Name: influenza a virus studies in a mouse model of infection
Uploaded: 2017-09-07T13:00:00
Description: Watch this Scientific Journal Video about Influenza A Virus Studies in a Mouse Model of Infection at JoVE.com

流感病毒在 LM 实验室的研究部分由纽约流感卓越中心 (NYICE) 提供, 这是 NIAID 流感研究和监测卓越中心 (CEIRS) 的成员。我们感谢温迪. 贝茨对手稿的修正表示支持。

这里描述的所有动物协议都是由机构动物保育和使用委员会 (IACUC) 和罗切斯特大学医学院和牙医学院的机构生物安全委员会 (IBC) 批准的, 并遵守与建议在指南为实验室动物的关心和用途全国研究委员会22。医学和牙科学院的 Vivarium 和实验动物医学科的设施和方案由 AAALAC 国际认证, 并符合国家法律、联邦法规和国家卫生研究院的政策。在每个机构都应采用类似的要求, 以遵守本手稿中所描述的动物...

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IAV 的小鼠模型广泛应用于体内的 IAV 病机、免疫原性和保护功效的研究。与其他动物模型 (如雪貂或豚鼠) 相比, 小老鼠可以使它们易于操作和储存。此外, 在动物成本、住房和生殖方面的便利使它们能够用于需要大量动物的前体免疫试验。值得注意的是, 由于小鼠已被用于多个研究学科, 一些分子和免疫学鼠试剂可以方便用于 IAV 研究。此外, 最小的寄主变异性和存在的免疫系统, 在进化上类?...

IAVs 被封装在Orthomyxoviridae系列3中的病毒分类。他们包含八单 RNA 分子与阴性极性3。在人类中, IAVs 导致季节性流行病和偶然大流行的重要后果, 当新的病毒在人类人口中引入4。此外, 季节性 IAVs 是高度和迅速传播之间的人产生了高的经济损失世界各地每年2,5。IAV 症状包括咳嗽、鼻塞、发热、不适、头痛、厌食和肌痛, 但病毒也会在免疫缺陷患者中产生更严重的疾病6。事实上, 世界卫生组织 (WHO) 计算, 季节性流感病毒每年造成30万至50万人死亡, 在世界各地1。目前只有两类药物被食品和药物管理局 (FDA) 批准用于人类的流感预防和治疗: 神经氨酸酶 (NA) 抑制剂 (例如, 奥司他韦) 和阻断剂的 M2 离子通道 (例如,金刚烷胺);然而, 耐药性病毒变种的出现日益引起人们的关注。因此, 疫苗接种仍然是保护人类免受 IAVs 感染的最佳医疗选择。迄今为止, 有三种类型的流感疫苗是由 FDA 授权供人使用的: 重组病毒血 (HA) 蛋白疫苗、灭活流感疫苗 (IIV) 和活减流感疫苗 (LAIV)5,7. 三种疫苗的设计目的是诱导对病毒 HA 蛋白的适应性免疫反应, 这是中和抗体对抗 IAVs 的主要目标。
一种验证的小鼠模型, 用于研究 IAV 感染在体内
动物模型被用于研究, 除其他以外, IAV 发病机制8,9,10,11, 导致疾病的病毒因素12和/或病毒传输13 ,14, 并测试新疫苗或抗病毒药物的功效9,10,15。老鼠 (小家鼠) 是最广泛使用的动物模型为 IAV 研究有几个原因: 1) 免疫系统在进化上类似于人类的存在;2) 低成本, 包括动物购买、住房和繁殖;3) 体积小, 易于操作和贮存;4) 最小主机的变异性, 以获得均匀的反应和结果;5) 大鼠生物学知识, 包括基因组序列;6) 许多可用的分子生物学和/或免疫学试剂;7) 可用的敲出 (KO) 小鼠研究某一宿主蛋白对病毒感染的贡献;和, 8) 多小鼠菌株, 可以利用, 以评估遗传基础的感染。
目前有几个老鼠菌株可用于研究 IAV在体内。年龄、免疫状况、性别、遗传背景和小鼠品系以及感染途径、剂量和病毒株均影响小鼠 IAV 感染的结果。IAV 研究中使用的最常见的小鼠菌株是 C57BL/6、balb/c/C 和最近的 DBA. 2 只小鼠, 因为它们比两个前菌株更容易 IAV 疾病16,17,18,19,20. 重要的是, 免疫应答也可以根据鼠标的应变情况而有所不同18,19,20。因此, 恢复所有关于鼠标和 IAV 应变的可用信息是非常重要的, 以选择最佳的实验方法进行。
虽然鼠标是一个良好的动物模型感染的在体内研究与 IAV, 他们有几个限制, 需要考虑的实验设计。例如, 使用小鼠进行体内研究的一个主要限制是, IAVs 不在小鼠之间传播。因而, 为传输研究, 更加被接受的动物模型 (例如, 雪貂或试验品) 使用16,17,21。此外, IAV 在小鼠和人类中的表现也存在着一些差异。与人类不同, 老鼠在 IAV 感染时不会发热;相反, 他们目前与低温16,17。在小鼠中, IAV 的复制集中在下呼吸道 (肺部) 而不是上层气管。因此, IAV 在小鼠中的毒力并不总是与人类所看到的相关联。总之, 由于优势大于有限的劣势, 鼠标代表了第一个动物模型, 用于评估流感病毒的发病机制, 免疫原性和保护功效的疫苗和抗病毒研究。此外, 在 IAV 感染的小动物模型中, 使用大型动物模型进行 IAV 研究, 并没有以前的证据, 在道德上是不能接受的。在这篇手稿中, 我们描述了如何感染小鼠鼻 (.) 与 IAV, 如何监测病毒感染的严重性和进展, 以及如何进行的实验, 以评估体液免疫反应和保护功效。

