Name: isolation, characterization and functional examination of the gingival immune cell network
Uploaded: 2016-02-16T15:00:00
Description: Watch this Scientific Journal Video about Isolation, Characterization and Functional Examination of the Gingival Immune Cell Network at JoVE.com

Authors were funded in part by the intramural program of NIDCR (N.M.M) and supported by a Wellcome Trust Stepping Stones Fellowship (097820/Z/11/B to J.E.K) and by a Manchester Collaborative Centre for Inflammation Research grant (to J.E.K). The authors thank Teresa Wild for critically reviewing the manuscript.

Alle experimentellen Verfahren in diesem Protokoll beschrieben gefolgt erforderlichen Richtlinien und wurden von der Institutional Animal Care und Use Committee, NIDCR / NIH genehmigt. 1. Bereiten Sie im Voraus <ol><li> Bereiten Sie die komplette Medien: RPMI mit 2 mM L-Glutamin, 100 Einheiten / ml Penicillin, 100 ug / ml Streptomycin und 10% FBS. </li><li> Bereiten DNase Mittel: 50 ml RPMI-Medium mit 7,5 ug DNase ergänzt (Make frisch, auf Eis halten zu allen Zeiten). </li><li> Bereiten Collagenase-DNase Medi...

<ol>
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Die aktuelle Technik erfolgreich eine große Zahl von Immunzellen führt (aus einer einzigen Maus) geeignet, nicht nur für die phänotypische Charakterisierung, sondern auch für funktionelle Studien ex vivo. Eine andere Gruppe war zuvor ein Protokoll veröffentlicht zu isolieren und murine gingival Immunzellen zu charakterisieren und stellte den Wert der mehrfarbigen mit Durchflusszytometrie in der Studie von Parodontitis in Tiermodellen 9. Nach erheblichen Fehlersuche den Schlüssel Änderung in d...

Zahnfleischgewebe umgeben den Menschen und der Maus Gebiss und werden ständig auf die komplexe Biofilm des Zahnes 1 ausgesetzt. Die Immunzellennetz der Zahnfleischschutz Überwachung ist entscheidend für Gewebeintegrität zu gewährleisten, mit den lokalen kommensale Bakterien gewährleisten Homöostase und, zur gleichen Zeit, zwei wirksame Immunität gegen pathogene Herausforderung bietet. Um das zu erreichen Homöostase wird das Immunsystem sorgfältig auf die Zahnfleischumgebung zugeschnitten eine hoch spezialisierte Immunzellen Netzwerk zu schaffen, noch so kleine Detail wird von den Zahnfleischimmunzellpopulationen bekannt und ihre Rolle bei der Aufrechterhaltung der Gewebe Immunität 2. Wenn Immunhomöostase auf dem Zahnfleisch gestört ist, entweder durch eine erhöhte Anfälligkeit Host und / oder das Vorhandensein von dysbiotischen mikrobiellen Gemeinschaften, eine entzündliche Erkrankung entsteht Parodontitis 3 4. Parodontitis ist eine weitverbreitete entzündliche Erkrankung, was zu einem Verlust des Zahnes supporting Strukturen. In seiner schweren Formen wird in ca. 10% der Bevölkerung im allgemeinen 5 ersichtlich. Die Schlüsselfaktoren in Parodontitis Anfälligkeit und Progression beteiligt Sezieren hat sich als schwierig erwiesen, 6. Allerdings Tiermodellen haben im Verständnis der Mechanismen der Parodontitis Initiierung und Progression 7 äußerst nützlich. Die Modelle können die wichtigsten Zellpopulationen und molekulare Mechanismen zu definieren, verwendet werden, die Immunhomöostase und Antrieb sind entscheidend Entwicklung einer Parodontitis zu halten. Eine solche Erkenntnis wird unser Verständnis von Zahnfleisch-spezifische Steuerung von Immunhomöostase und weiterhin unser gegenwärtiges Verständnis der Krankheitsentstehung zu transformieren.

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isolation, characterization and functional examination of the gingival immune cell network

Immune cell networks in tissues play a vital role in mediating local immunity and maintaining tissue homeostasis, yet little is known of the resident immune cell populations in the oral mucosa and gingiva. We have established a technique for the isolation and study of immune cells from murine gingival tissues, an area of constant microbial exposure and a vulnerable site to a common inflammatory disease, periodontitis. Our protocol allows for a detailed phenotypic characterization of the immune cell populations resident in the gingiva, even at steady state. Our procedure also yields sufficient cells with high viability for use in functional studies, such as the assessment of cytokine secretion ex vivo. This combination of phenotypic and functional characterization of the gingival immune cell network should aid towards investigating the mechanisms involved in oral immunity and periodontal homeostasis, but will also advance our understanding of the mechanisms involved in local immunopathology.

Zur Anwendung des Protokolls veranschaulichen, zeigen wir repräsentative Ergebnisse der Immunzellennetz in das Zahnfleisch von Mäusen mit und ohne Parodontitis untersucht (WT vs. LFA - / - 2A - C). Repräsentative FACS Plots zeigen Live-CD45 + hämatopoetische Zellen im Zahnfleisch (2A, 2C). Isolation und Verarbeitung von Immunzellen mit diesem Protokoll liefert genügend Zellen ex vivo S...

Watch this Scientific Journal Video about Isolation, Characterization and Functional Examination of the Gingival Immune Cell Network at JoVE.com

Isolation, Characterization and Functional Examination of the Gingival Immune Cell Network

Wir haben ein Verfahren zur Isolierung, phänotypische Charakterisierung und funktionelle Analyse von Immunzellen aus murinen Gingiva etabliert.

Isolierung, Charakterisierung und funktionelle Untersuchung des Zahnfleisch Immune Cell Network

Immune cell networks in tissues play a vital role in mediating local immunity and maintaining tissue homeostasis, yet little is known of the resident ...

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