Name: granulocyte-dependent autoantibody-induced skin blistering
Uploaded: 2012-10-12T13:00:00
Description: Watch this Scientific Journal Video about Granulocyte-dependent Autoantibody-induced Skin Blistering at JoVE.com

Forfatterne anerkender support af tilskud fra Deutsche Forschungsgemeinschaft SI-1281/4-1 og BIOSS fra det medicinske fakultet på Freiburg Universitet (til CS).

1. Fremstillinq af patogene antistof Bemærk: Rensning og koncentration af antistofferne bør udføres samme dag, som antistoffer ikke skal lagres i 0,1 M glycin pH 2,5-3 (elueringspuffer) natten over (ON). Affinitetsoprensning af IgG: anvende 25 ml immunt kaninserum: <ol><li> Optø kaninserum ON ved 4 ° C, blandes 1:1 med 1X PBS (løbebuffer) og centrifugeres den ved 1260 x g i 10 minutter ved 20 ° C. Eventuelt kan et filtreringstrin med filtrerpapir medtage...

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Den passive overførsel af autoantistoffer i forsøgsdyr er en vigtig fremgangsmåde til at demonstrere deres patogenicitet 7, -12. Dyremodeller, der opnås ved denne fremgangsmåde, foruden at være den indirekte bevis for autoimmunitet 13, tillader undersøgelsen af den efferente fase af den patogene mekanisme. Den passive transfer model af antistof-induceret granulocyt-afhængig hud blæredannelse epidermolysis bullosa acquisita (EBA) blev anvendt til at dissekere de mekanismer vævsskade ved au...

<table border="1"><tbody><tr><td> Navn på reagenset </td><td> Firma </td><td> Katalognummer </td><td> Kommentarer </td></tr><tr><td> rekombinant antigen </td><td></td><td></td><td> Plasmiderne kodende for de rekombinante former af murine collagen VII er tilgængelige fra den tilsvarende forfatter. </td></tr><tr><td> immunt kaninserum </td><td> <a href="http://www.eurogentec.com/" target="_blank">www.eurogentec.com</a> </td><td></td><td> Vi havde New Zealand hvide kaniner immuniseret med 200 ug af antigen, 3 gange med 2 ugers intervaller. Til dette formål har vi benyttet sig af Eurogentec SA, Belgien. </td></tr><tr><td> Protein G agarose </td><td> Roche Applied Science </td><td> 11243233001 </td><td></td></tr><tr><td> Balb / c mus </td><td> Charles River Laboratories </td><td></td><td></td&gt; </tr><tr><td> OCT-forbindelse, Tissue-Tek </td><td> Sakura Finetek </td><td> 4583 </td><td> OLT, optimal skæring temperatur </td></tr><tr><td> Kryoformen standard </td><td> Sakura Finetek </td><td> 4557 </td><td> 25 mm × 20 mm × 5 mm </td></tr><tr><td> Kryoformen mellemliggende </td><td> Sakura Finetek </td><td> 4566 </td><td> 15 mm × 15 mm × 5 mm </td></tr><tr><td> Uni-Link-Einbettkassette </td><td> R. Langenbrinck </td><td> 09-0503 </td><td> Histologi behandling og indlejring kassetter </td></tr><tr><td> Spezial-Tatowierfarbe Schwarz </td><td> H. Hauptnerund &amp; Richard Herberholz GmbH &amp; Co KG </td><td> 71492000 </td><td> tatovering pasta </td></tr><tr><td> Tatowierzange TZ1 </td><td> Ebeco E. Becker &amp; Co GmbH </td><td></td><td> tatovering anordning </td></tr><tr><td> Heparin </td><td> Carl Roth GmbH &amp; Co </td><td> 7692,1 </td><td></td></tr><tr><td> Formaldehyd 37% </td><td> Carl Roth GmbH &amp; Co </td><td> 7386 </td><td></td></tr><tr><td> Ketaminhydrochlorid </td><td> Sigma-Aldrich Chemie GmbH </td><td> K2753-1G </td><td></td></tr><tr><td> Xylazinhydrochlorid </td><td> Sigma-Aldrich Chemie GmbH </td><td> X1251-1G </td><td></td></tr><tr><td> sterilt PBS </td><td> Biochrom </td><td> L 182-50 </td><td></td></tr><tr><td> digitalkamera </td><td> Nikon </td><td> Coolpix 5400 </td><td></td></tr><tr><td> Sprøjte drevet filterenhed 0,45 pm </td><td> Millipore </td><td></td><td></td></tr><tr><td> Caliper </td><td> Mitutoyo 7309 </td><td> 1667338 </td><td> Farnell (forhandleren) </td></tr><tr><td> sygdom scoring ark </td><td></td><td></td><td> eksempel prøvelokaletsed </td></tr></tbody></table>

