Name: establishment of a surgically-induced model in mice to investigate the protective role of progranulin in osteoarthritis
Uploaded: 2014-02-25T13:45:00
Description: Watch this Scientific Journal Video about Establishment of a Surgically-induced Model in Mice to Investigate the Protective Role of Progranulin in Osteoarthritis at JoVE.com

Dette arbejde blev delvist støttet af NIH forskningstilskud R01AR062207, R01AR061484, R56AI100901, K01AR053210, og en sygdom Targeted Research Grant fra Rheumatology Research Foundation (til CJ Liu).

Alle de kirurgiske procedurer for dyrene skal godkendes af den lokale Institutionens Animal Care og etiske komité, med en indsats for at minimere smerte og ubehag forårsaget af operationen. 1.. Forberedelse <ol><li> Vælg 8-12 uger gamle mandlige C57BL / 6 mus med en kropsvægt på omkring 25 g for kirurgi. </li><li> Bedøve dyrene ved intraperitoneal injektion af en cocktail, der indeholder både xylazin (5 mg / kg) og ketamin (40 mg / kg). </li><li> Barbere knæet med barbermaskiner, s...

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Det forlyder, at den stamme af mus er meget vigtigt for DMM model induktion, har så forskellige stammer af mus varierende sværhedsgrad af OA efter DMM, med højeste niveau i 129/SvEv stamme, efterfulgt af C57BL6, 129/SvInJ og derefter FVB / n 26. En stor del af transgener er etableret i C57BL6-mus, såsom PGRN-/ - mus vi anvendt i den foreliggende undersøgelse, som er forholdsvis modtagelig for DMM. Hvis transgenet er baseret på ufølsom stamme såsom FVB / n mus, er det nødvendigt at backcross disse mus...

Slidgigt (OA), også kendt som slidgigt, berører 15% af verdens befolkning og mere end 46 mio mennesker i USA, og er kendetegnet ved synovitis, brusk degeneration og osteophyte dannelse 1.. Det kan være et resultat af et komplekst samspil af genetiske, metaboliske, biomekaniske og biokemiske faktorer. De underliggende mekanismer i OA fortsat at unddrage sig det videnskabelige samfund. Der er i øjeblikket mange dyremodeller, som kan efterligne patogenesen af OA 2,3. Det er vigtigt at skabe dyremodeller i mus på grund af både tilgængeligheden af ​​forskellige genetisk modificerede mus og omkostningseffektivitet eksperimenter. Blandt de forskellige typer af eksperimentelle OA modeller er kirurgisk induceret destabilisering af mediale menisk (DMM) model er et accepteret OA model på grund af dens gode reproducerbarhed og en relativt langsommere progression under OA udvikling. Begge disse egenskaber harværet nøglen til evaluering af OA progression i forskellige behandlinger eller transgener 3-8. Imidlertid er konsistensen af ​​kirurgisk OA model påvirkes af forskellige faktorer under operationen og som et resultat, er anvendelsen af ​​kirurgiske musemodel begrænset. Progranulin (PGRN) er en multi-funktionel vækstfaktor udtrykt i forskellige celler. Det vides, at PGRN spiller en kritisk rolle i forskellige fysiologiske processer og sygdomsprocesser, såsom sårheling 9 tumorigenese 10 og inflammation 11-15. Undersøgelser også vist, at utilstrækkelig PGRN kan forårsage degenerative sygdomme i nervesystemet hos både mennesker og mus 16-18. Det vides, at PGRN udtrykkes i human ledbrusk, og niveauet er signifikant forhøjet i brusk hos patienter med OA og rheumatoid arthritis 19. Derudover PGRN spiller også en afgørende rolle i chondrocyt proliferation 20, differentiering og endochondralforbening af vækst plade under udviklingen 21,22. Vi har for nylig rapporteret, at PGRN antagoniserede TNF-α ved binding til TNF-receptorer og udviser en anti-inflammatorisk funktion i inflammatoriske arthritis modeller 13,14,23,24. , Rolle PGRN i OA, især in vivo, er imidlertid stadig at være en gåde. Heri præsenterer vi proceduren for at fremkalde en kirurgisk DMM model, og undersøge den rolle PGRN i OA udvikling gennem oprettelse af DMM model i WT og PGRN-/ - mus.

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			<td height="21" style="height:21px;width:216px;">No. 10 Surgical blades</td>
			<td style="width:135px;">FEATHER</td>
			<td style="width:119px;">25-2976#10</td>
			<td style="width:167px;"></td>
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			<td height="21" style="height:21px;">6-0 suture</td>
			<td>APPLIED DENTAL</td>
			<td>WG-N53133</td>
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establishment of a surgically-induced model in mice to investigate the protective role of progranulin in osteoarthritis

Destabilisering af mediale menisk (DMM) model er et vigtigt redskab til at studere de patofysiologiske roller talrige arthritis associerede molekyler i patogenesen af slidgigt (OA) in vivo. Men den detaljerede, især visualiseret protokol for at etablere denne kompliceret model i mus, er ikke tilgængelig. Heri tog vi fordel af vildtype og progranulin (PGRN) - / - mus som eksempler for at indføre en protokol for at fremkalde DMM model i mus, og sammenlignede den begyndende OA efter oprettelsen af ​​denne kirurgisk induceret model. De operationer, der udføres på mus var enten fingeret operation, som netop åbnet ledkapsel eller DMM drift, hvilket skære menisco-tibiale ligament og forårsagede destabilisering af mediale menisk. Slidgigt sværhedsgrad blev vurderet ved hjælp af histologiske analyse (f.eks Safranin O farvning), udtryk for OA-associerede gener, nedbrydning af brusk ekstracellulære matrix molekyler og osteophyte formularation. DMM drift held induceret OA initiering og progression i både vildtype-og PGRN-/ - mus og tab af PGNR vækstfaktor ført til en mere alvorlig OA fænotype i dette kirurgisk induceret model.

DMM model blev succes etableret i mus, og mangel på PGRN overdrevet kirurgisk induceret OA udvikling. Sham og DMM operationer (figur 1) blev udført i WT og PGRN-/ - mus. 8 uger efter operationen blev musene aflivet, og Safranin O-farvning blev udført på snit fra knæleddene, efterfulgt af statistisk analyse af arthritis score baseret på histologi. Som vist i figur 2A, der var ingen indlysende degeneration af brusk i begge genotyper i sk...

Watch this Scientific Journal Video about Establishment of a Surgically-induced Model in Mice to Investigate the Protective Role of Progranulin in Osteoarthritis at JoVE.com

Establishment of a Surgically-induced Model in Mice to Investigate the Protective Role of Progranulin in Osteoarthritis

Etablering af et kirurgisk induceret model i mus for at undersøge den beskyttende rolle i Progranulin i Slidgigt

Vi beskriver en protokol for en destabilisering af den mediale menisk (DMM) model i mus, et effektivt redskab for slidgigt (OA) forskning. Desuden har vi påvist, at mangel på progranulincan overdrive OA udvikling og progression ved hjælp af denne model, hvilket indikerer, at progranulin spiller en beskyttende rolle i patogenesen for OA.

Destabilization of medial meniscus (DMM) model is an important tool for studying the pathophysiological roles of numerous arthritis associated molecules ...

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