程序。失效的主要潜在风险是角膜切口的SO泄漏。然而，使用SO的优点之一是，由于油滴是可见的和可测量的，我们可以很容易地识别接收的液滴太小，在注射后很快诱导稳定的眼高血压的小鼠，并将其排除在后续实验中。我们通常取得了80%的成功率，并排除了?...

视网膜结结细胞（RGCs）及其斧头的逐渐丧失是青光眼的标志，青光眼是视网膜1中常见的神经退行性疾病。到2040年，它将影响超过1亿40-80岁的人。IOP仍然是青光眼发展和进展中唯一可改变的风险因素。为了探索青光眼的发病机制、进展和潜在治疗，一种可靠、可重复和诱导的实验性眼高血压/青光眼模型，复制人类患者的关键特征势在必行。
IOP 取决于从后室的纤毛体流入前室的水性幽默，并在前室的角度通过眼网状物 （TM） 流出。达到稳定状态后，IOP 保持不变。当流入超过或小于流出时，IOP 将分别上升或下降。通过通过遮挡前腔室的角度或破坏TM来减少水流出，已经建立了3、4、5、6、7、8、9、10。这些模型通常与不可逆的眼组织损伤相关，前腔的高 IOP 也会导致不必要的并发症，如角膜水肿和眼内炎症，这使得视网膜成像和视觉功能测定难以执行和解释。
为了开发一个克服这些缺点的模型，我们专注于由硅油（SO）引起的二次青光眼，它作为人类视网膜手术的术后并发症发生11，12。SO因其高表面张力而被用作视网膜手术中的棉签。然而，SO可以物理遮挡学生，因为它比水液和vita流体轻，防止水流入前室。由于水性幽默积累，阻碍导致后室的IOP高程。这促使我们开发和描述基于宫内SO注射和小孔块13的新型眼高血压小鼠模型，其关键特征是继发性青光眼：有效的小毛块、在SO去除后可恢复正常的显著IOP高程，以及青光眼神经变性。
在这里，我们提出了一个详细的协议，SO诱导眼高血压在小鼠眼睛，包括SO注射和去除和IOP测量。

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		<tr height="21">
			<td height="21" style="height:21px;width:257px;">0.5% proparacaine hydrochloride</td>
			<td style="width:232px;">Akorn, Somerset</td>
			<td style="width:147px;"></td>
			<td style="width:279px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:257px;">10mL syinge</td>
			<td style="width:232px;">BD</td>
			<td style="width:147px;"></td>
			<td>Luer-Lok Tip</td>
		</tr>
		<tr height="25">
			<td height="25" style="height:26px;">18G needle</td>
			<td style="width:232px;">BD</td>
			<td style="width:147px;"></td>
			<td>with Regular Bevel, Needle Length:25.4 mm</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:257px;">2,2,2-Tribromoethanol (Avertin)</td>
			<td style="width:232px;">Fisher Scientific</td>
			<td style="width:147px;">CAS# 75-80-9</td>
			<td>50g</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:257px;">32G nano</td>
			<td style="width:232px;">BD</td>
			<td style="width:147px;">320122</td>
			<td>BD Nano Ultra Fine Pen Needle-32G 4mm</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:257px;">33G ophalmology needle</td>
			<td style="width:232px;">TSK/ VWR</td>
			<td style="width:147px;">TSK3313/ 10147-200</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:257px;">5mL syinge</td>
			<td style="width:232px;">BD</td>
			<td style="width:147px;"></td>
			<td>Luer-Lok Tip</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:257px;">AnaSed Injection (xylazine)</td>
			<td style="width:232px;">Butler Schein</td>
			<td style="width:147px;"></td>
			<td>100 mg/ml, 50 ml</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:257px;">artificial tears</td>
			<td style="width:232px;">Alcon Laboratories</td>
			<td style="width:147px;">300651431414</td>
			<td>Systane Ultra Lubricant Eye Drops</td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;">BSS PLUS Irrigating solution</td>
			<td style="width:232px;">Alcon Laboratories</td>
			<td style="width:147px;">65080050</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Dual-Stage Glass Micropipette Puller</td>
			<td style="width:232px;">NARISHIGE</td>
			<td style="width:147px;">PC-10</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:257px;">EZ-7000 Classic System</td>
			<td style="width:232px;">EZ system</td>
			<td style="width:147px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:257px;">Isoflurane</td>
			<td style="width:232px;">VetOne</td>
			<td style="width:147px;">502017</td>
			<td>isoflurane, USP, 250ml/bottle</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:257px;">IV Administration sets</td>
			<td style="width:232px;">EXELint/ Fisher</td>
			<td style="width:147px;">29081</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:257px;">KETAMINE HYDROCHLORIDE INJECTION</td>
			<td style="width:232px;">VEDCO</td>
			<td style="width:147px;">50989-996-06</td>
			<td>KETAVED 100mg/ml * 10ml</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">microgrind bevelling machine</td>
			<td style="width:232px;">NARISHIGE</td>
			<td style="width:147px;">EG-401</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:257px;">Miniature EVA Tubing</td>
			<td style="width:232px;">McMaster-Carr</td>
			<td style="width:147px;">1883T4</td>
			<td>0.05&#34; ID, 0.09&#34; OD, 10 ft. Length</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:257px;">silicon oil (SILIKON)</td>
			<td style="width:232px;">Alcon Laboratories</td>
			<td style="width:147px;">8065601185</td>
			<td>1,000 mPa.s</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:257px;">Standard Glass Capillaries</td>
			<td style="width:232px;">WPI/ Fisher</td>
			<td style="width:147px;">1B150-4</td>
			<td>4 in. (100mm) OD 1.5mm ID 0.84mm</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">TonoLab tonometer</td>
			<td style="width:232px;">Colonial Medical Supply, Finland</td>
			<td style="width:147px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:257px;">veterinary antibiotic ointment</td>
			<td style="width:232px;">Dechra Veterinary</td>
			<td style="width:147px;">1223RX</td>
			<td>BNP ophthalmic ointment, Vetropolycin</td>
		</tr>
	</tbody>
</table>

