Name: in vivo tracking of human adipose-derived mesenchymal stem cells in a rat knee osteoarthritis model with fluorescent lipophilic membrane dye
Uploaded: 2017-10-08T14:00:00
Description: Watch this Scientific Journal Video about In Vivo Tracking of Human Adipose-derived Mesenchymal Stem Cells in a Rat Knee Osteoarthritis Model with Fluorescent Lipophilic Membrane Dye at JoVE.com

Den nuværende undersøgelse blev støttet af Shanghai Innovation finansiering (1402H 294300) sponsoreret af videnskab og teknologi Kommissionen for Shanghai kommune (KN) til Dr. Wen Wang. Vi vil gerne takke Dr. Guangdong Zhou (nationale Tissue Engineering Center i Kina) for sin tekniske bistand og videnskabelig rådgivning for dette håndskrift. Vi vil også gerne takke Mr. Huitang Xia (Shanghai niende People's Hospital) for hans hjælp i dyrevelfærd.

procedurer, der involverer dyr emner blev godkendt af lokale institutionelle Animal Care og etiske komité, med et forsøg på at Minimer dyrs lidelser. Følgende protokol blev godkendt af institutionelle Animal Care og brug udvalg (IACUC) på Shanghai niende folk &#8217; s Hospital tilknyttet Shanghai JiaoTong University School of Medicine med protokollen nummer [2017] 063.
 1. etablering af en Surgically-Induced rotte knæet slidgigt Model <ol><li>For denne kirurgiske procedure, brug 8-12...

