Name: matrix-assisted autologous chondrocyte transplantation for remodeling and repair of chondral defects in a rabbit model
Uploaded: 2013-05-21T12:00:00
Description: Watch this Scientific Journal Video about Matrix-assisted Autologous Chondrocyte Transplantation for Remodeling and Repair of Chondral Defects in a Rabbit Model at JoVE.com

Dette prosjektet ble finansiert av den tyske Research Association (DFG, HE 4578/3-1).

A. Brusk Biopsi (Kirurgi Room; trinn 1-5 i ikke-sterile Forberedelse Room) <ol><li> Utfør en endelig vekt kontroll av kanin (New Zealand hvit kanin, kvinnelig, 3,5-4,0 kg kroppsvekt, 6 måneder gamle) for å være i stand til å dosere medikamenter skikkelig og for å overvåke vekten påfølgende til kirurgi. </li><li> Indusere anestesi til kaninen ved en intravenøs injeksjon av 10 mg / kg propofol. </li><li> Etter intubasjon opprettholde anestesi med 1,5 mg / kg / min propofol og 0,05 mg / kg intravenøst ​​fe...

<ol>
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M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+dependence+of+autologous+chondrocyte+transplantation+on+varying+cellular+passage,+yield+and+culture+duration.">The dependence of autologous chondrocyte transplantation on varying cellular passage, yield and culture duration.</a> Biomaterials. 32, 5810-5818 (2011).</li><li>Frohlich, M., Malicev, E., Gorensek, M., Knezevic, M., Kregar Velikonja, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evaluation+of+rabbit+auricular+chondrocyte+isolation+and+growth+parameters+in+cell+culture.">Evaluation of rabbit auricular chondrocyte isolation and growth parameters in cell culture.</a> Cell Biol. Int. 31, 620-625 (2007).</li><li>Willers, C., Chen, J., Wood, D., Xu, J., Zheng, M. 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J. Radiol. 57, 24-31 (2006).</li><li>Benya, P. D., Shaffer, J. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dedifferentiated+chondrocytes+reexpress+the+differentiated+collagen+phenotype+when+cultured+in+agarose+gels.">Dedifferentiated chondrocytes reexpress the differentiated collagen phenotype when cultured in agarose gels.</a> Cell. 30, 215-224 (1982).</li><li>Rudert, M., Hirschmann, F., Wirth, C. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Growth+behavior+of+chondrocytes+on+various+biomaterials.">Growth behavior of chondrocytes on various biomaterials.</a> Orthopade. 28, 68-75 (1999).</li><li>Hsu, S. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evaluation+of+biodegradable+polyesters+modified+by+type+II+collagen+and+Arg-Gly-Asp+as+Tissue+Engineering+scaffolding+materials+for+cartilage+regeneration.">Evaluation of biodegradable polyesters modified by type II collagen and Arg-Gly-Asp as Tissue Engineering scaffolding materials for cartilage regeneration.</a> Artificial Organs. 30, 42-55 (2006).</li><li>Brun, P., Cortivo, R., Zavan, B., Vecchiato, N., Abatangelo, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vitro++reconstructed+tissues+on+hyaluronan-based+temporary+scaffolding.">In vitro reconstructed tissues on hyaluronan-based temporary scaffolding.</a> J. Mater. Sci. Mater. 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Den presenterte protokollen gir en etablert 9,12,13 og lett reproduserbar teknikk for å isolere autologe chondrocytes for påfølgende spredning og re-implantering i kunstig skapt brusk defekter i kanin knærne. Bruken av autologe chondrocytes for ombygging og reparasjon av leddbruskskader er allerede i klinisk bruk gir tilfredsstillende langsiktige resultater seks. Store problemer som for eksempel periosteal hypertrofi og forkalkning, pode delaminering eller donor omr...

