We would like to acknowledge Beth Levine for critically reviewing the manuscript, Valerie Tassinari for developing the genotyping protocol, and Matt Ferola and Carlos Maya for their exceptional animal care.

所有实验都按照由卫生指南的国家机构的护理和使用动物的所述的协议进行，并批准了ALS TDI的机构动物护理和使用委员会（IACUC）。 1，动物，房屋和研究设计注：在SOD1-G93A小鼠菌落从高拷贝B6SJLTgN（SOD1G93A）1Gur应变最初由格尼等7产生的派生。菌落目前正在通过杂交的转基因C57BL / 6-SJL雄性野生型C57BL / 6-SJL F1雌性保持。通过杂交SJL雄性与雌性B6产生的F1动物。小鼠被运送到ALS TDI每周在35-45日龄。 <ol><li>允许小鼠至少一周分配给一个研究之前来适应环境的动物设施（12小时亮/暗循环，在18-23℃，40-60％湿度的温度下）。 </li><li>宅男SOD1-G93A麦克风每笼ê一个鼠标，因为群养的男性往往会打。宅女SOD1-G93A小鼠长达两个每笼。提供丰富环境，塑料小屋的形式。提供食物和水随意，除非小鼠是研究需要药物施用在饮用水/食品的一部分。在这种情况下，理性，记录食物和水。 </li><li>开始围绕日龄50药物疗效研究，其中包括64产仔匹配的性别平衡的小鼠（每个队列32）。最后，确保在进行研究的研究人员正在失明的实验条件。 注意：建议，一个典型的药物功效研究开始于年龄天50但是根据治疗介入的可能是优选的启动研究较早或较晚的类型。 注：研究用32只小鼠的队列通电检测性别特异性药物效果。如果没有性别区分的比较是必需的，每个队列24只应该足以检测药物的整体效果。</li></ol> 2.体重<ol><li>记录体重为所有小鼠在研究中每天用一克的十分之一的精度。注：体重是任何不适，可能会导致慢性药物治疗的疾病进展和的敏感指标。 </li><li>小鼠体重变化相当一天的过程中，作为在早晨更高，从而每天大约在同一时间记录下来。 </li></ol> 3.神经评分系统（NeuroScore） <ol><li>评估神经学得分（NS）的每只小鼠每日通过以下三种情况下观察鼠标顺序执行：a）该小鼠是由它的尾巴悬挂，b）该小鼠被允许行走，轻瘫C的发病后）的鼠标放置在其一侧。 <ol><li>对于悬尾实验，按住鼠标大约1.5“从尾巴在他们的家笼的线顶部的基地1-2秒，远离菜盆，而OBS欧文的后肢。重复测试悬浮液3次，并记录最一致的结果。 </li><li>对于行走测试，将鼠标放在一个干净的表面提供了一些牵引力和步行距离25厘米（例如，纸巾贴，以防止打滑）。让鼠标一共75厘米（纸巾的3倍长）边走边观察其步态。 </li><li>对于“翻正反射”试验，将鼠标在其左侧或右侧，并用秒表测量才能纠正自己独立站起双方的时间。注：本次测试中仅有一个实验是必需的。 </li></ol></li><li>确定每个后肢的NS（左或右）独立地在一个范围从0到4。注意：例如评分为0，1表示左后肢NS为0，而右后肢NS为1。 </li><li>分配对应的观测（摘要表 1中提供）下面的一组神经学得分（0-4）：<ol><li>分配NS 0（预症状）如果符合下列条件：当鼠标的尾巴悬吊时，后肢呈现正常喷射也就是说，它完全伸展远离横向中线和它保持在这个位置上，持续2秒或更长的时间。