<ol>
	<li>Loeser, R. F., Goldring, S. R., Scanzello, C. R., Goldring, M. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Osteoarthritis:+a+disease+of+the+joint+as+an+organ.">Osteoarthritis: a disease of the joint as an organ.</a> Arthritis Rheum. 64 (6), 1697-1707 (2012).</li><li>Lane, N. E., Shidara, K., Wise, B. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Osteoarthritis+year+in+review+2016:+clinical.">Osteoarthritis year in review 2016: clinical.</a> Osteoarthritis Cartilage. 25 (2), 209-215 (2017).</li><li>Wang, W., Cao, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Treatment+of+osteoarthritis+with+mesenchymal+stem+cells.">Treatment of osteoarthritis with mesenchymal stem cells.</a> Sci China Life Sci. 57 (6), 586-595 (2014).</li><li>Burke, J., 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=Therapeutic+potential+of+mesenchymal+stem+cell+based+therapy+for+osteoarthritis.">Therapeutic potential of mesenchymal stem cell based therapy for osteoarthritis.</a> Clin Transl Med. 5 (1), 27 (2016).</li><li>Bendele, A. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Animal+models+of+osteoarthritis.">Animal models of osteoarthritis.</a> J Musculoskelet Neuronal Interact. 1 (4), 363-376 (2001).</li><li>Gerwin, N., Bendele, A. M., Glasson, S., Carlson, C. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+OARSI+histopathology+initiative+-+recommendations+for+histological+assessments+of+osteoarthritis+in+the+rat.">The OARSI histopathology initiative - recommendations for histological assessments of osteoarthritis in the rat.</a> Osteoarthritis Cartilage. 18, S24-S34 (2010).</li><li>Janusz, M. J., 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=Induction+of+osteoarthritis+in+the+rat+by+surgical+tear+of+the+meniscus:+Inhibition+of+joint+damage+by+a+matrix+metalloproteinase+inhibitor.">Induction of osteoarthritis in the rat by surgical tear of the meniscus: Inhibition of joint damage by a matrix metalloproteinase inhibitor.</a> Osteoarthritis Cartilage. 10 (10), 785-791 (2002).</li><li>Li, 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=In+vivo+human+adipose-derived+mesenchymal+stem+cell+tracking+after+intra-articular+delivery+in+a+rat+osteoarthritis+model.">In vivo human adipose-derived mesenchymal stem cell tracking after intra-articular delivery in a rat osteoarthritis model.</a> Stem Cell Res Ther. 7 (1), 160 (2016).</li><li>Zhou, B., 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=Administering+human+adipose-derived+mesenchymal+stem+cells+to+prevent+and+treat+experimental+arthritis.">Administering human adipose-derived mesenchymal stem cells to prevent and treat experimental arthritis.</a> Clin Immunol. 141 (3), 328-337 (2011).</li><li>Desando, G., 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=Intra-articular+delivery+of+adipose+derived+stromal+cells+attenuates+osteoarthritis+progression+in+an+experimental+rabbit+model.">Intra-articular delivery of adipose derived stromal cells attenuates osteoarthritis progression in an experimental rabbit model.</a> Arthritis Res Ther. 15 (1), 22 (2013).</li><li>Riester, S. 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=Safety+Studies+for+Use+of+Adipose+Tissue-Derived+Mesenchymal+Stromal/Stem+Cells+in+a+Rabbit+Model+for+Osteoarthritis+to+Support+a+Phase+I+Clinical+Trial.">Safety Studies for Use of Adipose Tissue-Derived Mesenchymal Stromal/Stem Cells in a Rabbit Model for Osteoarthritis to Support a Phase I Clinical Trial.</a> Stem Cells Transl Med. 6 (3), 910-922 (2017).</li><li>Bai, X., 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=Tracking+long-term+survival+of+intramyocardially+delivered+human+adipose+tissue-derived+stem+cells+using+bioluminescence+imaging.">Tracking long-term survival of intramyocardially delivered human adipose tissue-derived stem cells using bioluminescence imaging.</a> Mol Imaging Biol. 13 (4), 633-645 (2011).</li><li>Wolbank, S., 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=Labelling+of+human+adipose-derived+stem+cells+for+non-invasive+in+vivo+cell+tracking.">Labelling of human adipose-derived stem cells for non-invasive in vivo cell tracking.</a> Cell Tissue Bank. 8 (3), 163-177 (2007).</li><li>Heymer, A., 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=Iron+oxide+labelling+of+human+mesenchymal+stem+cells+in+collagen+hydrogels+for+articular+cartilage+repair.">Iron oxide labelling of human mesenchymal stem cells in collagen hydrogels for articular cartilage repair.</a> Biomaterials. 29 (10), 1473-1483 (2008).