<table border="1">
	<tbody>
		<tr>
			<td>Name of reagent/equipment</td>
			<td>Company</td>
			<td>Catalogue Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>DMEM</td>
			<td>Biochrom AG</td>
			<td>F 0415</td>
			<td></td>
		</tr>
		<tr>
			<td>Collagenase A</td>
			<td>Roche</td>
			<td>10 103 586 001</td>
			<td>0.21 U/mg</td>
		</tr>
		<tr>
			<td>Fetal calf serum (FCS)</td>
			<td>PAN Biotech GmbH</td>
			<td>3702-P103009</td>
			<td></td>
		</tr>
		<tr>
			<td>Propofol</td>
			<td>Fresenius Kabi</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Penicillin/Streptomycin</td>
			<td>Biochrom AG</td>
			<td>A 2213</td>
			<td>10,000 U/ml/10,000 &mu;g/ml</td>
		</tr>
		<tr>
			<td>PBS Dulbecco (1X)</td>
			<td>Biochrom AG</td>
			<td>L1815</td>
			<td></td>
		</tr>
		<tr>
			<td>Ethanol (70%)</td>
			<td>Merck KgaA</td>
			<td>410230</td>
			<td></td>
		</tr>
		<tr>
			<td>Trypsin-EDTA 0.25 %/0.02 %</td>
			<td>Biochrom AG</td>
			<td>L2163</td>
			<td>in PBS w/o Ca2+, Mg2+</td>
		</tr>
		<tr>
			<td>Fentanyl</td>
			<td>Delta Select GmBH</td>
			<td>1819340</td>
			<td></td>
		</tr>
		<tr>
			<td>NaCl solution (0.9%)</td>
			<td>Bbraun</td>
			<td>8333A193</td>
			<td></td>
		</tr>
		<tr>
			<td>Tissue culture dishes 100 mm/150 mm</td>
			<td>TPP AG</td>
			<td>93100/93150</td>
			<td>Growth area 60.1 mm2/147.8 mm2</td>
		</tr>
		<tr>
			<td>Tissue culture flasks 25/75 mm2</td>
			<td>TPP AG</td>
			<td>90025/90075</td>
			<td>25 mm2, 75 mm2</td>
		</tr>
		<tr>
			<td>Centrifuge Tubes (50 ml)</td>
			<td>TPP AG</td>
			<td>91050</td>
			<td>Gamma-sterilized</td>
		</tr>
		<tr>
			<td>Hemocytometer</td>
			<td>Brand GmbH+Co KG</td>
			<td>717810</td>
			<td>Neubauer</td>
		</tr>
		<tr>
			<td>Trypan Blue Solution 0.4%</td>
			<td>Sigma-Aldrich</td>
			<td>L8154</td>
			<td></td>
		</tr>
		<tr>
			<td>Spray dressing (OpSite)</td>
			<td>Smith&amp;Nephew</td>
			<td>66004978</td>
			<td>Permeable for water vapor</td>
		</tr>
		<tr>
			<td>Chondro-Gide&Ograve;</td>
			<td>Geistlich Pharma AG</td>
			<td>30915.5</td>
			<td></td>
		</tr>
		<tr>
			<td>Biopsy Punch</td>
			<td>pfm medical ag</td>
			<td>48351</td>
			<td></td>
		</tr>
		<tr>
			<td>Tissucol Duo S</td>
			<td>Baxter</td>
			<td>3419627</td>
			<td>0.5 ml</td>
		</tr>
	</tbody>
</table>

matrix-assisted autologous chondrocyte transplantation for remodeling and repair of chondral defects in a rabbit model

