Name: bioprintable alginate/gelatin hydrogel 3d in vitro model systems induce cell spheroid formation
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Description: Watch this Scientific Journal Video about Bioprintable Alginate/Gelatin Hydrogel 3D In Vitro Model Systems Induce Cell Spheroid Formation at JoVE.com

Tao Jiang bedankt de China Scholarship Raad (201403170354) en het McGill doctorale techniekprijs (90025) voor de financiering van hun beurs. Jose G. Munguia-Lopez Bedankt CONACYT (250279, 290936 en 291168) en FRQNT (258421) voor de financiering van hun beurs. Salvador Flores-Torres Bedankt CONACYT voor hun beurs financiering (751540). Joseph M. Kinsella bedankt de National Science en Engineering Research Council, de Canadese Stichting voor innovatie, Townshend-Lamarre Family Foundation en McGill University voor hun financiering. We zouden graag bedanken Allen Ehrlicher voor het toestaan van ons om te gebruiken zijn rheometer, Dan Nicolau voor het toestaan van ons om te gebruiken zijn confocal microscoop en Morag Park voor de toekenning van ons toegang tot fluorescently geëtiketteerde cellijnen.

1. voorbereiding van de materialen, Hydrogel en cel cultuur materialen
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Voorbereiding op het materiaal en de oplossing
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<li>Wassen en drogen van 250 mL en 100 mL glazen bekers Magneetroerders, spatels, 10 mL cartridges, 25 G cilindrische sproeiers met (een lengte van 0.5 in) en een inwendige diameter van 250 µm. De materialen steriliseren in autoclaaf bij 121 ° C/15 min. hen/1 atm. de materialen onder steriele omstandigheden houden tot gebruik. Nota: Verwijs naar de Tabel van ...

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	<li>Cui, X., Hartanto, Y., Zhang, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Advances+in+multicellular+spheroids+formation.">Advances in multicellular spheroids formation.</a> Journal of the Royal Society Interface. 14 (127), (2017).</li><li>Yip, D., Cho, C. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+multicellular+3D+heterospheroid+model+of+liver+tumor+and+stromal+cells+in+collagen+gel+for+anti-cancer+drug+testing.">A multicellular 3D heterospheroid model of liver tumor and stromal cells in collagen gel for anti-cancer drug testing.</a> Biochemical and Biophysical Research Communications. 433 (3), 327-332 (2013).</li><li>Breslin, S., O'Driscoll, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+relevance+of+using+3D+cell+cultures,+in+addition+to+2D+monolayer+cultures,+when+evaluating+breast+cancer+drug+sensitivity+and+resistance.">The relevance of using 3D cell cultures, in addition to 2D monolayer cultures, when evaluating breast cancer drug sensitivity and resistance.</a> Oncotarget. 7 (29), 45745-45756 (2016).</li><li>Yue, X., Lukowski, J. 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V., Atala, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=3D+bioprinting+of+tissues+and+organs.">3D bioprinting of tissues and organs.</a> Nature Biotechnology. 32 (8), 773-785 (2014).</li><li>Kang, H. 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=A+3D+bioprinting+system+to+produce+human-scale+tissue+constructs+with+structural+integrity.">A 3D bioprinting system to produce human-scale tissue constructs with structural integrity.</a> Nature Biotechnology. 34 (3), 312-319 (2016).</li><li>Miller, J. 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=Rapid+casting+of+patterned+vascular+networks+for+perfusable+engineered+three-dimensional+tissues.">Rapid casting of patterned vascular networks for perfusable engineered three-dimensional tissues.</a> Nature Materials. 11 (9), 768-774 (2012).