</li><li>Hemmrich, K., Meersch, M., von Heimburg, D., Pallua, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Applicability+of+the+dyes+CFSE,+CM-DiI+and+PKH26+for+tracking+of+human+preadipocytes+to+evaluate+adipose+tissue+engineering.">Applicability of the dyes CFSE, CM-DiI and PKH26 for tracking of human preadipocytes to evaluate adipose tissue engineering.</a> Cells Tissues Organs. 184 (3-4), 117-127 (2006).</li><li>Shim, G., 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=Pharmacokinetics+and+in+vivo+fate+of+intra-articularly+transplanted+human+bone+marrow-derived+clonal+mesenchymal+stem+cells.">Pharmacokinetics and in vivo fate of intra-articularly transplanted human bone marrow-derived clonal mesenchymal stem cells.</a> Stem Cells Dev. 24 (9), 1124-1132 (2015).</li><li>Chen, B. K., 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=A+safety+study+on+intrathecal+delivery+of+autologous+mesenchymal+stromal+cells+in+rabbits+directly+supporting+Phase+I+human+trials.">A safety study on intrathecal delivery of autologous mesenchymal stromal cells in rabbits directly supporting Phase I human trials.</a> Transfusion. 55 (5), 1013-1020 (2015).</li><li>Chan, M. M., Gray, B. D., Pak, K. Y., Fong, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Non-invasive+in+vivo+imaging+of+arthritis+in+a+collagen-induced+murine+model+with+phosphatidylserine-binding+near-infrared+(NIR)+dye.">Non-invasive in vivo imaging of arthritis in a collagen-induced murine model with phosphatidylserine-binding near-infrared (NIR) dye.</a> Arthritis Res Ther. 17, 50 (2015).</li><li>Texier, I., 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=Cyanine-loaded+lipid+nanoparticles+for+improved+in+vivo+fluorescence+imaging.">Cyanine-loaded lipid nanoparticles for improved in vivo fluorescence imaging.</a> J Biomed Opt. 14 (5), 054005 (2009).</li><li>Honig, M. G., Hume, R. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fluorescent+carbocyanine+dyes+allow+living+neurons+of+identified+origin+to+be+studied+in+long-term+cultures.">Fluorescent carbocyanine dyes allow living neurons of identified origin to be studied in long-term cultures.</a> J Cell Biol. 103 (1), 171-187 (1986).</li><li>Rahmati, M., Mobasheri, A., Mozafari, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Inflammatory+mediators+in+osteoarthritis:+A+critical+review+of+the+state-of-the-art,+current+prospects,+and+future+challenges.">Inflammatory mediators in osteoarthritis: A critical review of the state-of-the-art, current prospects, and future challenges.</a> Bone. 85, 81-90 (2016).</li><li>Detante, O., 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=Intravenous+administration+of+99mTc-HMPAO-labeled+human+mesenchymal+stem+cells+after+stroke:+in+vivo+imaging+and+biodistribution.">Intravenous administration of 99mTc-HMPAO-labeled human mesenchymal stem cells after stroke: in vivo imaging and biodistribution.</a> Cell Transplant. 18 (12), 1369-1379 (2009).</li><li>Hu, S. L., 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=In+vivo+magnetic+resonance+imaging+tracking+of+SPIO-labeled+human+umbilical+cord+mesenchymal+stem+cells.">In vivo magnetic resonance imaging tracking of SPIO-labeled human umbilical cord mesenchymal stem cells.</a> J Cell Biochem. 113 (3), 1005-1012 (2012).</li><li>Xia, Q., 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=Intra-articular+transplantation+of+atsttrin-transduced+mesenchymal+stem+cells+ameliorate+osteoarthritis+development.">Intra-articular transplantation of atsttrin-transduced mesenchymal stem cells ameliorate osteoarthritis development.</a> Stem Cells Transl Med. 4 (5), 523-531 (2015).</li><li>Jasmin, , 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=Optimized+labeling+of+bone+marrow+mesenchymal+cells+with+superparamagnetic+iron+oxide+nanoparticles+and+in+vivo+visualization+by+magnetic+resonance+imaging.">Optimized labeling of bone marrow mesenchymal cells with superparamagnetic iron oxide nanoparticles and in vivo visualization by magnetic resonance imaging.</a> J Nanobiotechnology. 9, 4 (2011).</li><li>Lehmann, T. P., 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=Coculture+of+human+nucleus+pulposus+cells+with+multipotent+mesenchymal+stromal+cells+from+human+bone+marrow+reveals+formation+of+tunnelling+nanotubes.">Coculture of human nucleus pulposus cells with multipotent mesenchymal stromal cells from human bone marrow reveals formation of tunnelling nanotubes.</a> Mol Med Rep. 9 (2), 574-582 (2014).</li><li>Wang, W., 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=Human+adipose-derived+mesenchymal+progenitor+cells+engraft+into+rabbit+articular+cartilage.">Human adipose-derived mesenchymal progenitor cells engraft into rabbit articular cartilage.</a> Int J Mol Sci. 16 (6), 12076-12091 (2015).</li></ol>