当鼠标被允许行走，正常步态是观察。 </li><li>分配NS 1（首发症状）是否遵守以下规定：当鼠标的尾巴悬挂，后肢呈现异常扇， 也就是说，它被折叠或部分向外侧中线倒塌或在悬尾它颤抖或是缩回/抱住。当鼠标被允许行走，正常或稍缓慢步态是观察。 </li><li>分配NS 2（麻痹性痴呆的发病），如果符合下列条件：当鼠标的尾巴悬挂，后肢部分或完全倒塌，很多不延长。 （有可能仍然是关节运动）。当鼠标被允许行走，后肢用于向前MOT然而离子脚趾在一张90厘米的步行向下卷曲至少两次或任何部分的脚的沿笼底/表拖动。当鼠标被放置在它的左侧和右侧，它能够从两侧在10秒内权利本身。 </li><li>分配的NS 3（麻痹）如果符合下列条件：当鼠标的尾巴悬吊，有刚性的麻痹的后肢或最小关节运动。当鼠标被允许行走有向前然而后肢不被用于向前运动的运动。当鼠标被放置在它的左侧和右侧，它能够从两侧在10秒内权利本身。注：很少，瘫发病后，尿液中的水分可能会出现在后肢。如果不进行治疗尿水分，可能会导致尿“烧钱”和皮肤损伤。治疗尿水分被裁剪的头发，应用温水浸泡去除尿液，并轻轻擦干。如果皮肤损伤是存在适用antibio抽动软膏。 </li><li>指定NS 4（人文终点），如果符合下列条件：当鼠标的尾巴悬挂，有刚性瘫痪在后肢。当鼠标被允许行走，没有向前运动。当鼠标被放置在其左，右手边是不是能够在来自任何一方的10秒权利本身。 也就是说，没有翻正反射。 </li></ol></li></ol> 4.数据录入和分析<ol><li>期间药效研究的过程中，进入收集每只小鼠成实验室信息管理系统（LIMS）或到市售的表计算软件的数据。 注：ALS TDI开发的LIMS软件是从内部研究收集的所有数据的中央存储库，使这些数据随着时间的推移ALS TDI研究者方便地访问。 LIMS跟踪任何个人的鼠标，凭借其独特的ID，回到它的血统，体重或神经功能评分在所有研究日，包括在其一生中指出的其他任何意见。 LIMS最终产生其用于后续分析的电子表格。 </li><li>当完成一个药效研究， 即，当所有的小鼠到达人道终点，使用电子表格软件（见表材料）通过平均在左右后肢的NS来计算合并的NS每只小鼠，每天的。接着，计算平均年龄在每个NS对每个小鼠每只小鼠花费在特定的NS的天数。 </li><li>通过从电子表格软件导入数据的统计软件分析在一段时间内的NS总体变化（见表材料）。符合这些数据的使用顺序逻辑回归的模型估计预期的NS，中位年龄在NS和药物治疗作为回归的影响。使用对数秩卡方检验估计的影响似然概率，并确定在NS进展的药物治疗效果是否是统计LY显著（见一步一步的指示补充编码文件）。 注：p值&lt;0.05被认为是统计显著。 注意：统计分析的该方法尊重的NS数据并向NS轨迹单个小鼠的可变性质的序号性质。这是首选，因为个人NS轨迹可以明显不同导致NS时关联组平均数是不充分的反映NS响应。此外，在某些情况下，NS会波动，特别是当在零到一的范围内。 </li><li>最后，使用图形软件（见表材料）生成“序预计NS VS平均年龄NS”图表药物治疗组和对照组动物（ 图2，见补充编码文件一步一步的说明）。使用这些图形由y轴插补疾病发作（ 我的年龄中位数为方便评估药物治疗组和对照组之间的变化 。即，NS 2）上x轴。 注意：在两组中位年龄的差异代表药物引起的移位和被当作效果幅度的实际措施。 注：在预期NS与年龄中位数曲线表明减缓症状的疾病进展右移。相反，在预期的NS对位年龄曲线向左偏移指示加速症状的疾病的进展。 </li></ol>

