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 Arbeidet ble støttet av forskningsmidler ID-BOF 2006 ved Universitetet i Antwerpen (gis til PPO og AVdL), ved forskningsstipend G.0136.11 og G.0130.11 (gis til AVdL, ZB og PPO) og 1.5.021.09. N.00 (gitt til PPO) av fondet for Scientific Research-Flandern (FWO-Vlaanderen, Belgia), ved SBO forskningsstipend IWT-60838: BRAINSTIM av den flamske Institute for Science and Technology (gis til ZB og AVDL), i del av en Methusalem forskningsstipend fra den flamske regjeringen (bevilget til ZB), dels ved EC-FP6-NoE DiMI (LSHB-CT-2005-512146), EC-FP6-NoE EMIL (LSHC-CT-2004-503569) , og ved Inter Universitetet attraksjon polakker IUAP-NIMI-P6/38 (gitt til AVDL). Nathalie De Vocht har en PhD-studentship fra FWO-Vlaanderen. Peter Ponsaerts er en postdoktor i FWO-Vlaanderen.</p

1. Celleforberedelsen <ol><li> Forsøk bør igangsettes ved hjelp av ex vivo dyrkede stamceller populasjoner genetisk konstruert til å uttrykke luciferase og eGFP reporter proteiner. Her bruker vi luciferase / eGFP-uttrykker murine benmarg-deriverte stromale celler (BMSC-Luc/eGFP) som tidligere beskrevet av Bergwerf et al. 2, 5. </li><li> To dager før celle merking, plate BMSC-Luc/eGFP celler ved en tetthet på 8 x 10 5 celler per T75 kultur kolbe i 15 ml komplett ekspansjon medium (CE...

<ol>
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I denne rapporten beskriver vi en optimalisert protokoll for kombinasjonen av tre komplementære bildediagnostikk (BLI, MR og histologi) for detaljert karakterisering av mobilnettet implantater i CNS av immun kompetente mus. En kombinasjon av reporter gen merking av celler, basert på genetisk modifisering med reporteren gener ildflue luciferase og eGFP, og en direkte celle merking med GB MPIO, fører til en nøyaktig vurdering av stamcelleforskningen grafts in vivo. For partikkel m...

<table border="1"><tbody><tr><td> Navn på reagensen </td><td> Firma </td><td> Katalognummer </td><td> Kommentarer </td></tr><tr><td> IMDM </td><td> Lonza </td><td> BE12-722F </td><td> Komponenten av cellevekst medium CEM </td></tr><tr><td> Fetal bovin serum </td><td> Gibco </td><td> 10270-106 </td><td> Komponenten av cellevekst medium CEM </td></tr><tr><td> Horse serum </td><td> Gibco </td><td> 1605-122 </td><td> Komponenten av cellevekst medium CEM </td></tr><tr><td> Penicillin-streptomycin </td><td> Gibco </td><td> 15140 </td><td> Komponenten av cellevekst medium CEM </td></tr><tr><td> Fungizone </td><td> Gibco </td><td> 15290-018 </td><td> Komponenten av cellevekst medium 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 Health omsorg </td><td></td><td></td></tr><tr><td> Isofluran </td><td> Isoflo </td><td> 05260-05 </td><td></td></tr><tr><td> 0,9% NaCl-løsning </td><td> Baxter </td><td></td><td></td></tr><tr><td> paraformaldehyde </td><td> Merck </td><td> 1.04005.1000 </td><td></td></tr><tr><td> sukrose </td><td> Industry </td><td> A1125 </td><td></td></tr><tr><td> Micro-innsprøytningspumpe </td><td> KD vitenskapelig </td><td> KDS100 </td><td></td></tr><tr><td> Photon imager </td><td> Biospace Lab </td><td></td><td></td></tr><tr&gt; <td> 9.4T MR skanner </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 cryostat </td><td> Prosan </td><td> HM525 </td><td></td></tr><tr><td> sprøyte </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 programvare </td><td> Biospace Lab </td><td></td><td></td></tr><tr><td> M3vision programvare </td><td> Biospace Lab </td><td></td><td></td></tr><tr><td> Paravision 5.1 programvare </td><td> Bruker Biospin </td><td></td><td></td></tr><tr><td> Amira 4.0 programvare </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 løpet av det siste tiåret har stamcelletransplantasjon fått økende interesse som primær eller sekundær terapeutisk modalitet for en rekke sykdommer, både i prekliniske og kliniske studier. Men hittil resultater mht funksjonelle resultatet og / eller vev regenerering følgende stamcelletransplantasjon er ganske mangfoldig. Vanligvis er en klinisk nytte observert uten dyp forståelse av de underliggende mekanisme (er) 1. Derfor har flere innsats førte til utviklingen av ulike molekylære imaging modaliteter å overvåke stamcelleforskningen pode med det endelige målet til nøyaktig evaluere overlevelse, skjebne og fysiologi av podet stamceller og / eller deres mikro-miljø. Observerte endringene i én eller flere parametre bestemt av molekylær bildediagnostikk kan være relatert til den observerte kliniske effekten. I denne sammenheng våre studier fokuserer på den kombinerte bruken av bioluminesens imaging (BLI), magnetisk resonans imaging (MRI) og histologiske analysis å evaluere stamcelleforskningen poding. BLI brukes ofte til ikke-invasiv utføre celle sporing og overvåke celle overlevelse i tiden etter 2-7 transplantasjon, basert på en biokjemisk reaksjon der cellene uttrykker luciferase-reporter genet er i stand til å sende ut lys etter samspill med underlaget sin (for eksempel D- luciferin 8), 9. MR derimot er en non-invasiv teknikk som er klinisk relevant 10 og kan brukes til nøyaktig lokalisere cellulære grafts med svært høy oppløsning 11-15, selv om følsomheten svært avhengig av kontrasten generert etter celle merking med en MR kontrastmiddel . Endelig er post-mortem histologisk analyse metoden for valg for å validere forskningsresultater oppnådd med ikke-invasive teknikker med høyest oppløsning og følsomhet. Videre endepunkt histologisk analyse tillater oss å utføre detaljerte fenotypisk analyse av podet celler og / eller omkringliggende vev, BAsed på bruk av fluorescerende reporter proteiner og / eller direkte celle merking med spesifikke antistoffer. I sammendraget, vi her visuelt demonstrere komplementaritet i BLI, MR og histologi for å avdekke ulike stamcelleforskningen-og / eller miljø-assosiert egenskaper etter stamcelle transplantasjon i CNS av mus. Som et eksempel, benmarg-avledet stromale celler, genmanipulerte å uttrykke den forbedrede grønt fluorescerende protein (eGFP) og Firefly luciferase (svingninger), og merket med blå lysstoffrør mikron-størrelse jernoksid partikler (MPIOs), vil bli podet i CNS av immun-kompetente mus og utfallet vil bli overvåket av BLI, MR 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 artikkelen beskriver en optimalisert sekvens av hendelser for multimodal imaging av cellulære grafts i gnager hjernen ved hjelp: (i) in vivo bioluminesens og magnetisk resonans imaging, og (ii) post mortem histologisk analyse. Ved å kombinere disse bildediagnostikk på et enkelt dyr lar mobilnettet pode evaluering med høy oppløsning, sensitivitet og spesifisitet.

Multimodal Imaging of Stem Cell implantasjon i sentralnervesystemet av 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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