Sikkerhedsstandarder og biodistributionen undersøgelser af stamcelleterapi er tvingende nødvendigt før vi kan bringe regenerativ stamcelle behandling for KOA fra bænken til sengen. Men den patologiske miljø af sygdom spiller en vigtig rolle i persistens og biodistributionen transplanterede haMSCs10. For nylig, viste vores gruppe, at intra-artikulær injektion af haMSCs varet længere i en patologisk KOA miljø end gjorde injektioner under normale forhold8. Det er mulig...

Knæet slidgigt (KOA) er en degenerativ sygdom som følge af tab af ledbrusk og progressiv inflammation, som er blevet en større kroniske sygdom hos ældre omkring verden1. Nuværende behandlinger ved hjælp af anti-inflammatoriske lægemidler, fysiske kosttilskud og kirurgiske procedurer kan imidlertid kun yde midlertidig lindring for symptomatisk smerte2.
Menneskelige fedt-afledt mesenkymale stamceller (haMSCs) er blevet en lovende regenerativ middel mod knæ slidgigt, på grund af deres Multipotente differentiering potentiale for brusk regenerering og immunmodulerende egenskaber3, 4. Sammenlignet med farmakologiske ruter til at undersøge mekanismerne i aktion i vivo, er tracking live haMSCs i små KOA dyremodeller i øjeblikket lærerigt at etablere begrundelsen for og gennemførligheden af haMSC terapi før kliniske anvendelse. For præklinisk afprøvning, destabiliserer mediale meniscectomy (MM) den mekaniske belastning af fælles at fremkalde KOA i rotter, som giver et relativt realistisk model med konsekvent reproducerbarhed5. Udbrud af KOA induceret af MM er tidligere end forreste korsbånd transection alene eller kombineret med delvis mediale meniscectomy6. De langsigtede interaktioner mellem injiceres haMSCs med den patologiske mikromiljø af KOA vurderes derfor ofte i rotter induceret af MM7,8.
Om den terapeutiske virkning af haMSCs har været flittigt rapporteret, relevante viden på er implanteret haMSCs via intra-artikulær (IA) indsprøjtning i vivo persistens knappe9,10. Dermed er forskellige cellulære mærkning metoder blevet udviklet, herunder immunohistology11, luciferase12, grøn fluorescerende protein13 Transfektion, jernoxid mærkning for magnetisk resonans imaging (MR)14 , og talrige fluorescerende celle farvestoffer8,15,16. Sammenlignet med arbejdskrævende histologi analyser, beskæftiger i vivo ikke-invasive billeddannelse optiske enheder til at registrere real-time distribution og dynamikken i celler mærket med fluorescerende signaler10,17. For funktionelle levende celle imaging er cytocompatible fluorescerende mærkning en sofistikeret radioaktivt-fri tracking teknik til at afsløre cellulære aktiviteter efter stamcelle transplantation18. Derudover besidder multicolor fluorescerende lipofile farvestoffer fordele over hydrofile amino-reaktive farvestoffer eller fluorescerende proteiner, herunder deres forbedret celle permeabilitet og forbedret fluorescens quantum udbytter19.
Således protokollerne her udnytter en rød laser til at ophidse celler mærket med lipofile carbocyanines gjorde (DilC18(5)), som er en far-red fluorescerende Dil (dialkylcarbocyanines) analog20. De rød-forskudt excitations- og spektre af undgår autofluorescent indblanding og giver dyb væv imaging over en lang periode i levende dyr8. Denne metode til sporing celler i vivo mærket med gjorde er gyldig for overvågning transplanterede stamceller, såsom haMSCs, i dyremodeller, som er afgørende for at forstå og forbedre nuværende regenerativ stamcelleterapi.