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			<td height="40" style="height:40px;width:295px;">Madin-Darby Canine Kidney (MDCK) epithelial cells</td>
			<td style="width:119px;">ATCC</td>
			<td style="width:123px;">CCL-34</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;width:295px;">Six- to eight-week-old female C57BL/6 mice</td>
			<td style="width:119px;">National Cancer Institute (NCI)</td>
			<td>01XBE</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:295px;">Turckey red blod cells</td>
			<td style="width:119px;">Biolink Inc</td>
			<td style="width:123px;"></td>
			<td style="width:121px;">Store at 4&#176;C</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:295px;">Dulbecco&#8217;s modified Eagle&#8217;s medium (DMEM)</td>
			<td style="width:119px;">Corning Cellgro</td>
			<td style="width:123px;">15-013-CV</td>
			<td style="width:121px;">Store at 4&#176;C</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:295px;">Fetal Bovine Serum (FBS)</td>
			<td style="width:119px;">Seradigm</td>
			<td style="width:123px;">1500-050</td>
			<td style="width:121px;">Store at -20&#176;C</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;width:295px;">Penicillin/Streptomycin/L-Glutamine (PSG) 100X</td>
			<td style="width:119px;">Corning</td>
			<td style="width:123px;">30-009-CI</td>
			<td style="width:121px;">Store at -20&#176;C</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:295px;">Penicillin/Streptomycin (PS) 100X</td>
			<td style="width:119px;">Corning</td>
			<td style="width:123px;">30-00-CI</td>
			<td style="width:121px;">Store at -20&#176;C</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:295px;">Bovin Albumin solution (BA)</td>
			<td style="width:119px;">Sigma-Aldrich</td>
			<td style="width:123px;">A7409</td>
			<td style="width:121px;">Store at 4&#176;C</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:295px;">Bovin Serum Albumin (BSA)</td>
			<td style="width:119px;">Sigma-Aldrich</td>
			<td style="width:123px;">A9647</td>
			<td style="width:121px;">Store at 4&#176;C</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;width:295px;">Tosylsulfonyl phenylalanyl chloromethyl ketone (TPCK)-treated trypsin</td>
			<td style="width:119px;">Sigma-Aldrich</td>
			<td style="width:123px;">T8802</td>
			<td style="width:121px;">Store at -20&#176;C</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:295px;">Neutral Buffered Formalin 10%</td>
			<td style="width:119px;">EMD</td>
			<td style="width:123px;">65346-85</td>
			<td style="width:121px;">Store at RT</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:295px;">Triton X-100</td>
			<td style="width:119px;">J.T.Baker</td>
			<td style="width:123px;">X198-07</td>
			<td style="width:121px;">Store at RT</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;width:295px;">Monoclonal Antibody anti-NP Influenza A Virus HB-65</td>
			<td style="width:119px;">ATTC</td>
			<td style="width:123px;">H16-L10-4R5</td>
			<td style="width:121px;">Store at -20&#176;C</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;width:295px;">Polyclonal rabbit anti-mouse immunoglobulins/FITC</td>
			<td style="width:119px;">Dako</td>
			<td style="width:123px;">F0261</td>
			<td style="width:121px;">Store at 4&#176;C</td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:295px;">ECL Anti-mouse IgG, Horseradish Peroxidase linked whole antibody</td>
			<td style="width:119px;">GE Healthcare</td>
			<td style="width:123px;">LNA931V/AG</td>
			<td style="width:121px;">Store at 4&#176;C</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:295px;">TMB substrate set</td>
			<td style="width:119px;">BioLegend</td>
			<td style="width:123px;">421101</td>
			<td style="width:121px;">Store at 4&#176;C</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;width:295px;">Vmax Kinetic plate reader</td>
			<td style="width:119px;">Molecular Devices</td>
			<td style="width:123px;"></td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:295px;">Dounce Tissue Grinders</td>
			<td style="width:119px;">Thomas Scientific</td>
			<td style="width:123px;">7722-7</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:295px;">Receptor destroying enzyme, RDE (II)</td>
			<td style="width:119px;">Denka Seiken Co.</td>
			<td style="width:123px;">370013</td>
			<td style="width:121px;">Store at -20&#176;C</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:295px;">Crystal Violet</td>
			<td style="width:119px;">Fisher Scienctific</td>
			<td style="width:123px;">C581-100</td>
			<td style="width:121px;">Store at RT</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:295px;">96-well Cell Culture Plate</td>
			<td style="width:119px;">Greiner Bio-one</td>
			<td style="width:123px;">655-180</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:295px;">Cell Culture dishes 100mm</td>
			<td style="width:119px;">Greiner Bio-one</td>
			<td style="width:123px;">664-160</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;width:295px;">Nunc MicroWell 96-Well Microplates</td>
			<td style="width:119px;">Thermo Fisher Scienctific</td>
			<td style="width:123px;">269620</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;width:295px;">Nunc 96-Well Polystyrene Conical Bottom MicroWell Plates</td>
			<td style="width:119px;">Thermo Fisher Scienctific</td>
			<td style="width:123px;">249570</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:295px;">Puralub Vet Ointment</td>
			<td style="width:119px;">Dechra</td>
			<td style="width:123px;">9N-76855</td>
			<td style="width:121px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:295px;">Fluorescent microscope</td>
			<td style="width:119px;">Olympus</td>
			<td>Olympus IX81</td>
			<td style="width:121px;"></td>
		</tr>
	</tbody>
</table>