Leddbrusk defekter er ansett som et stort helseproblem fordi leddbrusk har en begrenset evne til selv-regenerering en. Ubehandlet brusklesjoner føre til pågående smerte, negativt påvirke livskvalitet og disponere for slitasjegikt. I løpet av de siste tiårene har flere kirurgiske teknikker blitt utviklet for å behandle slike lesjoner. Men inntil nå var det ikke mulig å oppnå en fullstendig reparasjon når det gjelder å dekke defekten med hyalint artikulær brusk eller for å gi tilfredsstillende langvarig utvinning 2-4. Derfor leddbrusk skader fortsatt et hovedmål for regenerativ teknikker som Tissue Engineering. I motsetning til andre kirurgiske teknikker, noe som ofte fører til dannelse av fibrøst vev eller fibrocartilaginous, tar sikte Vev Engineering på fullt ut å gjenopprette den komplekse strukturen og egenskapene av den opprinnelige leddbrusk ved hjelp av chondrogenic potensialet av transplanterte celler. Recent utviklingen åpnet opp lovende muligheter for regenerative brusk terapi. Den første cellen tilnærming for behandling av full-tykkelse brusk eller osteochondral lesjoner ble utført i 1994 av Peterson og Mats Lars Brittberg som utviklet klinisk autolog kondrocytt implantasjon (ACI) 5. I dag er den teknikk klinisk godt etablerte for behandling av store hyalin brusk-defekter i kne, opprettholder gode kliniske resultater selv 10 til 20 år etter implantasjon 6.. I de senere årene, gjennomgikk implantasjon av autologe chondrocytes en rask progresjon. Bruken av en kunstig tredimensjonal kollagen-matriks på hvilken cellene blir deretter omplantet ble mer og mer populær 7-9. MACT består av to operasjoner: Først, for å samle kondrocytter, trenger en biopsi brusk som skal utføres fra et ikke vektbærende brusk område av than kneleddet. Deretter blir kondrocytter blir ekstrahert, renset og utvidet til et tilstrekkelig antall celle in vitro. Kondrocytter blir deretter sådd ut på en tredimensjonal matrise og kan senere re-implantert. Når du forbereder en vev-konstruert implantat, proliferasjonsrate og differensiering kapasitet er avgjørende for en vellykket vev gjenfødelse 10. Bruken av en tredimensjonal matrise som en celle bærer er tenkt å støtte disse cellulære egenskaper 11. Følgende protokoll vil summere og demonstrerer en teknikk for isolering av kondrocytter fra brusk biopsier, deres proliferasjon in vitro og deres poding på en 3D-matrise (Chondro-Gides, Geistlich Biomaterials, Wollhusen, Sveits). Endelig vil implantering av celle-matriksinteraksjonsprosesser-konstrukter inn i kunstig skapt chondral defekter av en kanin kneleddet bli beskrevet. Denne teknikken kan brukes som en eksperimentell innstillingfor videre eksperimenter av brusk reparasjon.

Den beskrevne kirurgiske teknikken tillater en vellykket isolasjon og implantasjon av autologe chondrocytes inn en kunstig chondral defekt. Det eksperimentelle oppsettet som resulterte i en vellykket integrering av implantatet til det omgivende brusk. Etter 12 uker in vivo, ble det chondral defekten fylt med reparasjon vev med et homogent og intakt overflate, som reduserte skjærspenning og skade på implantatet (figur 4). Dessuten ble ingen hypertrofi eller forkalk...

Watch this Scientific Journal Video about Matrix-assisted Autologous Chondrocyte Transplantation for Remodeling and Repair of Chondral Defects in a Rabbit Model at JoVE.com

Matrix-assisted Autologous Chondrocyte Transplantation for Remodeling and Repair of Chondral Defects in a Rabbit Model

En eksperimentell teknikk for behandling av chondral defekter i kaninens kneleddet er beskrevet. Implantasjon av autologe chondrocytes seeded på en matrise er en godt akseptert metode for ombygging og reparasjon av leddbruskskader gi tilfredsstillende langsiktige resultater. Matrix-assistert autolog chondrocyttransplantasjon transplantasjon (MACT) tilbyr et standardisert og klinisk etablert implantasjon metoden.

Matrix-assistert Autolog kondrocytt Transplantasjon for ombygging og reparasjon av chondral Defekter i et Rabbit Model

Articular cartilage defects are considered a major health problem because articular cartilage has a limited capacity for self-regeneration 1. Untreated ...

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