Name: multimodal imaging of stem cell implantation in the central nervous system of mice
Uploaded: 2012-06-13T13:00:00
Description: Watch this Scientific Journal Video about Multimodal Imaging of Stem Cell Implantation in the Central Nervous System of Mice at JoVE.com

<p class="jove_content"&gt; Forfatterne arbejde blev støttet af forskningsbevilling ID-BOF 2006 fra universitetet i Antwerpen (indrømmet PPO og AVdL), ved forskningsbevilling G.0136.11 og G.0130.11 (tildelt AVdL, ZB og PPO) og 1.5.021.09. N.00 (tildelt til PPO) af Fonden for Videnskabelig Forskning-Flandern (FWO-Vlaanderen, Belgien), ved SBO forskningsbevilling IWT-60.838: BRAINSTIM af det flamske Institut for Videnskab og Teknologi (indrømmet ZB og AVDL), i del af en Methusalem forskningsbevilling fra den flamske regering (tildelt ZB), dels ved EF-FP6-NoE dimi (LSHB-CT-2005-512146), EF-FP6-NoE EMIL (LSHC-CT-2004-503569) , og ved Inter University seværdighed polakker IUAP-NIMI-P6/38 (tildelt AVDL). Nathalie De Vocht har en ph.d.-studentship fra FWO-Vlaanderen. Peter Ponsaerts er en post-ph.d.-stipendiat i FWO-Vlaanderen.</p

1. Cellepræparat <ol><li> Forsøgene skal initieres ved hjælp af ex vivo-dyrket stamcellepopulationer genetisk manipuleret til at udtrykke luciferase og EGFP reporter-proteiner. Her anvender Luciferase / EGFP-udtrykkende murine knoglemarv-afledte stromaceller (BMSC-Luc/eGFP) som tidligere beskrevet af Bergwerf et al. 2, 5. </li><li> To dage før cellemærkning, plade BMSC-Luc/eGFP celler ved en densitet på 8 x 10 5 celler pr T75 dyrkningskolbe i 15 ml komplet ekspansionsmedium (CEM) s...

<ol>
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I denne rapport beskriver vi en optimeret protokol til en kombination af tre komplementære billeddiagnostiske metoder (BLI, MRI og histologi) for detaljeret karakterisering af cellulære implantater i CNS i immunkompetente mus. En kombination af reportergen mærkning af celler, baseret på genetisk modifikation med reportergener ildflueluciferase og EGFP og en direkte cellemærkning med GB MPIO, fører til en nøjagtig vurdering af stamceller transplantater in vivo. For partikel m?...

<table border="1"><tbody><tr><td> Navn på reagenset </td><td> Firma </td><td> Katalog nummer </td><td> Kommentarer </td></tr><tr><td> IMDM </td><td> Lonza </td><td> BE12-722F </td><td> Komponent af cellevækstmedium CEM </td></tr><tr><td> Føtalt bovint serum </td><td> Gibco </td><td> 10270-106 </td><td> Komponent af cellevækstmedium CEM </td></tr><tr><td> Hesteserum </td><td> Gibco </td><td> 1605-122 </td><td> Komponent af cellevækstmedium CEM </td></tr><tr><td> Penicillin-streptomycin </td><td> Gibco </td><td> 15140 </td><td> Komponent af cellevækstmedium CEM </td></tr><tr><td> Fungizone </td><td> Gibco </td><td> 15290-018 </td><td> Komponent af cellevækstmedium CEM </td></tr><tr><td> PBS </td><td> Gibco </td><td> 14190 </td><td></td></tr><tr><td> Puromycine </td><td> Invivogen </td><td> ant-PR-1 </td><td></td></tr><tr><td> trypsin </td&gt; <td> Gibco </td><td> 25300 </td><td></td></tr><tr><td> GB MPIO </td><td> Bangs Laboratories </td><td> ME04F/7833 </td><td></td></tr><tr><td> D-luciferin </td><td> Promega </td><td> E1601 </td><td></td></tr><tr><td> Ketamin (Ketalar) </td><td> Pfizer </td><td></td><td></td></tr><tr><td> Xylazin (Rompun) </td><td> Bayer Sundhedspleje </td><td></td><td></td></tr><tr><td> Isofluran </td><td> Isoflo </td><td> 05260-05 </td><td></td></tr><tr><td> 0,9% NaCI-opløsning </td><td> Baxter </td><td></td><td></td></tr><tr><td> paraformaldehyd </td><td> Merck </td><td> 1.04005.1000 </td><td></td></tr><tr><td> saccharose </td><td> Applichem </td><td> A1125 </td><td></td></tr><tr><td> Micro-indsprøjtningspumpe </td><td> KD videnskabelig </td><td> KDS100 </td><td></td></tr><tr><td> Foton Imager </td><td> Biospace Lab </td><td></td><td></td></tr><tr&gt; <td> 9.4T MR scanner </td><td> Bruker Biospin </td><td> Biospec 94/20 USR </td><td></td></tr><tr><td> BX51 mikroskop </td><td> Olympus </td><td> BX51 </td><td></td></tr><tr><td> Mycrom HM kryostat </td><td> Prosan </td><td> HM525 </td><td></td></tr><tr><td> sprøjte </td><td> Hamilton </td><td> 7635-01 </td><td></td></tr><tr><td> 30 gauge nål </td><td> Hamilton </td><td> 7762-03 </td><td></td></tr><tr><td> Foto Vision software </td><td> Biospace Lab </td><td></td><td></td></tr><tr><td> M3vision software </td><td> Biospace Lab </td><td></td><td></td></tr><tr><td> Paravision 5,1 software </td><td> Bruker Biospin </td><td></td><td></td></tr><tr><td> Amira 4,0 software </td><td> Visage Imaging </td><td></td><td></td></tr></tbody></table>

