Name: use of the epiairway model for characterizing long-term host-pathogen interactions
Uploaded: 2011-09-02T13:00:00
Description: Watch this Scientific Journal Video about Use of the EpiAirway Model for Characterizing Long-term Host-pathogen Interactions at JoVE.com

我们想感谢帕特里克海登（MatTek），有益的讨论和利比的佐治亚州健康科学大学为他们的EM技能的罗伯特史密斯和佩里。这项研究是由NIDCD授予DC010187到DAD

1。为EpiAirway组织准备生物安全柜<ol><li>穿着专用的白大褂，头发绑背和手套，开始层流。 5分钟后，将在内阁（移液器，小技巧，离心管等），任何一方的工具，喷洒生物安全柜内部和窗框用70％乙醇，并用干净的纸巾擦拭。用70％乙醇再次喷洒清洁的室内，并留下来干。干净的一侧移动工具和重复乙醇的过程​​。 </li><li>在内阁中的气溶胶屏障枪头和专用的移液器。要使用单独包装的无菌血清...

<ol>
	<li>Murphy, T. F., Apicella, M. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nontypeable+Haemophilus+influenzae:+a+review+of+clinical+aspects,+surface+antigens,+and+the+human+immune+response+to+infection.">Nontypeable Haemophilus influenzae: a review of clinical aspects, surface antigens, and the human immune response to infection.</a> Rev. Infect. Dis. 9, 1-15 (1987).</li><li>Murphy, T. F., Faden, H., Bakaletz, L. O., Kyd, J. M., Forsgren, A., Campos, J., Virji, M., Pelton, S. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nontypeable+Haemophilus+influenzae+as+a+pathogen+in+children.">Nontypeable Haemophilus influenzae as a pathogen in children.</a> Ped. Infect. Dis. J. 28, 43-48 (2009).</li><li>Hotomi, M., Arai, J., Billal, D. S., Takei, S., KIkeda, Y., Ogami, M., Kono, M., Beder, L. B., Toya, K., Kimura, M., Yamanaka, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nontypeable+Haemophilus+influenzae+isolated+from+intractable+acute+otitis+media+internalized+into+cultured+human+epithelial+cells.">Nontypeable Haemophilus influenzae isolated from intractable acute otitis media internalized into cultured human epithelial cells.</a> Auris Nasus Larynx. 37, 137-144 (2010).</li><li>Chemuturi, N. V., Hayden, P., Kalausner, M., Donovan, M. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparison+of+human+tracheal/bronchial+epithelial+cell+culture+and+bovine+nasal+respiratory+explants+for+nasal+drug+transport+studies.">Comparison of human tracheal/bronchial epithelial cell culture and bovine nasal respiratory explants for nasal drug transport studies.</a> J. Pharm. Sci. 94, 1976-1985 (2005).</li><li>Sharma, M., Schoop, R., Hudson, J. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+efficacy+of+Echinacea+in+a+3-D+tissue+model+of+human+airway+epithelium.">The efficacy of Echinacea in a 3-D tissue model of human airway epithelium.</a> Phytother. Res. 24, 900-904 (2010).</li><li>Sexton, K., Balharry, D., BeruBe, K. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genomic+biomarkers+of+pulmonary+exposure+to+tobacco+smoke+components.">Genomic biomarkers of pulmonary exposure to tobacco smoke components.</a> Pharmacogenet Genomics. 10, 853-860 (2008).</li><li>Babu, R. J., Dayal, P., Singh, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+cyclodextrins+on+the+complexation+and+nasal+permeation+of+melatonin.">Effect of cyclodextrins on the complexation and nasal permeation of melatonin.</a> Drug Deliv. 6, 381-388 (2008).</li><li>Balharry, D., Sexton, K., BeruBe, K. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+in+vitro+approach+to+assess+the+toxicity+of+inhaled+tobacco+smoke+components:+nicotine,+cadmium,+formaldehyde+and+urethane.">An in vitro approach to assess the toxicity of inhaled tobacco smoke components: nicotine, cadmium, formaldehyde and urethane.</a> Toxicology. 244, 66-76 (2008).</li><li>Fahy, J. V., Dickey, M. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Airway+mucus+function+and+dysfunction.">Airway mucus function and dysfunction.</a> N. Engl. J. Med. 363, 2233-2247 (2010).</li><li>Huang, Y., Mikami, F., Jono, H., Zhang, W., Weng, X., Koga, T., Xu, H., Yan, C., Kai, H., Li, J. -. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Opposing+roles+of+PAK2+and+PAK4+in+synergistic+induction+of+MUC5AC+mucin+by+bacterium+NTHi.">Opposing roles of PAK2 and PAK4 in synergistic induction of MUC5AC mucin by bacterium NTHi.</a> 359, 691-696 (2007).</li></ol>

这种方法允许的长期宿主 - 病原体相互作用的首要人权组织在气 - 液界面的呼吸生物学相关背景调查。在这里，我们使用了感染的有机体NTHI，但用这种方法可以量化的任何细菌的相互作用，不引入不可接受的细胞毒作用随着时间的推移。 EpiAirway模式也可以被用于研究病毒，药物或化学物质，影响人类的上呼吸道5 6 7 8。我们一直保持着超过40天，与NTHI感染的组织至少10天未感染的组织。我...