</li><li>Jia, 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=Direct+3D+bioprinting+of+perfusable+vascular+constructs+using+a+blend+bioink.">Direct 3D bioprinting of perfusable vascular constructs using a blend bioink.</a> Biomaterials. 106, 58-68 (2016).</li><li>Kolesky, D. B., Homan, K. A., Skylar-Scott, M. A., Lewis, J. 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H., Zhao, J. J., Tasoglu, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bioprinting+for+cancer+research.">Bioprinting for cancer research.</a> Trends in Biotechnology. 33 (9), 504-513 (2015).</li><li>Derby, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Printing+and+prototyping+of+tissues+and+scaffolds.">Printing and prototyping of tissues and scaffolds.</a> Science. 338 (6109), 921-926 (2012).</li><li>Nair, 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=Characterization+of+cell+viability+during+bioprinting+processes.">Characterization of cell viability during bioprinting processes.</a> Biotechnology Journal. 4 (8), 1168-1177 (2009).</li><li>Costa, E. C., 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=3D+tumor+spheroids:+an+overview+on+the+tools+and+techniques+used+for+their+analysis.">3D tumor spheroids: an overview on the tools and techniques used for their analysis.</a> Biotechnology Advances. 34 (8), 1427-1441 (2016).</li><li>Zhao, Y., 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=Three-dimensional+printing+of+Hela+cells+for+cervical+tumor+model+in+vitro.">Three-dimensional printing of Hela cells for cervical tumor model in vitro.</a> Biofabrication. 6 (3), 035001 (2014).</li><li>Ling, 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=Bioprinting-based+high-throughput+fabrication+of+three-dimensional+MCF-7+human+breast+cancer+cellular+spheroids.">Bioprinting-based high-throughput fabrication of three-dimensional MCF-7 human breast cancer cellular spheroids.</a> Engineering. 1 (2), 269-274 (2015).</li><li>Liang, Y., 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+cell-instructive+hydrogel+to+regulate+malignancy+of+3D+tumor+spheroids+with+matrix+rigidity.">A cell-instructive hydrogel to regulate malignancy of 3D tumor spheroids with matrix rigidity.</a> Biomaterials. 32 (35), 9308-9315 (2011).</li><li>Szot, C. S., Buchanan, C. F., Freeman, J. W., Rylander, M. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=3D+in+vitro+bioengineered+tumors+based+on+collagen+I+hydrogels.">3D in vitro bioengineered tumors based on collagen I hydrogels.</a> Biomaterials. 32 (31), 7905-7912 (2011).</li><li>Carey, S. P., Kraning-Rush, C. M., Williams, R. M., Reinhart-King, C. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biophysical+control+of+invasive+tumor+cell+behavior+by+extracellular+matrix+microarchitecture.">Biophysical control of invasive tumor cell behavior by extracellular matrix microarchitecture.</a> Biomaterials. 33 (16), 4157-4165 (2012).</li><li>Hospodiuk, M., Dey, M., Sosnoski, D., Ozbolat, I. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+bioink:+a+comprehensive+review+on+bioprintable+materials.">The bioink: a comprehensive review on bioprintable materials.</a> Biotechnology Advances. 35 (2), 217-239 (2017).</li><li>Caliari, S. R., Burdick, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+practical+guide+to+hydrogels+for+cell+culture.">A practical guide to hydrogels for cell culture.</a> Nature Methods. 13 (5), 405-414 (2016).</li><li>Bhutani, U., Laha, A., Mitra, K., Majumdar, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sodium+alginate+and+gelatin+hydrogels:+viscosity+effect+on+hydrophobic+drug+release.">Sodium alginate and gelatin hydrogels: viscosity effect on hydrophobic drug release.</a> Materials Letters. 164, 76-79 (2016).