multimodal imaging of stem cell implantation in the central nervous system of mice

I det sidste årti har stamcelletransplantation vundet stigende interesse som primær eller sekundær terapeutisk modalitet for en række sygdomme, både i prækliniske og kliniske undersøgelser. Men til dato resultater med hensyn til det funktionelle resultat og / eller vævsregeneration følgende stamcelletransplantation er ret forskellige. Generelt er en klinisk fordel observeres uden dybere forståelse af de underliggende mekanisme (r) 1. Derfor har flere indsats førte til udviklingen af ​​forskellige molekylære billeddiagnostiske metoder til at overvåge stamceller podning med det endelige mål at præcist vurdere overlevelse, skæbne og fysiologi af podede stamceller og / eller deres mikro-miljø. Observerede ændringer i en eller flere parametre bestemmes ved molekylær billeddannelse kan være relateret til den observerede kliniske effekt. I denne sammenhæng fokusere vores undersøgelser på kombineret anvendelse af bioluminescens imaging (BLI), magnetisk resonans imaging (MRI) og histologiske analysis vurdere stamcelle podning. BLI er almindeligt anvendt til ikke-invasivt at udføre cellesporing og overvåge celleoverlevelse i tid efter transplantation 2-7, baseret på en biokemisk reaktion, hvor celler, der udtrykker luciferase-reportergenet, er i stand til at udsende lys efter interaktion med dets substrat (fx D- luciferin) 8, 9. MRI på den anden side er en ikke-invasiv teknik, som er klinisk relevant 10 og kan anvendes til præcist at lokalisere cellulære transplantater med meget høj opløsning 11-15, selv om dens følsomhed afhænger i høj grad kontrast genereret efter cellemærkning med et MRI-kontrastmiddel . Endelig post mortem histologisk analyse er den foretrukne fremgangsmåde til validering forskningsresultater opnået med ikke-invasive teknikker med den højeste opløsning og følsomhed. Endvidere endepunkt histologisk analyse gør det muligt at udføre detaljeret fænotypisk analyse af podede celler og / eller det omgivende væv, baSED om brugen af ​​fluorescerende reporter-proteiner og / eller direkte cellemærkning med specifikke antistoffer. Sammenfattende, her er vi visuelt påvise komplementariteten af ​​BLI, MRI og histologi at udrede forskellige stamcelle-og / eller miljø-associerede egenskaber som følge stamceller podning i centralnervesystemet hos mus. Som et eksempel marv-afledte knogle stromaceller, genetisk manipuleret til at udtrykke forøget Green Fluorescent Protein (EGFP) og ildflueluciferase (udsving), og mærkes med fluorescerende blå mikrometerstore jernoxidpartikler (MPIOs) vil blive podet på CNS af immun-kompetente mus, og resultaterne vil blive overvåget af BLI, MRI og histologi (figur 1).

Watch this Scientific Journal Video about Multimodal Imaging of Stem Cell Implantation in the Central Nervous System of Mice at JoVE.com

Multimodal Imaging of Stem Cell Implantation in the Central Nervous System of Mice

Denne artikel beskriver en optimeret sekvens af begivenheder for multimodal billeddannelse af cellulære transplantater i gnaverhjerne med: (i) in vivo bioluminescens og magnetisk resonansimagografi, og (ii) post mortem histologisk analyse. Ved at kombinere disse billeddiagnostiske metoder på et enkelt dyr giver cellulære graft evaluering med høj opløsning, følsomhed og specificitet.

Multimodal Imaging af stamcelle implantation i det centrale nervesystem mus

During the past decade, stem cell transplantation has gained increasing interest as primary or secondary therapeutic modality for a variety of diseases, ...

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Electrophysiological Recording of The Central Nervous System Activity of Third-Instar  Drosophila Melanogaster

Elektrofysiologiske optagelse af centralnervesystemet aktivitet af tredje-Instar Drosophila Melanogaster

The majority of the currently available insecticides target the nervous system and genetic mutations of invertebrate neural proteins oftentimes yield ...