a reversible silicon oil-induced ocular hypertension model in mice

眼内压力升高 （IOP） 是青光眼的一个详细记录的危险因素。在这里，我们描述了一种在小鼠中持续诱导稳定的IOP高程的新方法，该方法模仿了在人类视网膜手术中使用硅油（SO）作为棉布剂的术后并发症。在此协议中，SO被注射到小鼠眼睛的前室，以阻止学生，防止水性幽默的流入。后室积累水性幽默，这反过来又增加了后段的IOP。单次SO注射可产生可靠、充足和稳定的IOP高程，从而诱发显著的青光眼神经退化。此模型是眼科诊所中继发性青光眼的真实复制。为了进一步模拟临床设置，SO 可以从前室中取出，以重开排水通道，并允许水性幽默的流入，水性幽默通过前室的角度通过眼网状工程 （TM） 排出。由于 IOP 很快恢复正常，该模型可用于测试降低 IOP 对青光眼视网膜性结条细胞的影响。该方法简单明了，不需要特殊设备或重复程序，密切模拟临床情况，可适用于多种动物物种。但是，可能需要稍作修改。

注射后不久，我们可以很容易地识别小鼠，不产生稳定的眼部高血压，因为SO液滴太小（±1.5毫米）13。这些动物被排除在后续实验中。按照注射程序，超过80%的SO注射小鼠最终滴大于1.6毫米。在一次SO注射后，我们每周测量一次这些小鼠眼睛的IOP，持续8周。眼睛接收SO的IOP仍然很高，一般是反向控制眼的IOP的两倍，表明有效的孔阻滞（图1）。小鼠角膜水肿?...

Watch this Scientific Journal Video about A Reversible Silicon Oil-Induced Ocular Hypertension Model in Mice at JoVE.com

A Reversible Silicon Oil-Induced Ocular Hypertension Model in Mice

在这里，我们提出了一个方案，通过硅胶油的宫内注射和从前腔中取出硅油的程序，诱导小鼠眼睛的眼内高血压和青光眼神经变性， 以返回升高的眼内压力正常。

小鼠中可逆硅油诱发眼部高血压模型

Elevated intraocular pressure (IOP) is a well-documented risk factor for glaucoma. Here we describe a novel, effective method for consistently inducing ...