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The authors have nothing to disclose.

在这份报告中，我们描述了一个快速和简单的视频协议，如果应用得当，能够可靠地评估SOD1-G93A小鼠的疾病进展和确定药物引起的变化。尽管不同群体开发的表型的评分系统对SOD1-G93A小鼠8-10，他们往往不提供关于该过程足够的细节和不足进行复制。其结果，评分系统很少使用，开发他们的特定的组之外。在这份报告中，用视频详细手稿组合最大程度地减少评分含糊和跨实验者变异从而允?...

自从它在90年代中期开发的SOD1-G93A转基因小鼠模型已经肌萎缩侧索硬化症（ALS）1的最广泛使用的动物模型。这种转基因小鼠模型被基因工程化以过表达人铜/锌超氧化物歧化酶1（SOD1）基因携带有ALS相关甘氨酸至丙氨酸的突变在氨基酸93（G93A）的突变体形式。的G93A突变是许多突 ​​变的SOD1基因，它们共同化妆大约五分之一的家族性ALS例2之一。 在SOD1-G93A模式已经成为了ALS药物开发研究的一个主力，因为除了要ALS其明显的遗传链接，这概括了许多ALS在人体内的病理特点，如运动神经元丧失，肌肉逐渐萎缩无力，与最终麻痹而死亡3。虽然，这是常有用的转基因动物模型的情况下，有一种内在的双向与SOD1-G93A小鼠相关ological变化。 Scott等。已经确定，需要在药物功效的研究设计，以克服由生物变异4中创建的噪声至少24垫料匹配性别平衡小鼠的队列。大量在这些研究中与侵袭性疾病进展和需要日常监测组合需要的动物的禁止使用的精心设计的，耗时的和潜在的压力的技术，用于测量神经肌肉强度和功能（例如，肌电图，握力，旋转棒等）。相反，它强调需要一个体内筛选工具，可靠地评估了一个快速的方式SOD1-G93A小鼠的神经肌肉功能。 在ALS TDI的协议被开发，允许SOD1-G93A小鼠的神经肌肉功能的每只小鼠不到30秒，平均可靠的评估。该协议encompa小规模企业后肢功能的使用是该报告在媒体和视频描述了一个简单的神经评分系统（NeuroScore）每天评估。 NeuroScore专注于后肢功能，因为后肢赤字是最早报道的疾病在SOD1-G93A小鼠5,6神经的迹象。此外，通过ALS TDI开发的协议提供了发病轻瘫（轻微或局部麻痹），进展和麻痹的严重程度的公正评估，这是不够敏感，以确定在疾病进展的药物引起的变化。

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>B6SJLTgN(SOD1G93A)1Gur strain</td>
			<td>The Jackson Laboratory, Bar Harbor, ME</td>
			<td>002726</td>
			<td>Strain of mice used in this study</td>
		</tr>
		<tr>
			<td>Breeders</td>
			<td>Biomedical Research Models, Inc., Worcester, MA</td>
			<td></td>
			<td>Colony maintainance</td>
		</tr>
		<tr>
			<td>Scale</td>
			<td>Navigator OHAUS, Parsippany, NJ</td>
			<td>Model N14120</td>
			<td>Used to weigh mice</td>
		</tr>
		<tr>
			<td>Nutra-GelH</td>
			<td>Bio-Serv, Flemington, NJ</td>
			<td>#S4798</td>
			<td>Wet food provided to sick mice</td>
		</tr>
		<tr>
			<td>Irradiated Lab Animal Diet</td>
			<td>Harlan, Indianapolis, IN</td>
			<td>2918-111914M</td>
			<td>Chow diet</td>
		</tr>
		<tr>
			<td>Lab-grade Sani-chips</td>
			<td>Harlan Teklad, Indianapolis, IN</td>
			<td>7090</td>
			<td>Bedding</td>
		</tr>
		<tr>
			<td>JMPH&#160;</td>
			<td>SAS Institute, Inc., SAS Campus Drive, Cary, NC</td>
			<td>&#160;v10.0.2</td>
			<td>Statistical software</td>
		</tr>
		<tr>
			<td>Microsoft Excel</td>
			<td>Microsoft, One Microsoft Way, Redmond, WA</td>
			<td>Excel 2013 (v 15.0)</td>
			<td>Spreadsheet software</td>
		</tr>
		<tr>
			<td>GraphPad Prism 5</td>
			<td>GraphPad software Inc., La Jolla, CA</td>
			<td>v5.4</td>
			<td>Graphing software</td>
		</tr>
		<tr>
			<td>Mouse Huts</td>
			<td>Bio-Serv, Flemington, NJ</td>
			<td>K3272</td>
			<td>For environmental enrichment</td>
		</tr>
	</tbody>
</table>

a quick phenotypic neurological scoring system for evaluating disease progression in the sod1-g93a mouse model of als