</li><li>Biswal, D., 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=Effect+of+mechanical+and+electrical+behavior+of+gelatin+hydrogels+on+drug+release+and+cell+proliferation.">Effect of mechanical and electrical behavior of gelatin hydrogels on drug release and cell proliferation.</a> Journal of the Mechanical Behavior of Biomedical Materials. 53, 174-186 (2016).</li><li>Rowley, J. A., Madlambayan, G., Mooney, D. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Alginate+hydrogels+as+synthetic+extracellular+matrix+materials.">Alginate hydrogels as synthetic extracellular matrix materials.</a> Biomaterials. 20 (1), 45-53 (1999).</li><li>Djabourov, M., Leblond, J., Papon, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gelation+of+aqueous+gelatin+solutions.+I.+Structural+investigation.">Gelation of aqueous gelatin solutions. I. Structural investigation.</a> Journal de Physique (France). 49 (2), 319-332 (1988).</li><li>Djabourov, M., Leblond, J., Papon, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gelation+of+aqueous+gelatin+solutions.+II.+Rheology+of+the+sol-gel+transition.">Gelation of aqueous gelatin solutions. II. Rheology of the sol-gel transition.</a> Journal de Physique (France). 49 (2), 333-343 (1988).</li><li>Coussot, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Rheometry of Pastes, Suspensions, and Granular Materials: Applications in Industry and Environment. , (2005).</li><li>Ouyang, L., Yao, R., Zhao, Y., Sun, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+bioink+properties+on+printability+and+cell+viability+for+3D+bioplotting+of+embryonic+stem+cells.">Effect of bioink properties on printability and cell viability for 3D bioplotting of embryonic stem cells.</a> Biofabrication. 8 (3), 035020 (2016).</li><li>Michon, C., Cuvelier, G., Launay, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Concentration+dependence+of+the+critical+viscoelastic+properties+of+gelatin+at+the+gel+point.">Concentration dependence of the critical viscoelastic properties of gelatin at the gel point.</a> Rheologica Acta Rheologica Acta: An International Journal of Rheology. 32 (1), 94-103 (1993).</li><li>Mouser, V. 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Cel-beladen structuren kunnen worden aangetast als er verontreiniging (biologische of chemische) optreedt op elk punt in het proces. Meestal, biologische verontreiniging wordt gezien na twee of drie dagen van de cultuur als een kleur wijzigen in de cultuur-media of de structuur van de bioprinted. De sterilisatie (fysische en chemische desinfectie) is daarom een belangrijke stap voor alle cel-gerelateerde processen. Opmerkelijk, autoclaaf gelatine verandert de gelerend eigenschappen, waardoor het langzamer gel in de proev...

Hoewel 2D celkweek wijd in kankeronderzoek gebruikt wordt, bestaan beperkingen als de cellen worden gekweekt in een opmaak monolayer met een uniforme concentratie van voedingsstoffen en zuurstof. Deze culturen ontbreken belangrijke cel-cel en cel-matrix interacties in de communicatie van de oorspronkelijke tumor (TME). Daarom recapituleren deze modellen slecht fysiologische omstandigheden, wat resulteert in de afwijkende cel gedrag, met inbegrip van onnatuurlijke morphologies, onregelmatige receptor organisatie, membraan polarisatie en abnormale genexpressie, onder andere voorwaarden1,2,3,4. Aan de andere kant, biedt 3D-celkweek, waar cellen worden uitgevouwen in een volumetrische ruimte als aggregaten, spheroïden of organoids, een alternatieve techniek om nauwkeuriger in vitro omgevingen inrichten om te bestuderen fundamentele celbiologie en fysiologie. 3D cel cultuur modellen kunnen ook het aanmoedigen van cel-ECM interacties die kritische fysiologische kenmerken van de inheemse TME in vitro1,4,5 zijn. De opkomende 3D bioprinting technologie biedt mogelijkheden om te bouwen van modellen die de heterogene TME na te bootsen.
