Name: ex vivo organotypic corneal model of acute epithelial herpes simplex virus type i infection
Uploaded: 2012-11-03T13:00:00
Description: Watch this Scientific Journal Video about Ex Vivo Organotypic Corneal Model of Acute Epithelial Herpes Simplex Virus Type I Infection at JoVE.com

Den støtte fra Lions Eye Bank of Delaware Valley var uvurderlig for dette arbejde. Primært muse-monoklonalt antistof mod ICP8 var en venlig gave fra Dr. David Knipe (Harvard Medical School). Vi takker også Dr. Stephen Jennings (Drexel University College of Medicine) og Dr. Peter Laibson (Wills Eye Institute) for personer diskussioner og ekspertise.

Alle reagenser og udstyr blev indkøbt sterile eller blev steriliseret forud for proceduren. Denne artikel beskriver de trin for oprettelsen af den ex vivo model begynder med en pre-kerneløs øje. I vores eksperimenter, anvender vi friske hele øjne fra unge (8-12 wks) albinokaniner (Pel-Freez Biologicals, Rogers, AR). Derudover anvender vi humane hornhinder opnået fra den lokale Eye Bank. Hvis du udfører enucleation selv, sørge for at undgå at beskadige hornhinden eller limbus under proceduren. Kanin enuc...

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	<li>Kaye, S., Choudhary, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Herpes+simplex+keratitis.">Herpes simplex keratitis.</a> Prog. Retin. Eye Res. 25, 355-380 (2006).</li><li>Holmberg, B. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Enucleation+of+exotic+pets.">Enucleation of exotic pets.</a> Journal of Exotic Pet Medicine. 16, 88-94 (2007).</li><li>Remeijer, L., Osterhaus, A., Verjans, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Human+herpes+simplex+virus+keratitis:+the+pathogenesis+revisited.">Human herpes simplex virus keratitis: the pathogenesis revisited.</a> Ocul. Immunol. Inflamm. 12, 255-285 (2004).</li><li>Foreman, D. M., Pancholi, S., Jarvis-Evans, J., McLeod, D., Boulton, M. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+simple+organ+culture+model+for+assessing+the+effects+of+growth+factors+on+corneal+re-epithelialization.">A simple organ culture model for assessing the effects of growth factors on corneal re-epithelialization.</a> Exp. Eye Res. 62, 555-564 (1996).</li><li>Xu, K. P., Li, X. F., Yu, F. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Corneal+organ+culture+model+for+assessing+epithelial+responses+to+surfactants.">Corneal organ culture model for assessing epithelial responses to surfactants.</a> Toxicol. Sci. 58, 306-314 (2000).</li><li>Klausner, E. A., Peer, D., Chapman, R. L., Multack, R. F., Andurkar, S. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Corneal+gene+therapy.">Corneal gene therapy.</a> J. Control Release. 124, 107-133 (2007).</li><li>Halford, W. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=ICP0+antagonizes+Stat+1-dependent+repression+of+herpes+simplex+virus:+implications+for+the+regulation+of+viral+latency.">ICP0 antagonizes Stat 1-dependent repression of herpes simplex virus: implications for the regulation of viral latency.</a> Virol. J. 3, 44 (2006).</li><li>Bhattacharjee, P. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effective+treatment+of+ocular+HSK+with+a+human+apolipoprotein+E+mimetic+peptide+in+a+mouse+eye+model.">Effective treatment of ocular HSK with a human apolipoprotein E mimetic peptide in a mouse eye model.</a> Invest Ophthalmol Vis. Sci. 49, 4263-4268 (2008).</li><li>Novitskaya, E. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Difficulties+imaging+herpes+simplex+keratitis+with+fluorescein+isothiocynate-labeled+anti-HSV-1+antibodies+in+an+ex+vivo+model.">Difficulties imaging herpes simplex keratitis with fluorescein isothiocynate-labeled anti-HSV-1 antibodies in an ex vivo model.</a> Cornea. 28, 421-425 (2009).</li><li>Sharma, A., Shimeld, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vivo+immunofluorescence+to+diagnose+herpes+simplex+virus+keratitis+in+mice.">In vivo immunofluorescence to diagnose herpes simplex virus keratitis in mice.</a> Br. J. Ophthalmol. 81, 785-788 (1997).</li></ol>

I denne video artikel beskriver vi en fremgangsmåde til undersøgelse af akut hornhindeepitel HSV-1 infektion i en ex vivo model. Denne model er en nyttig skridt på vejen mellem in vitro og in vivo modeller, fordi det giver mulighed for fysiologisk nøjagtig validering af cellekultur resultater og derved begrænser mængden af dyreforsøg, der skal udføres. Desuden giver den ex vivo model en unik og uvurderlig mulighed for at studere epitel herpes infektion hos intakte humane</...