The SOD1-G93A transgenic mouse is the most widely used animal model of amyotrophic lateral sclerosis (ALS). At ALS TDI we developed a phenotypic screening protocol, demonstrated in video herein, which reliably assesses the neuromuscular function of SOD1-G93A mice in a quick manner. This protocol encompasses a simple neurological scoring system (NeuroScore) designed to assess hindlimb function. NeuroScore is focused on hindlimb function because hindlimb deficits are the earliest reported neurological sign of disease in SOD1-G93A mice. The protocol developed by ALS TDI provides an unbiased assessment of onset of paresis (slight or partial paralysis), progression and severity of paralysis and it is sensitive enough to identify drug-induced changes in disease progression. In this report, the combination of a detailed manuscript with video minimizes scoring ambiguities and inter-experimenter variability thus allowing for the protocol to be adopted by other laboratories and enabling comparisons between studies taking place at different settings. We believe that this video protocol can serve as an excellent training tool for present and future ALS researchers.

神经功能评分数据为90男，94女未治疗的SOD1-G93A小鼠就读于ALS TDI在2014年进行了评价。结果表明，雄性小鼠通常具有比雌性小鼠更具侵略性的疾病进展，如由在NS 1的比例更大其寿命相比，女性。 NS 2和NS 3发生跨性别大致相等的频率。 NS 0被从分析中排除，因为它被认为是一个“正常”表型。 NS 4也被排除在外，因为根据定义，它只能出现一次在SOD1-G93A小鼠的生存期（ 表2和图1）。 <p c...

Watch this Scientific Journal Video about A Quick Phenotypic Neurological Scoring System for Evaluating Disease Progression in the SOD1-G93A Mouse Model of ALS at JoVE.com

A Quick Phenotypic Neurological Scoring System for Evaluating Disease Progression in the SOD1-G93A Mouse Model of ALS

This video protocol describes a sensitive, reliable, and quick method for evaluating the neuromuscular deficits in a transgenic mouse model of amyotrophic lateral sclerosis.

一个快速的表型神经评分系统在ALS的SOD1-G93A小鼠模型评估疾病进展

The SOD1-G93A transgenic mouse is the most widely used animal model of amyotrophic lateral sclerosis (ALS). At ALS TDI we developed a phenotypic screening ...

a-quick-phenotypic-neurological-scoring-system-for-evaluating-disease

Research

JoVE Journal

Medicine

19.2K 次观看.  ALS Therapy Development Institute. This video protocol describes a sensitive, reliable, and quick method for evaluating the neuromuscular deficits in a transgenic mouse model of amyotrophic lateral sclerosis.

视频: A Quick Phenotypic Neurological Scoring System for Evaluating Disease Progression in the SOD1-G93A Mouse Model of ALS

Development of a Unilaterally-lesioned 6-OHDA Mouse Model of Parkinson's Disease

The unilaterally lesioned 6-hyroxydopamine (6-OHDA)-lesioned rat model of Parkinson's disease (PD) has proved to be invaluable in advancing our understanding of the mechanisms underlying parkinsonian symptoms, since it recapitulates the changes in basal ganglia circuitry and pharmacology observed in parkinsonian patients1-4. However, the precise cellular and molecular changes occurring at cortico-striatal synapses of the output pathways within the striatum, which is the major input region of the basal ganglia remain elusive, and this is believed to be site where pathological abnormalities underlying parkinsonian symptoms arise3,5.
In PD, understanding the mechanisms underlying changes in basal ganglia circuitry following degeneration of the nigro-striatal pathway has been greatly advanced by the development of bacterial artificial chromosome (BAC) mice over-expressing green fluorescent proteins driven by promoters specific for the two striatal output pathways (direct pathway: eGFP-D1; indirect pathway: eGFP-D2 and eGFP-A2a)8, allowing them to be studied in isolation. For example, recent studies have suggested that there are pathological changes in synaptic plasticity in parkinsonian mice9,10. However, these studies utilised juvenile mice and acute models of parkinsonism. It is unclear whether the changes described in adult rats with stable 6-OHDA lesions also occur in these models. Other groups have attempted to generate a stable unilaterally-lesioned 6-OHDA adult mouse model of PD by lesioning the medial forebrain bundle (MFB), unfortunately, the mortality rate in this study was extremely high, with only 14% surviving the surgery for 21 days or longer11. More recent studies have generated intra-nigral lesions with both a low mortality rate &gt;80% loss of dopaminergic neurons, however expression of L-DOPA induced dyskinesia11,12,13,14 was variable in these studies. Another well established mouse model of PD is the MPTP-lesioned mouse15. Whilst this model has proven useful in the assessment of potential neuroprotective agents16, it is less suitable for understanding mechanisms underlying symptoms of PD, as this model often fails to induce motor deficits, and shows a wide variability in the extent of lesion17, 18.
Here we have developed a stable unilateral 6-OHDA-lesioned mouse model of PD by direct administration of 6-OHDA into the MFB, which consistently causes &gt;95% loss of striatal dopamine (as measured by HPLC), as well as producing the behavioural imbalances observed in the well characterised unilateral 6-OHDA-lesioned rat model of PD. This newly developed mouse model of PD will prove a valuable tool in understanding the mechanisms underlying generation of parkinsonian symptoms.