influenza a virus studies in a mouse model of infection

流感病毒在全球范围内造成超过50万人死亡1 , 并与每年的费用 12-140亿 USD 在美国单独考虑直接医疗和住院费用和工作缺勤的2。动物模型是关键的流感 A 病毒 (IAV) 研究评估病毒发病机制, 宿主-病原体相互作用, 免疫反应, 以及目前和/或新的疫苗方法的功效, 以及抗病毒药物。小鼠是一种有利的小动物模型, 因为它们的免疫系统在进化上与人类相似, 它们可从商业供应商那里获得, 作为基因上相同的学科, 有多种菌株可以利用来评估感染的遗传基础, 它们相对便宜, 易于操作。为了总结 IAV 感染的人通过呼吸道, 小鼠首先被麻醉之前, 鼻腔接种与传染性 IAVs 在适当的生物安全控制下。感染后, IAVs 的发病机制是通过每日监测发病率 (体重损失) 和死亡率 (存活率) 来确定的。此外, 病毒病机还可以评估病毒复制在上 (鼻黏膜) 或较低 (肺) 呼吸道感染小鼠。对 IAV 感染的体液反应可以通过无创性出血和二级抗体检测方法快速评估, 目的是检测总或中和抗体的存在。在这里, 我们描述了常用的方法感染小鼠鼻 (i. n) 与 IAV 和评估发病机制, 体液免疫反应和保护功效。

小鼠病毒病机的特征
IAV 的发病机制与其感染引起的发病率和死亡率有关。这两个参数可以很容易地在小鼠中进行评估: IAV 发病率与受感染小鼠的体重下降有关, 存活率的百分比将表明死亡率 (图 1)。IAV 感染小鼠的体重和生存率通常在感染8、9、<sup class="xr...