<table>
	<tbody>
		<tr>
			<td>Matrx VMR animal anesthesia system</td>
			<td>Midmark</td>
			<td>VIP3000</td>
			<td></td>
		</tr>
		<tr>
			<td>4-0 suture</td>
			<td>Shanghai Jinhuan</td>
			<td>KC439</td>
			<td></td>
		</tr>
		<tr>
			<td>Razor</td>
			<td>Pritech</td>
			<td>LD-9987</td>
			<td></td>
		</tr>
		<tr>
			<td>Gentamicin</td>
			<td>Zhejiang Jindakang Animal Health Product Co., Ltd.</td>
			<td>None</td>
			<td></td>
		</tr>
		<tr>
			<td>0.9% Sodium chloride solution</td>
			<td>Hunan Kelun Pharmaceutical Co., Ltd.</td>
			<td>H43020455</td>
			<td></td>
		</tr>
		<tr>
			<td>Penicillin</td>
			<td>Shanghai Kangfu chemical pharmaceutical Co., Ltd.</td>
			<td>None</td>
			<td></td>
		</tr>
		<tr>
			<td>Buprenorphine</td>
			<td>Tianjin Pharmaceutical Research Institute Pharmaceutical Co., Ltd.</td>
			<td>None</td>
			<td></td>
		</tr>
		<tr>
			<td>Paraformaldehyde</td>
			<td>Sigma-Aldrich</td>
			<td>16005</td>
			<td>Dilute to final concentration of 10% in PBS</td>
		</tr>
		<tr>
			<td>EDTA</td>
			<td>Sigma-Aldrich</td>
			<td>E9884</td>
			<td>Dilute to final concentration of 20% in PBS</td>
		</tr>
		<tr>
			<td>0.1% Hematoxylin Solution, Mayer&#8217;s</td>
			<td>Sigma-Aldrich</td>
			<td>MHS16</td>
			<td></td>
		</tr>
		<tr>
			<td>0.5% Eosin Y solution, alcoholic</td>
			<td>Sigma-Aldrich</td>
			<td>HT110116</td>
			<td></td>
		</tr>
		<tr>
			<td>Safranin O</td>
			<td>Sigma-Aldrich</td>
			<td>S8884</td>
			<td></td>
		</tr>
		<tr>
			<td>Fast Green</td>
			<td>Sigma-Aldrich</td>
			<td>F7258</td>
			<td></td>
		</tr>
		<tr>
			<td>Shandon Excelsior ESTM Tissue Processor</td>
			<td>Thermo Fisher</td>
			<td>A78400006</td>
			<td></td>
		</tr>
		<tr>
			<td>Shandon Histocentre&#8482; 3 Tissue Embedding Center</td>
			<td>Thermo Fisher</td>
			<td>B64100010</td>
			<td></td>
		</tr>
		<tr>
			<td>Fully Automated Rotary Microtome</td>
			<td>Leica</td>
			<td>RM2255</td>
			<td></td>
		</tr>
		<tr>
			<td>DiD</td>
			<td>Molecular Probes, Life 
			Technologies</td>
			<td>V-22887</td>
			<td></td>
		</tr>
		<tr>
			<td>D-MEM High Glucose</td>
			<td>Sigma-Aldrich</td>
			<td>D5648</td>
			<td></td>
		</tr>
		<tr>
			<td>PBS</td>
			<td>GIBCO, Life Technologies</td>
			<td>14190-144</td>
			<td></td>
		</tr>
		<tr>
			<td>0.25% Trypsin-EDTA</td>
			<td>Invitrogen</td>
			<td>25200-114</td>
			<td></td>
		</tr>
		<tr>
			<td>10 cm Petri Dish</td>
			<td>Corning</td>
			<td>V118877</td>
			<td></td>
		</tr>
		<tr>
			<td>Centrifuge</td>
			<td>Beckman</td>
			<td>Optima MAX-TL</td>
			<td></td>
		</tr>
		<tr>
			<td>Fluorescent microscope</td>
			<td>Olympus</td>
			<td>BX53</td>
			<td></td>
		</tr>
		<tr>
			<td>0.4% Trypan Blue solution</td>
			<td>Sigma-Aldrich</td>
			<td>93595</td>
			<td></td>
		</tr>
		<tr>
			<td>Titetamme</td>
			<td>Virbac (Zoletil 50)</td>
			<td>1000000188</td>
			<td></td>
		</tr>
		<tr>
			<td>Zolazepam</td>
			<td>Virbac (Zoletil 50)</td>
			<td>1000000188</td>
			<td></td>
		</tr>
		<tr>
			<td>Sterile hyposermic syringe for single use 26G</td>
			<td>Shanghai Misawa Medical Industry</td>
			<td>None</td>
			<td></td>
		</tr>
		<tr>
			<td>IVIS Spectrum In Vivo Imaging System</td>
			<td>PerkinElmer</td>
			<td>124262</td>
			<td></td>
		</tr>
		<tr>
			<td>Living Imaging 4.0 software</td>
			<td>PerkinElmer</td>
			<td>None</td>
			<td></td>
		</tr>
	</tbody>
</table>