Development of a Unilaterally-lesioned 6-OHDA Mouse Model of Parkinson&#039;s Disease

A protocol for performing unilateral 6-OHDA lesions of the medial forebrain bundle in mice is described. This method has a low mortality rate (13.3 %) with 89% of the surviving animals showing &#62;95% loss of striatal dopamine and 90.63&#177;-4.02 % ipsiversive rotational bias towards the side of the lesion.

1单侧损毁6  - 羟基多巴胺帕金森病小鼠模型的发展

The unilaterally lesioned 6-hyroxydopamine (6-OHDA)-lesioned rat model of Parkinson's disease (PD) has proved to be invaluable in advancing our ...

科研

医学

Utilizing Transcranial Magnetic Stimulation to Study the Human Neuromuscular System

Transcranial magnetic stimulation (TMS) has been in use for more than 20 years 1, and has grown exponentially in popularity over the past decade. While the use of TMS has expanded to the study of many systems and processes during this time, the original application and perhaps one of the most common uses of TMS involves studying the physiology, plasticity and function of the human neuromuscular system. Single pulse TMS applied to the motor cortex excites pyramidal neurons transsynaptically 2 (Figure 1) and results in a measurable electromyographic response that can be used to study and evaluate the integrity and excitability of the corticospinal tract in humans 3. Additionally, recent advances in magnetic stimulation now allows for partitioning of cortical versus spinal excitability 4,5. For example, paired-pulse TMS can be used to assess intracortical facilitatory and inhibitory properties by combining a conditioning stimulus and a test stimulus at different interstimulus intervals 3,4,6-8. In this video article we will demonstrate the methodological and technical aspects of these techniques. Specifically, we will demonstrate single-pulse and paired-pulse TMS techniques as applied to the flexor carpi radialis (FCR) muscle as well as the erector spinae (ES) musculature. Our laboratory studies the FCR muscle as it is of interest to our research on the effects of wrist-hand cast immobilization on reduced muscle performance6,9, and we study the ES muscles due to these muscles clinical relevance as it relates to low back pain8. With this stated, we should note that TMS has been used to study many muscles of the hand, arm and legs, and should iterate that our demonstrations in the FCR and ES muscle groups are only selected examples of TMS being used to study the human neuromuscular system.

Transcranial magnetic stimulation (TMS) is a non-invasive tool to gain insight on the physiology and function of the human nervous system. Here, we present our TMS techniques to study cortical excitability of the upper limb and lumbar musculature.

利用经颅磁刺激，以研究人类的神经肌肉系统

Transcranial magnetic stimulation (TMS) has been in use for more than 20 years 1, and has grown exponentially in popularity over the past decade.  While ...