<table border="1"><tbody><tr><th>试剂名称</th><th>公司</th><th>目录＃ </th><th>评论</th></tr><tr><td>皂素</td><td> Calbiochem </td><td> 558255 - 25GM </td><td>在D - PBS无钙或镁的1％，过滤消毒</td></tr><tr><td> 1 + X贝科的磷酸盐缓冲生理盐水与钙和镁</td><td>龙沙</td><td> 17 - 513Q </td><td></td></tr><tr><td> 1 + X贝科的磷酸盐缓冲生理盐水无钙或镁</td><td>龙沙</td><td> 17 - 515Q </td><td></td></tr><tr><td> EpiAirway抗生素自由组织</td><td> MatTek </td><td> AIR - 100 - ABF </td><td></td></tr><tr><td> EpiAirway抗生素免费维修媒体</td><td> MatTek </td><td> AIR - 100 - MM - ABF </td><td>提供试剂盒</td></tr><tr><td> 10X磷酸盐缓冲盐溶液</td><td>电解二氧化锰</td><td> 6506 </td><td>使用前稀释成1X </td></tr><tr><td>明胶</td><td> JT贝克</td><td> 2124-01 </td><td>在1 × PBS和高压灭菌，加入终浓度为0.1％ </td></tr><tr><td> DIFCO GC中等基地（巧克力琼脂） </td><td> VWR </td><td> 90002-016 </td><td>高压灭菌36克在500毫升DDH 2 O和冷却到60 ° C </td></tr><tr><td> BBL血红蛋白（巧克力琼脂） </td><td> VWR </td><td> 90000-662 </td><td>高压灭菌10克，500毫升DDH 2 O，冷却至60 ° C和混合与GC以上中型基地</td></tr><tr><td> BD BBL IsoVitaleX浓缩（巧克力琼脂） </td><td> VWR </td><td> 90000-414 </td><td> GC中型基地和血红蛋白的混合物冷却至55℃，添加10毫升水合IsoVitaleX，倒入巧克力琼脂平板</td></tr><tr><td>解剖钳，尖端细，弯曲</td><td> VWR </td><td> 82027-406 </td><td></td></tr><tr><td>自密封的消毒袋</td><td> VWR </td><td> 89140-802 </td><td></td></tr><tr><td>硫酸庆大霉素，10毫克/毫升</td><td>龙沙</td><td> 17 - 519Z </td><td>加入10微升/毫升EpiAirway的MM庆大霉素杀死</td></tr></tbody></table>

use of the epiairway model for characterizing long-term host-pathogen interactions

Nontypeable 流感嗜血杆菌 （NTHI）是人类适应革兰氏阴性菌，可引起复发和慢性感染的呼吸道粘膜1 2。要研究的机制，这些生物生存和呼吸组织内，一个是必需的，可以进行长期成功的细菌和人类细胞的共培养模式。我们使用初级人类呼吸道上皮组织，提高气 - 液界面，EpiAirway模型（MatTek，阿什兰，马萨诸塞州）。这些都是非永生化，分化良好，3维的组织，包含紧密连接，纤毛和nonciliated细胞，杯状细胞产生的粘蛋白，并保留产生细胞因子对感染的反应能力。 有关这种生物在人类上呼吸道的体外模型可以用在许多方面;这种方法的总体目标是，随着时间的推移执行长期NTHI和定量EpiAirway组织共培养细胞相关的和内在的细菌。同时，受感染的共同文化的生产和粘蛋白，细胞因子的个人资料才能确定。这种方法提高了，许多当前的协议后，现有的方法，利用人体细胞淹没单层或Transwell小文化，这是不能够支持长时间的细菌感染超过3。例如，如果一个有机体，可以在覆媒体复制，这可能导致在宿主细胞的细胞毒性和损失不可接受的水平，逮捕了实验。 EpiAirway模型允许长期宿主 - 病原体相互作用的特性。此外，由于EpiAirway源是人体正常细胞，而不是一个永生行气管支气管，每一个实际人类上呼吸道组织的优秀代表，在结构和功能4 ，。 对于这种方法，EpiAirway组织断奶2天交货前的抗微生物和抗真菌化合物，和所有的程序都无抗生素的条件下进行。这就需要特殊的考虑，因为这两种细菌和小学的人体组织在同一生物安全柜使用，并长时间共同培养。