in vivo tracking of human adipose-derived mesenchymal stem cells in a rat knee osteoarthritis model with fluorescent lipophilic membrane dye

For at støtte den kliniske anvendelse af humane fedt-afledt mesenkymale stamceller (haMSC) terapi til knæet osteoarthritis (KOA), undersøgte vi effekten af celle persistens og biodistributionen af haMSCs i dyremodeller. Vi demonstrerede en metode til at mærke cellemembranen af haMSCs med lipofile fluorescerende farvestof. Efterfølgende, blev intra-artikulær injektion af mærket cellerne i rotter med kirurgisk induceret KOA overvåget dynamisk af en i vivo imaging system. Vi ansat den lipofile carbocyanines gjorde (DilC18 (5)), en far-red fluorescerende Dil (dialkylcarbocyanines) analog, som udnyttede en rød laser for at undgå excitation af den naturlige grønne autofluorescence fra omkringliggende væv. Desuden rød-forskudt emission spektre af tilladt dybe væv imaging med levende dyr og den mærkning procedure forårsagede ingen cytotoksiske virkninger eller funktionelle skader på haMSCs. Denne tilgang har vist sig at være en effektiv sporingsmetode for haMSCs i en rotte KOA model. Anvendelsen af denne metode kan også bruges til at bestemme optimal administration rute og dosering af MSCs fra andre kilder i prækliniske studier.

For at fremkalde KOA model, blev MM udført i den rigtige knæleddet af SD rotter (figur 3). Otte uger efter kirurgi, rotter blev ofret og de serielle sektioner af knæled blev vurderet med både H &#38; E og Safranin O/hurtig grøn farvning (figur 4). For H &#38; E farvning, overfladen af ledbrusken udstillet grovere grænser i kirurgi knæ end den normale fælles uden kirurgi. Safranin-O/hurtig grøn farvning, vi observeret ned...

Watch this Scientific Journal Video about In Vivo Tracking of Human Adipose-derived Mesenchymal Stem Cells in a Rat Knee Osteoarthritis Model with Fluorescent Lipophilic Membrane Dye at JoVE.com

In Vivo Tracking of Human Adipose-derived Mesenchymal Stem Cells in a Rat Knee Osteoarthritis Model with Fluorescent Lipophilic Membrane Dye

In Vivo Sporing af menneskers fedt-afledt mesenkymale stamceller i en rotte knæet slidgigt Model med fluorescerende lipofile membran farvestof

Denne protokol beskriver en effektiv måde at overvåge celle persistens og biodistributionen af menneskers fedt-afledt mesenkymale stamceller (haMSCs) af far-red fluorescens mærkning i en rotte knæet slidgigt (KOA) model via intra-artikulær (IA) injektion.

In order to support the clinical application of human adipose-derived mesenchymal stem cell (haMSC) therapy for knee osteoarthritis (KOA), we examined the ...

Research

JoVE Journal

Developmental Biology

In Vivo Tracking of Human Adipose-derived Mesenchymal Stem Cells in a Rat Knee Osteoarthritis Model with Fluorescent Lipophilic Membrane Dye

Isolation and Enrichment of Human Adipose-derived Stromal Cells for Enhanced Osteogenesis

Isolering og Berigelse af human adipøst afledte stromaceller til Enhanced Osteogenesis

Bone marrow-derived mesenchymal stromal cells (BM-MSCs) are considered the gold standard for stem cell-based tissue engineering applications. However, the ...