青光眼是不可逆性失明的主要原因，其特征是视网膜结节细胞死亡和光神经退化。眼内压力的升高，也称为眼部高血压，是青光眼最公认的危险因素。在实验动物（如小鼠）中，一个简单而有效的眼部高血压模型对于研究神经退化中的光度计和神经保护的试验治疗非常重要。因此，通过结果，我们开发一个小鼠眼部高血压模型，密切模仿人类患者观察到的继发青光眼。该模型忠实地复制了硅油引起的二次青光眼，导致瞳孔阻塞。程序简单，IOP高程稳定，如果与眼部高血压相关的神经退化在小鼠眼睛中严重，更重要的是，眼部高血压是可逆的，因为硅油可以很容易地从小鼠眼睛中去除。演示这个程序的将是方方博士，一个来自我实验室的手术后研究员。首先，使用移液器拉拔器拉制玻璃毛细管，用于卡内硅油喷射的玻璃微管。在微管尖端切割开口，并使用微磨床斜面机锐化尖端，使斜面为 35 至 40 度。抛光斜面的边缘，用水清洗，清除所有碎屑。使用前先将微管中去，然后进行中。要为角膜进入准备准中心针，请将 32 测量针连接到诱饵锁上的 5 毫升注射器上，然后用胶带将其固定。弯曲针斜头，面朝上，在30度。要准备硅油喷油器，请将钝端 18 表针固定在 10 毫升注射器上。然后将塑料管与 18 量表针连接在一端，并根据需要通过另一端加注硅油。麻醉9至10周老的C57黑色6J小鼠后，检查对手趾捏没有反应，胡须或尾巴缺乏运动。将鼠标放在手术平台上的横向位置。注射前在角膜上涂抹一滴5%的盐酸丙酸，以降低其在手术过程中的敏感性。使用 32 量表副中心针在超时间象限（远离肢体约 5 毫米）进行入口切口。穿过角膜层约3毫米，然后穿入前室，小心不要接触镜头或虹膜。慢慢取针，通过隧道从前室释放约一到两个微升水性幽默，等待约八分钟，以进一步降低眼内压力。测量反向控制眼以确定压力。将用硅油预装的玻璃微管通过角膜隧道插入前室，斜面朝下插入虹膜表面。缓慢地推动注射器柱塞，将硅油注入前室，直到油滴覆盖大部分虹膜表面，直径约为 2.3 至 2.4 毫米。将微管留在前室中再保留 10 秒钟，然后缓慢取出。轻轻推动上眼睑关闭角膜切口，以尽量减少硅油泄漏。然后将抗生素软膏涂抹在眼部。将鼠标放在加热垫上，直到完全从麻醉中恢复过来。要从前室中去除硅油，请给鼠标麻醉，然后检查对手趾捏的缺乏反应，以确定麻醉强度和胡须或尾巴的运动不足。将鼠标放在手术平台上，用胶带将鼠标固定在横向位置。在角膜上涂抹一滴5%的盐酸丙酸，以降低其敏感性。使用预制的 32 量表准中心针，在大约 2 到 5 点钟的角膜时间象限中，在硅油滴的边缘进行两个切口。以最大速度通过一个角膜切口插入连接到灌溉溶液的 33 量表灌溉针。将另一根 33 量表排水针插入注射器，无需柱塞通过其他角膜切口，以使硅油滴在用灌溉溶液灌溉的同时退出前室。将排水针取走，然后取灌针。使用连接到汉密尔顿注射器的玻璃微管将气泡注入前室，以保持其正常深度，并按下以关闭角膜切口。在两只眼睛上涂抹抗生素软膏，将鼠标放在加热恢复垫上，直到完全从麻醉中恢复。将鼠标放入感应室。麻醉它如手稿中描述的，并在整个过程中应用人工眼泪来保持角膜湿润。等待约15分钟，让瞳孔完全扩张。根据产品说明使用吨位计测量双眼的 IOP。将测速表靠近鼠标眼，将探头尖端到小鼠角膜的距离保持约三到四毫米。按下测量按钮六次可生成一个读数，然后从每只眼睛获取三个机器生成的读数，以获得平均 IOP。对全座视网膜视网膜神经神经半薄部分进行免疫组织化学，对在硅油注射后八周内牺牲的动物进行存活轴的定量。在一次硅油注注后，每周测量一次，对大于1.6毫米的硅油滴的小鼠眼睛的眼内压力进行一次测量，为期八周。眼睛接受油的压力仍然很高，大部分是反面控制眼的两倍，表明有效的瞳孔阻塞。在另一组小鼠中，硅胶油在注射两周后从前室冲洗。等待一周后，让角膜恢复，眼内压力恢复正常，与控制眼相同。为了确定硅油引起的眼部低血压对视网膜结节细胞的影响，RBPMS染色对视网膜整体坐管周围区域中幸存的索马塔进行量化，显示硅油注射后视网膜结节细胞死亡剧烈。注射后八周的多溴二苯醚染色，对视神经半薄横截面中幸存的轴子进行量化。硅油引起的眼部高血压中轴松退化很大。动物可以使用SLOOCT成像，以确定视网膜的变化。动物的视觉功能也可以通过模式ERG和OKR进行检测。这种眼部高血压模型将有助于研究高 IOP 诱导的冰川计神经退化。它也可以用来测试有希望的神经保护疗法临床前。