Watch this Scientific Journal Video about Influenza A Virus Studies in a Mouse Model of Infection at JoVE.com

Influenza A Virus Studies in a Mouse Model of Infection

甲型流感病毒 (IAVs) 是人类重要的呼吸道病原体。为了了解 IAVs 的致病性和对新疫苗方法进行临床前测试, 需要模仿人体生理学的动物模型。在这里, 我们描述的技术, 以评估 IAV 病机, 体液反应和疫苗的效果使用小鼠感染模型。

流感病毒在小鼠感染模型中的研究

Influenza viruses cause over 500,000 deaths worldwide1 and are associated with an annual cost of 12 - 14 billion USD in the United States alone ...

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SCIENTISTS IN ACTION

Using Bioluminescent Imaging to Investigate Synergism Between Streptococcus pneumoniae and Influenza A Virus in Infant Mice

Using Bioluminescent Imaging to Investigate Synergism Between Streptococcus pneumoniae and Influenza A Virus in Infant Mice

利用生物发光成像之间调查的协同作用肺炎链球菌 A型流感病毒的婴儿小鼠

During the 1918 influenza virus pandemic, which killed approximately 50 million people worldwide, the majority of fatalities were not the result of ...

科研

免疫与感染

Influenza Virus Propagation in Embryonated Chicken Eggs

在鸡胚的流感病毒传播

Influenza infection is associated with about 36,000 deaths and more than 200,000 hospitalizations every year in the United States. The continuous ...

Co-immunoprecipitation of the Mouse Mx1 Protein with the Influenza A Virus Nucleoprotein

免疫共沉淀鼠标MX1蛋白与流感病毒核蛋白

Studying the interaction between proteins is key in understanding their function(s). A very powerful method that is frequently used to study interactions ...

Nasal Wipes for Influenza A Virus Detection and Isolation from Swine

鼻湿巾从猪流感病毒检测和​​隔离

Surveillance for influenza A viruses in swine is critical to human and animal health because influenza A virus rapidly evolves in swine populations and ...

Measuring Attachment and Internalization of Influenza A Virus in A549 Cells by Flow Cytometry

通过流式细胞仪测量附件和流感的内在病毒在A549细胞

Attachment to target cells followed by internalization are the very first steps of the life cycle of influenza A virus (IAV). We provide here a detailed ...

Neutrophil Isolation and Analysis to Determine their Role in Lymphoma Cell Sensitivity to Therapeutic Agents

中性粒细胞分离和分析，以确定他们的淋巴瘤细胞的敏感性对治疗药物的作用

Neutrophils are the most abundant (40% to 75%) type of white blood cells and among the first inflammatory cells to migrate towards the site of ...

A Miniaturized Glycan Microarray Assay for Assessing Avidity and Specificity of Influenza A Virus Hemagglutinins

小型化聚糖微阵列测定评定A型流感病毒血凝素的亲和力和特异性

Influenza A virus (IAV) hemagglutinins recognize sialic acids on the cell surface as functional receptors to gain entry into cells. Wild waterfowl are the ...

In Vitro Disassembly of Influenza A Virus Capsids by Gradient Centrifugation

In Vitro Disassembly of Influenza A Virus Capsids by Gradient Centrifugation

在甲型流感病毒的衣壳的体外拆卸梯度离心

Acid-triggered molecular processes closely control cell entry of many viruses that enter through the endocytic system. In the case of influenza A virus ...

Imaging Cell Interaction in Tracheal Mucosa During Influenza Virus Infection Using Two-photon Intravital Microscopy

双光子活体显微镜感病毒感染时气管黏膜的影像细胞相互作用

The analysis of cell-cell or cell-pathogen interaction in vivo is an important tool to understand the dynamics of the immune response to infection. ...

A Luciferase-fluorescent Reporter Influenza Virus for Live Imaging and Quantification of Viral Infection

用于病毒感染活成像和定量的路西酶-荧光报告流感病毒

Influenza A viruses (IAVs) cause human respiratory disease that is associated with significant health and economic consequences. As with other viruses, ...