Analysis of Zebrafish Larvae Skeletal Muscle Integrity with Evans Blue Dye

Analyse af zebrafisk Larver Skeletmuskel Integritet med Evans blå farvestof

The zebrafish model is an emerging system for the study of neuromuscular disorders.  In the study of neuromuscular diseases, the integrity of the muscle ...

Isolation and Differentiation of Adipose-Derived Stem Cells from Porcine Subcutaneous Adipose Tissues

Isolering og differentiering af adipøst afledte stamceller fra Porcine subkutane fedtvæv

Obesity is an unconstrained worldwide epidemic. Unraveling molecular controls in adipose tissue development holds promise to treat obesity or diabetes. ...

Immunomagnetic Separation of Fat Depot-specific Sca1high Adipose-derived Stem Cells (ASCs)

Immunomagnetic Separation of Fat Depot-specific Sca1high Adipose-derived Stem Cells ASCs

Immunomagnetisk Adskillelse af Fat Depot-specifikke Sca1<sup&gt; høj</sup&gt; adipøst afledte stamceller (ASC&#39;er)

The isolation of adipose-derived stem cells (ASCs) is an important method in the field of adipose tissue biology, adipogenesis, and extracellular matrix ...

Isolation and Expansion of Mesenchymal Stem/Stromal Cells Derived from Human Placenta Tissue

Isolation og udvidelse af mesenchymstamceller / stromacellers afledt af humane Placenta Tissue

Mesenchymal stem/stromal cells (MSC) are promising candidates for use in cell-based therapies. In most cases, therapeutic response appears to be cell-dose ...

Chondrogenic Differentiation Induction of Adipose-derived Stem Cells by Centrifugal Gravity

Chondrogene differentiering Induktion af adipøst afledte stamceller ved centrifugal Gravity

Impaired cartilage cannot heal naturally. Currently, the most advanced therapy for defects in cartilage is the transplantation of chondrocytes ...

Rapid Isolation of BMPR-IB+ Adipose-Derived Stromal Cells for Use in a Calvarial Defect Healing Model

Hurtig Isolering af BMPR-IB + Adipose-afledte stromaceller til brug i en calvariale Defekt Healing Model

Invasive cancers, major injuries, and infection can cause bone defects that are too large to be reconstructed with preexisting bone from the patient's own ...

Directed Differentiation of Primitive and Definitive Hematopoietic Progenitors from Human Pluripotent Stem Cells

Instrueret differentiering af Primitive og endelige hæmatopoietisk ophav fra humane pluripotente stamceller

One of the major goals for regenerative medicine is the generation and maintenance of hematopoietic stem cells (HSCs) derived from human pluripotent stem ...

In Vitro Differentiation of Human Mesenchymal Stem Cells into Functional Cardiomyocyte-like Cells

In Vitro Differentiation of Human Mesenchymal Stem Cells into Functional Cardiomyocyte-like Cells

In Vitro Differentiering af humane mesenkymale stamceller i funktionel Cardiomyocyte-lignende celler

Myocardial infarction and the subsequent ischemic cascade result in the extensive loss of cardiomyocytes, leading to congestive heart failure, the leading ...

Culturing of Retinal Pigment Epithelial Cells on an Ex Vivo Model of Aged Human Bruch's Membrane

Culturing of Retinal Pigment Epithelial Cells on an Ex Vivo Model of Aged Human Bruch's Membrane

Dyrkning af retinale Pigment epitel celler på en Ex Vivo Model af alderen menneskelige Bruchs membran

Aside from vitamins and antioxidants recommended by the Age-Related Eye Disease Study, there is no effective therapy for &#34;dry,&#34;&#160;or atrophic ...