Watch this Scientific Journal Video about Use of the EpiAirway Model for Characterizing Long-term Host-pathogen Interactions at JoVE.com

Use of the EpiAirway Model for Characterizing Long-term Host-pathogen Interactions

此方法允许EpiAirways，初级人类呼吸道上皮组织生长在气 - 液界面，生物有关延长细菌共同培养的表征<em&gt;在体外</em&gt;车型。该方法可用于任何微生物，是经得起长期的合作文化。

使用的EpiAirway模型，用于描述长期宿主 - 病原体相互作用

Nontypeable Haemophilus influenzae  (NTHi) are human-adapted Gram-negative bacteria that can cause recurrent and chronic infections of the respiratory ...

Research

JoVE Journal

Immunology and Infection

Use of an Optical Trap for Study of Host-Pathogen Interactions for Dynamic Live Cell Imaging

动态活细胞成像的宿主 - 病原体相互作用的研究使用的光学陷阱

Dynamic live cell imaging allows direct visualization of real-time interactions between cells of the immune system1, 2; however, the lack of spatial and ...

科研

免疫与感染

A Comparative Approach to Characterize the Landscape of Host-Pathogen Protein-Protein Interactions

比较方法来表征景观的宿主 - 病原体蛋白 - 蛋白相互作用

Significant efforts were gathered to generate  large-scale comprehensive protein-protein interaction network maps. This is  instrumental to understand the ...

Long Term Chronic Pseudomonas aeruginosa Airway Infection in Mice

Long Term Chronic Pseudomonas aeruginosa Airway Infection in Mice

长期慢性<em&gt;铜绿假单胞菌</em&gt;在呼吸道感染小鼠

A mouse model of chronic airway infection is a key asset in cystic fibrosis (CF) research, although there are a number of concerns regarding the model ...

Use of Shigella flexneri to Study Autophagy-Cytoskeleton Interactions

Use of Shigella flexneri to Study Autophagy-Cytoskeleton Interactions

使用<em&gt;福氏痢疾杆菌</em&gt;研究自噬，细胞骨架相互作用

Shigella flexneri is an intracellular pathogen that can escape from phagosomes to reach the cytosol, and polymerize the host actin cytoskeleton to promote ...

Long Term Intravital Multiphoton Microscopy Imaging of Immune Cells in Healthy and Diseased Liver Using CXCR6.Gfp Reporter Mice

免疫细胞健康和疾病肝脏使用CXCR6.Gfp记者小鼠长期活体多光子显微镜成像

Liver inflammation as a response to injury is a highly dynamic process involving the infiltration of distinct subtypes of leukocytes including monocytes, ...

Evaluation of the Efficacy And Toxicity of RNAs Targeting HIV-1 Production for Use in Gene or Drug Therapy

RNA的疗效和毒性的评价基因或药物治疗靶向HIV-1的生产使用

Small RNA therapies targeting post-integration steps in the HIV-1 replication cycle are among the top candidates for gene therapy and have the potential ...

An IL-8 Transiently Transgenized Mouse Model for the In Vivo Long-term Monitoring of Inflammatory Responses

An IL-8 Transiently Transgenized Mouse Model for the In Vivo Long-term Monitoring of Inflammatory Responses

IL-8瞬时转化的小鼠模型<em&gt;体内</em&gt;长期监测炎症反应

Airway inflammation is often associated with bacterial infections and represents a major determinant of lung disease. The in vivo determination of the ...

Long-term In Vivo Tracking of Inflammatory Cell Dynamics Within Drosophila Pupae

Long-term In Vivo Tracking of Inflammatory Cell Dynamics Within Drosophila Pupae

果蝇蛹中炎症细胞动力学的长期体内追踪

During the rapid inflammatory response to tissue damage, cells of the innate immune system are quickly recruited to the injury site. Once at the wound, ...

Complementary Use of Microscopic Techniques and Fluorescence Reading in Studying Cryptococcus-Amoeba Interactions

Complementary Use of Microscopic Techniques and Fluorescence Reading in Studying Cryptococcus-Amoeba Interactions

在研究隐球菌-阿米巴相互作用中,显微技术和荧光读数的补充应用

To simulate Cryptococcus infection, amoeba, which is the natural predator of cryptococcal cells in the environment, can be used as a model for ...

Evaluation of Host-Pathogen Responses and Vaccine Efficacy in Mice

宿主病原学反应及疫苗效果评价

Vaccines are a 20th century medical marvel. They have dramatically reduced the morbidity and mortality caused by infectious diseases and contributed to a ...