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SCIENTISTS IN ACTION

Creation of Reversible Cholestatic Rat Model

可逆的胆汁淤积大鼠模型的建立

Cholestasis is a clinical condition commonly encountered by both surgeons and gastroenterologists. Cholestasis can cause various physiological changes and ...

科研

医学

5/6th Nephrectomy in Combination with High Salt Diet and Nitric Oxide Synthase Inhibition to Induce Chronic Kidney Disease in the Lewis Rat

5/6th联合切除术，高盐饮食和一氧化氮合酶抑制剂诱导Lewis大鼠慢性肾脏病

Chronic kidney disease (CKD) is a global problem. Slowing CKD progression is a major health priority. Since CKD is characterized by complex derangements ...

A Laser-induced Mouse Model of Chronic Ocular Hypertension to Characterize Visual Defects

激光诱导的慢性高眼压表征视觉缺陷的小鼠模型

Glaucoma, frequently associated with elevated intraocular pressure (IOP), is one of the leading causes of blindness. We sought to establish a mouse model ...

Carotid Artery Infusions for Pharmacokinetic and Pharmacodynamic Analysis of Taxanes in Mice

颈动脉输液的紫杉烷类化合物在小鼠体内的药代动力学和药效学分析

When proposing the use of a drug, drug combination, or drug delivery into a novel system, one must assess the pharmacokinetics of the drug in the study ...

A Simple Critical-sized Femoral Defect Model in Mice

一个简单的临界尺寸股骨缺损模型小鼠

While bone has a remarkable capacity for regeneration, serious bone trauma often results in damage that does not properly heal. In fact, one tenth of all ...

A Murine Model of Irreversible and Reversible Unilateral Ureteric Obstruction

的不可逆和可逆单侧输尿管梗阻小鼠模型

Obstruction of the kidney may affect native or transplanted kidneys and results in kidney injury and scarring. Presented here is a model of obstructive ...

A Magnetic Microbead Occlusion Model to Induce Ocular Hypertension-Dependent Glaucoma in Mice

A磁条微珠缺血模型，以诱导小鼠高眼压，青光眼相关

The use of rodent models of glaucoma has been essential to understand the molecular mechanisms that underlie the pathophysiology of this multifactorial ...

A Repetitive Concussive Head Injury Model in Mice

重复震荡颅脑损伤模型小鼠

Despite the concussion/ mild traumatic brain injury (mTBI) being the most frequent occurrence of traumatic brain injury, there is still a lack of ...

Shunt Surgery, Right Heart Catheterization, and Vascular Morphometry in a Rat Model for Flow-induced Pulmonary Arterial Hypertension

分流手术，右心导管，血管形态计量学在大鼠模型中的流量引起的肺动脉高压

In this protocol, PAH is induced by combining a 60 mg/kg monocrotalin (MCT) injection with increased pulmonary blood flow through an aorto-caval shunt ...

Corneal Epithelial Abrasion with Ocular Burr As a Model for Cornea Wound Healing

眼毛刺角膜上皮磨损与角膜创面愈合的模型研究

The murine cornea provides an excellent model to study wound healing. The cornea is the outermost layer of the eye, and thus is the first defense to ...