Use of Animal Model of Sepsis to Evaluate Novel Herbal Therapies

Sepsis refers to a systemic inflammatory response syndrome resulting from a microbial infection. It has been routinely simulated in animals by several techniques, including infusion of exogenous bacterial toxin (endotoxemia) or bacteria (bacteremia), as well as surgical perforation of the cecum by cecal ligation and puncture (CLP)1-3. CLP allows bacteria spillage and fecal contamination of the peritoneal cavity, mimicking the human clinical disease of perforated appendicitis or diverticulitis. The severity of sepsis, as reflected by the eventual mortality rates, can be controlled surgically by varying the size of the needle used for cecal puncture2. In animals, CLP induces similar, biphasic hemodynamic cardiovascular, metabolic, and immunological responses as observed during the clinical course of human sepsis3. Thus, the CLP model is considered as one of the most clinically relevant models for experimental sepsis1-3.
Various animal models have been used to elucidate the intricate mechanisms underlying the pathogenesis of experimental sepsis. The lethal consequence of sepsis is attributable partly to an excessive accumulation of early cytokines (such as TNF, IL-1 and IFN-&gamma;)4-6 and late proinflammatory mediators (e.g., HMGB1)7. Compared with early proinflammatory cytokines, late-acting mediators have a wider therapeutic window for clinical applications. For instance, delayed administration of HMGB1-neutralizing antibodies beginning 24 hours after CLP, still rescued mice from lethality8,9, establishing HMGB1 as a late mediator of lethal sepsis. The discovery of HMGB1 as a late-acting mediator has initiated a new field of investigation for the development of sepsis therapies using Traditional Chinese Herbal Medicine. In this paper, we describe a procedure of CLP-induced sepsis, and its usage in screening herbal medicine for HMGB1-targeting therapies.

Sepsis refers to a systemic inflammatory response syndrome resulting from a microbial infection, and can be simulated by a surgical technique termed cecal ligation and puncture (CLP). Here we describe a method to use CLP-induced animal model to screen medicinal herbs for therapeutic agents.

脓毒症动物模型，用于评估新型草药疗法

Sepsis refers to a systemic inflammatory response syndrome resulting from a microbial infection. It has been routinely simulated in animals by several ...

Clinical Testing and Spinal Cord Removal in a Mouse Model for Amyotrophic Lateral Sclerosis (ALS)

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder resulting in progressive degeneration of motoneurons. Peak of onset is around 60 years for the sporadic disease and around 50 years for the familial disease. Due to its progressive course, 50% of the patients die within 30 months of symptom onset. In order to evaluate novel treatment options for this disease, genetic mouse models of ALS have been generated based on human familial mutations in the SOD gene, such as the SOD1 (G93A) mutation. Most important aspects that have to be evaluated in the model are overall survival, clinical course and motor function. Here, we demonstrate the clinical evaluation, show the conduction of two behavioural motor tests and provide quantitative scoring systems for all parameters. Because an in depth analysis of the ALS mouse model usually requires an immunohistochemical examination of the spinal cord, we demonstrate its preparation in detail applying the dorsal laminectomy method. Exemplary histological findings are demonstrated. The comprehensive application of the depicted examination methods in studies on the mouse model of ALS will enable the researcher to reliably test future therapeutic options which can provide a basis for later human clinical trials.

Clinical Testing and Spinal Cord Removal in a Mouse Model for Amyotrophic Lateral Sclerosis ALS

A mouse model for amyotrophic lateral sclerosis (ALS) is examined clinically and behaviorally. As a prerequisite for an accompanying immunohistological analysis the preparation of the spinal cord is depicted in detail.

临床试验和脊髓去除在肌萎缩性侧索硬化症的小鼠模型（ALS）

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder resulting in progressive degeneration of motoneurons. Peak of onset is around 60 ...

A Simple Method of Mouse Lung Intubation

A simple procedure to intubate mice for pulmonary function measurements would have several advantages in longitudinal studies with limited numbers or expensive animal. One of the reasons that this is not done more routinely is that it is relatively difficult, despite there being several published studies that describe ways to achieve it. In this paper we demonstrate a procedure that eliminates one of the major hurdles associated with this intubation, that of visualizing the trachea during the entire time of intubation. The approach uses a 0.5 mm fiberoptic light source that serves as an introducer to direct the intubation cannula into the mouse trachea. We show that it is possible to use this procedure to measure lung mechanics in individual mice over a time course of at least several weeks. The technique can be set up with relatively little expense and expertise, and it can be routinely accomplished with relatively little training. This should make it possible for any laboratory to routinely carry out this intubation, thereby allowing longitudinal studies in individual mice, thereby minimizing the number of mice needed and increasing the statistical power by using each mouse as its own control.

This paper describes a striaghforward and efficient method of intubating mice for pulmonary function measurements or pulmonary instillation, that allows the mice to recover and be studied at later times. The procedure involves an inexpensive fiberoptic light source that directly illuminates the trachea.