<table border="1"><tbody><tr><td> Reagenser / Supplies </td><td> Firma </td><td> Katalog # </td></tr><tr><td> Minimum Essential Medium (MEM) </td><td> Mediatech </td><td> 10-010-CV </td></tr><tr><td> Ikke-essentielle aminosyrer (100x opløsning) </td><td> Mediatech </td><td> 25-025-CI </td></tr><tr><td> L-Glutamin </td><td> Mediatech </td><td> 25 til 005-CI </td></tr><tr><td> Penicillin G natriumsalt </td><td> Sigma </td><td> P3032 </td></tr><tr><td> Streptomycinsulfat Salt </td><td> Sigma </td><td> S9137 </td></tr><tr><td> Agarose </td><td> Invitrogen </td><td> 15510-027 </td></tr><tr><td> Phosphatbufret saltvand (PBS) </td><td> Udarbejdet i lab </td><td> N / A </td></tr><tr><td> Trypsin EDTA (1x) &lt;/ Td&gt; <td> Mediatech </td><td> 25 til 053-CI </td></tr><tr><td> DNeasy Blood &amp; Tissue Kit </td><td> QIAGEN </td><td> 69.504 </td></tr><tr><td> RNeasy Mini Kit </td><td> QIAGEN </td><td> 74.104 </td></tr><tr><td> Laemmli puffer </td><td> Udarbejdet i lab </td><td> N / A </td></tr><tr><td> Optimal Cutting Temperatur (OCT) Compound </td><td> Tissue-Tek </td><td> 4583 </td></tr><tr><td> Hele kaninøjne (Young) </td><td> Pel-Freez </td><td> 41.211-2 </td></tr><tr><td> Menneskelige Hornhinder og hele Eyes </td><td> Lokale Eye Bank </td><td> N / A </td></tr><tr><td> 12 Well Porcelæn Spot Plate </td><td> Avogadros Lab Supply </td><td> 1877 </td></tr><tr><td> 35 mm vævskulturskål </td><td> Falcon </td><td> 353.001 </td></tr><tr><td> 6 WellCell Culture Plate </td><td> Greiner Bio-One </td><td> 657.160 </td></tr><tr><td> Kryoformen Intermediate </td><td> Tissue-Tek </td><td> 4566 </td></tr></tbody></table>

ex vivo organotypic corneal model of acute epithelial herpes simplex virus type i infection

Herpes keratitis er en af ​​de mest alvorlige patologier forbundet med herpes simplex virus-type 1 (HSV-1). Herpes keratitis er i øjeblikket den hyppigste årsag til både hornhinde-afledt og infektion-associeret blindhed i den udviklede verden. Typisk præsentation af herpes keratitis omfatter infektion af det corneale epithel og undertiden dybere hornhindens stroma og endothel, hvilket fører til sådanne permanente hornhinden patologier som ardannelse, udtynding, og opacitet 1. Corneal HSV-1-infektion er traditionelt undersøgt i to typer af eksperimentelle modeller. Den in vitro-model, hvor dyrkede monolag af hornhindeepithelceller inficeres i en petriskål, tilbyder enkelthed, højt formeringsevne hurtige forsøg og relativt lave omkostninger. På den anden side byder in vivo-model, hvor dyr, såsom kaniner eller mus inokuleres direkte i hornhinden, en meget avanceret fysiologisksystem, men har højere omkostninger, længere eksperimenter, nødvendig pasning af dyr samt en højere grad af variabilitet. I denne video artiklen, giver vi et detaljeret demonstration af en ny ex vivo model for hornhinde-epitel HSV-1 infektion, der forener styrken i både in vitro og in vivo modeller. Den ex vivo model benytter intakte hornhinder organotypically opretholdt i kultur og inficeret med HSV-1. Anvendelsen af ex vivo model giver mulighed for stærkt fysiologisk baserede konklusioner, men det er temmelig billig og kræver tid engagement sammenlignes med den in vitro model.

Watch this Scientific Journal Video about Ex Vivo Organotypic Corneal Model of Acute Epithelial Herpes Simplex Virus Type I Infection at JoVE.com

Ex Vivo Organotypic Corneal Model of Acute Epithelial Herpes Simplex Virus Type I Infection

I denne video artikel beskriver vi anvendelsen af ​​en ny<em&gt; Ex vivo</em&gt; Model af akut herpes simplex virus type I hornhinde-epitel infektion.

Ex vivo organotypisk Corneal model af akut Epiteliale Herpes Simplex Virus Type I Infektion

Herpes keratitis is one of the most severe pathologies associated with the herpes simplex virus-type 1 (HSV-1).  Herpes keratitis is currently the leading ...

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Neuroscience

Ex Vivo Organotypic Corneal Model of Acute Epithelial Herpes Simplex Virus Type I Infection

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