granulocyte-dependent autoantibody-induced skin blistering

Autoimmune fænomener forekommer hos raske personer, men når selvtolerance mislykkes, kan det autoimmune respons resultere i specifikke patologi. Ifølge Witebsky s postulater, et af kriterierne i diagnosticering af en sygdom som autoimmune er reproduktion af sygdommen hos forsøgsdyr ved passiv overførsel af autoantistoffer. For epidermolysis bullosa acquisita (EBA), en prototypisk organspecifik autoimmun sygdom i hud og slimhinder, blev adskillige eksperimentelle modeller for nylig fastslået. I dyremodellen beskrevet i vores foreliggende arbejde, oprenset IgG-antistoffer mod en strækning på 200 aminosyrer (aa 757-967) af kollagen VII injiceres gentagne gange i mus gengiver blærer fænotype såvel som histo-og immunpatologiske karakteristiske for menneskers EBA 1. Egentligt udbredt sygdom ses sædvanligvis 5-6 dage efter den første injektion og omfanget af sygdommen korrelerer med dosen af ​​den indgivne collagen VII-specifikt IgG. Den vævsbeskadigelse (blæredannelse) i den eksperimentelle EBA er afhængig af rekruttering og aktivering af granulocytter ved vævs-bundne autoantistoffer 2, -4. Derfor er denne model muliggør dissektion af granulocyt-afhængige inflammatoriske pathway involveret i den autoimmune vævsskade, som model reproducerer kun den T-celle-uafhængig fase af efferente autoimmunrespons. Endvidere er dens værdi understreges af en række undersøgelser, som viser blister-fremkaldende potentiale af autoantistoffer in vivo og undersøge mekanismen for blæredannelse i EBA 1,3, -6. Endelig vil denne model høj grad lette udviklingen af ​​nye antiinflammatoriske behandlinger i autoantistof-inducerede sygdomme. Samlet set passiv overførsel dyremodel af EBA er en tilgængelig og lærerig sygdom model og vil hjælpe forskerne til at analysere ikke blot EBA patogenese, men at besvare fundamentale biologisk og kliniskvæsentlige autoimmunitet spørgsmål.

Den passive overførsel af antigen-specifikke antistoffer resulterer i en fuldstændig blæst sygdom i mus, der ligner på kliniske, histologiske og immunpatologiske niveauer den humane EBA. Vabler, skorper, erosioner og alopeci udvikle sig på ørerne, snude, poter, ben, ryg og omkring øjnene af mus. De første kliniske tegn på sygdommen vil sandsynligvis blive vist på ørerne og / eller hoved-området. Deposition af specifik kanin IgG, og mus komplement C3 påvises ved direkte IF på den dermale epidermale krydset ...

Watch this Scientific Journal Video about Granulocyte-dependent Autoantibody-induced Skin Blistering at JoVE.com

Granulocyte-dependent Autoantibody-induced Skin Blistering

I dyremodellen beskrevet i vores foreliggende arbejde, oprenset IgG-antistoffer mod en strækning på 200 aminosyrer (aa 757-967) af kollagen VII injiceres gentagne gange i mus gengiver blærer fænotype såvel som histo-og immunpatologiske karakteristiske for menneskers epidermolysis bullosa acquisita (EBA)<sup&gt; 1</sup&gt;.

Granulocyt-afhængig autoantistof-induceret vabler

Autoimmune phenomena occur in healthy individuals, but when self-tolerance fails, the autoimmune response may result in specific pathology. According to ...

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