一个简单的方法小鼠肺插管

A  simple procedure to intubate mice for pulmonary function measurements would  have several advantages in longitudinal studies with limited numbers or  ...

Bioluminescent Orthotopic Model of Pancreatic Cancer Progression

Pancreatic cancer has an extremely poor five-year survival rate of 4-6%. New therapeutic options are critically needed and depend on improved understanding of pancreatic cancer biology. To better understand the interaction of cancer cells with the pancreatic microenvironment, we demonstrate an orthotopic model of pancreatic cancer that permits non-invasive monitoring of cancer progression. Luciferase-tagged pancreatic cancer cells are resuspended in Matrigel and delivered into the pancreatic tail during laparotomy. Matrigel solidifies at body temperature to prevent leakage of cancer cells during injection. Primary tumor growth and metastasis to distant organs are monitored following injection of the luciferase substrate luciferin, using in vivo imaging of bioluminescence emission from the cancer cells. In vivo imaging also may be used to track primary tumor recurrence after resection. This orthotopic model is suited to both syngeneic and xenograft models and may be used in pre-clinical trials to investigate the impact of novel anti-cancer therapeutics on the growth of the primary pancreatic tumor and metastasis.

Improved understanding of pancreatic cancer biology is critically needed to enable the development of better therapeutic options to treat pancreatic cancer. To address this need, we demonstrate an orthotopic model of pancreatic cancer that permits non-invasive monitoring of cancer progression using in vivo bioluminescence imaging.

生物荧光原位模型胰腺癌进展

Pancreatic cancer has an extremely poor five-year survival rate of 4-6%.  New therapeutic options are critically needed and depend on improved ...

Flexible Colonoscopy in Mice to Evaluate the Severity of Colitis and Colorectal Tumors Using a Validated Endoscopic Scoring System

The use of modern endoscopy for research purposes has greatly facilitated our understanding of gastrointestinal pathologies. In particular, experimental endoscopy has been highly useful for studies that require repeated assessments in a single laboratory animal, such as those evaluating mechanisms of chronic inflammatory bowel disease and the progression of colorectal cancer. However, the methods used across studies are highly variable. At least three endoscopic scoring systems have been published for murine colitis and published protocols for the assessment of colorectal tumors fail to address the presence of concomitant colonic inflammation. This study develops and validates a reproducible endoscopic scoring system that integrates evaluation of both inflammation and tumors simultaneously. This novel scoring system has three major components: 1) assessment of the extent and severity of colorectal inflammation (based on perianal findings, transparency of the wall, mucosal bleeding, and focal lesions), 2) quantitative recording of tumor lesions (grid map and bar graph), and 3) numerical sorting of clinical cases by their pathological and research relevance based on decimal units with assigned categories of observed lesions and endoscopic complications (decimal identifiers). The video and manuscript presented herein were prepared, following IACUC-approved protocols, to allow investigators to score their own experimental mice using a well-validated and highly reproducible endoscopic methodology, with the system option to differentiate distal from proximal endoscopic colitis (D-PECS).

Over the past few years, new generation endoscopes have emerged as important diagnostic research aids for evaluating murine colitis and colorectal tumors. We present herein a detailed protocol for endoscopic assessment of inflammation and colorectal tumors in mice, as well as a novel scoring system that uses decimal identifiers to document the endoscopic severity of colitis and colorectal tumors.

灵活的结肠镜检查在小鼠使用经过验证的内镜评分系统来评估结肠炎和结肠直肠肿瘤的严重程度

The use of modern endoscopy for research purposes has greatly facilitated our understanding of gastrointestinal pathologies. In particular, experimental ...