3D bioprinting is afgeleid van snelle prototyping en maakt de fabricage van 3D microstructuren die kunnen nabootsen van sommige van de complexiteit van levende weefsel monsters6,7. De huidige bioprinting methoden omvatten inkjet-, extrusie- en laser-assisted afdrukken8. Onder hen kunt de methode van de extrusie de heterogeniteit binnen de afgedrukte matrices worden gecontroleerd door het juist positioneren van verschillende soorten materialen op verschillende locaties van de eerste. Daarom is het de beste aanpak te fabriceren van heterogene in vitro modellen waarbij meerdere soorten cellen of matrices. Extrusie bioprinting is met succes gebruikt om te bouwen van auricular gevormde steigers9, vasculaire structuren10,11,12, en de huid weefsels13, wat resulteert in hoge afdrukken trouw en cel levensvatbaarheid. De technologie beschikt ook over veelzijdige materiële selecties, de mogelijkheid om te storten materialen met cellen ingebed met een bekende dichtheid, en hoge reproduceerbaarheid14,15,16,17 . Natuurlijke en synthetische hydrogels worden vaak gebruikt als bioinks voor 3D bioprinting als gevolg van hun biocompatibiliteit, topicale en hun hydrofiele netwerken die kunnen worden ontworpen om te lijken structureel op de ECM7,18 ,19,20,21,22,23. Hydrogels zijn ook voordelig aangezien zij zelfklevende sites voor cellen, structurele elementen, permeabiliteit voor voedingsstoffen en gassen opnemen kunnen en de juiste mechanische eigenschappen te moedigen cel ontwikkeling24. Bijvoorbeeld, bieden collageen hydrogels integrine anchorage sites die cellen gebruiken kunnen om te koppelen aan de matrix. Gelatine, gedenatureerde collageen, behoudt vergelijkbare cel adhesie sites. Daarentegen alginaat is bioinert maar geeft een mechanische integriteit door de vorming van crosslinks met divalente ionen25,26,27,28.
In dit werk ontwikkelden we een samengestelde hydrogel als een bioink, samengesteld uit alginaat en gelatine, met overeenkomsten met de microscopische architectuur van een inheemse tumor stroma. Borstkankercellen en fibroblasten werden ingesloten in de hydrogels en afgedrukt via een extrusie gebaseerde bioprinter om te maken van een 3D-model dat de communicatie in vivo nabootst. De gemanipuleerde 3D-omgeving kunnen kankercellen aan vormen van meercellige tumor spheroïden (MCTS) met een hoge levensvatbaarheid voor lange periodes van cultuur van de cel (> 30 dagen). Dit protocol laat zien van de methoden van samengestelde hydrogels synthese, karakterisering van de materialen microstructuur en bedrukbaarheid, bioprinting cellulaire heterogene modellen, en observeren van de vorming van MCTS. Deze methoden kunnen worden toegepast op andere bioinks in de bioprinting van de extrusie ook over verschillende ontwerpen van heterogene weefsel modellen met mogelijke toepassingen in drug screening, cel migratie tests en studies die gericht zijn op fundamentele cel fysiologische functies.