Establishment and Characterization of UTI and CAUTI in a Mouse Model

Urinary tract infections (UTI) are highly prevalent, a significant cause of morbidity and are increasingly resistant to treatment with antibiotics. Females are disproportionately afflicted by UTI: 50% of all women will have a UTI in their lifetime. Additionally, 20-40% of these women who have an initial UTI will suffer a recurrence with some suffering frequent recurrences with serious deterioration in the quality of life, pain and discomfort, disruption of daily activities, increased healthcare costs, and few treatment options other than long-term antibiotic prophylaxis. Uropathogenic Escherichia coli (UPEC) is the primary causative agent of community acquired UTI. Catheter-associated UTI (CAUTI) is the most common hospital acquired infection accounting for a million occurrences in the US annually and dramatic healthcare costs. While UPEC is also the primary cause of CAUTI, other causative agents are of increased significance including Enterococcus faecalis. Here we utilize two well-established mouse models that recapitulate many of the clinical characteristics of these human diseases. For UTI, a C3H/HeN model recapitulates many of the features of UPEC virulence observed in humans including host responses, IBC formation and filamentation. For CAUTI, a model using C57BL/6 mice, which retain catheter bladder implants, has been shown to be susceptible to E. faecalis bladder infection. These representative models are being used to gain striking new insights into the pathogenesis of UTI disease, which is leading to the development of novel therapeutics and management or prevention strategies.

The ability to model urinary tract infections (UTI) is crucial in order to be able to understand bacterial pathogenesis and spawn the development of novel therapeutics. This work&#8217;s goal is to demonstrate mouse models of experimental UTI and catheter associated UTI that recapitulate and predict findings seen in humans.

建立和UTI和CAUTI表征在小鼠模型

Urinary tract infections (UTI) are highly prevalent, a significant cause of morbidity and are increasingly resistant to treatment with antibiotics. ...

Echocardiographic Approaches and Protocols for Comprehensive Phenotypic Characterization of Valvular Heart Disease in Mice

The aim of this manuscript and accompanying video is to provide an overview of the methods and approaches used for imaging heart valve function in rodents, with detailed descriptions of the appropriate methods for anesthesia, the echocardiographic windows used, the imaging planes and probe orientations for image acquisition, the methods for data analysis, and the limitations of emerging technologies for the evaluation of cardiac and valvular function. Importantly, we also highlight several future areas of research in cardiac and heart valve imaging that may be leveraged to gain insights into the pathogenesis of valve disease in preclinical animal models. We propose that using a systematic approach to evaluating cardiac and heart valve function in mice can result in more robust and reproducible data, as well as facilitate the discovery of previously underappreciated phenotypes in genetically-altered and/or physiologically-stressed mice.

This protocol provides a detailed description of the echocardiographic approach for comprehensive phenotyping of heart and heart valve function in mice.

超声心动图方法和协议心脏瓣膜疾病的综合表型特征小鼠

The aim of this manuscript and accompanying video is to provide an overview of the methods and approaches used for imaging heart valve function in ...

Evaluating the Procedure for Performing Awake Cystometry in a Mouse Model

Awake filling cystometry has been used for a long time to evaluate bladder function in freely moving mice, however, the specific methods used, vary among laboratories. The goal of this study was to describe the microsurgical procedure used to implant an intravesical tube and the experimental technique for recording urinary bladder pressure in an awake, freely moving mouse. In addition, experimental data is presented to show how surgery, as well as tubing type and size, affect lower urinary tract function and recording sensitivity. The effect of tube diameter on pressure recording was assessed in both polyethylene and polyurethane tubing with different internal diameters. Subsequently, the best performing tube from both materials was surgically implanted into the dome of the urinary bladder of male C57BL/6 mice. Twelve-hour, overnight micturition frequency was recorded in healthy, intact animals and animals 2, 3, 5, and 7 days post-surgery. At harvest, bladders were assessed for signs of swelling using gross observation and were subsequently processed for pathological analysis. The greatest extent of bladder swelling was observed on day 2 and 3, which correlated with behavioral voiding data showing significantly impaired bladder function. By day 5, bladder histology and voiding frequency had normalized. Based on the literature and evidence provided by our studies, we propose the following steps for in vivo recording of intravesical pressure and voided volume in an awake mouse: 1) Perform the surgery using an operating microscope and microsurgical tools, 2) Use polyethylene-10 tubing to minimize movement artifacts, and 3) Perform cystometry on post-operative day 5, when bladder swelling resolves.

This study describes the surgical procedures and experimental techniques for performing awake cystometry in a freely moving mouse. In addition, it provides experimental evidence to support its optimization and standardization.

评估在鼠标模型中执行唤醒膀胱测压的过程

Awake filling cystometry has been used for a long time to evaluate bladder function in freely moving mice, however, the specific methods used, vary among ...