<table border="0" cellpadding="0" cellspacing="0" width="925">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">Sodium alginate</td>
			<td style="width:213px;">FMC BioPolymer</td>
			<td style="width:173px;">CAS-No: 9005-38-3</td>
			<td style="width:167px;">Protanal LF 10/60 FT</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">Gelatin</td>
			<td style="width:213px;">Sigma-Aldrich</td>
			<td style="width:173px;">G9391</td>
			<td>Type B gelatin from bovine skin</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">Dubelcco&#39;s phosphate buffered saline (DPBS 1X)</td>
			<td style="width:213px;">Gibco</td>
			<td style="width:173px;">LS14190136</td>
			<td>1&#215;, w/o calcium, w/o magnesium</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">Magnetic hotplate</td>
			<td style="width:213px;">Corning&#160;</td>
			<td style="width:173px;">N/A</td>
			<td>Stirrer/hot plate model PC-420</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">50 mL centrifuge tubes</td>
			<td style="width:213px;">Corning</td>
			<td style="width:173px;">352098</td>
			<td>Falcon&#174; 50mL High Clarity PP Centrifuge Tube, Conical Bottom, Sterile</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">Centrifuge</td>
			<td style="width:213px;">GMI</td>
			<td style="width:173px;">N/A</td>
			<td>Sorvall RT6000D, GMI, USA</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Calcium chloride anhydrous</td>
			<td style="width:213px;">Sigma-Aldrich</td>
			<td>C1016</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">MilliQ water</td>
			<td style="width:213px;">Millipore</td>
			<td style="width:173px;">N/A</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">Millipore 0.22 &#181;m filters</td>
			<td style="width:213px;">Millipore</td>
			<td style="width:173px;">SLGS033SB</td>
			<td>Millex-GS Syringe Filter Unit, 0.22&#160;&#181;m, mixed cellulose esters, 33&#160;mm, ethylene oxide sterilized</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">Oscillation rheometer MCR 302</td>
			<td style="width:213px;">Anton Paar</td>
			<td style="width:173px;">N/A</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">Rheometer measuring tool CP25</td>
			<td style="width:213px;">Anton Paar</td>
			<td style="width:173px;">79038</td>
			<td>Conical plate geometry for rheometer</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">RheoCompass</td>
			<td style="width:213px;">Anton Paar</td>
			<td style="width:173px;">N/A</td>
			<td>Software controlling rheometer MCR 302</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">Scanning electron microscope</td>
			<td style="width:213px;">Hitachi</td>
			<td style="width:173px;">N/A</td>
			<td>SEM, Hitachi SU-3500 Variable Pressure</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">Paraformaldehyde, 96%, extra pure</td>
			<td style="width:213px;">Acros Organics</td>
			<td style="width:173px;">416785000</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">Dulbecco modified eagle medium (DMEM)</td>
			<td style="width:213px;">Gibco</td>
			<td>11965092</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Antibiotic/Antimycotic solution (100X) stabilized</td>
			<td style="width:213px;">Sigma</td>
			<td>A5955</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Fetal bovine serum</td>
			<td>Wisent Bioproducts</td>
			<td style="width:173px;">080-150</td>
			<td></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:372px;">Cell culture T-75 flasks</td>
			<td style="width:213px;">Sigma-Aldrich</td>
			<td style="width:173px;">CLS430641</td>
			<td>75 cm2 TC-Treated surface treatment</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">3D bioprinter BioScaffolder 3.1</td>
			<td style="width:213px;">GeSiM</td>
			<td style="width:173px;">N/A</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">GeSim software</td>
			<td style="width:213px;">GeSiM</td>
			<td style="width:173px;">N/A</td>
			<td>Software controlling BioScaffolder 3.1</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">10cc cartridge UV resist</td>
			<td>EFD Nordson</td>
			<td>7012126</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">End cap</td>
			<td>EFD Nordson</td>
			<td>7014472</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Tip cap</td>
			<td>EFD Nordson</td>
			<td>7014469</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Piston&#160;</td>
			<td>EFD Nordson</td>
			<td>7012182</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Stainless nozzle G25</td>
			<td>EFD Nordson</td>
			<td>7018345</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">Water bath</td>
			<td style="width:213px;">VWR</td>
			<td style="width:173px;">N/A</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">Agarose</td>
			<td style="width:213px;">Sigma-Aldrich</td>
			<td style="width:173px;">A9539</td>
			<td>Bioreagent, for molecular biology</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">Costar 6-well plates&#160;</td>
			<td style="width:213px;">Corning</td>
			<td style="width:173px;">3516</td>
			<td>TC-Treated Multiple Well Plates, Individually Wrapped, Sterile&#160;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">Confocal spinning disk inverted microscope</td>
			<td style="width:213px;">Olympus Life Science</td>
			<td style="width:173px;">N/A</td>
			<td>Olympus IX83</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">MTS assay kit</td>
			<td style="width:213px;">Promega</td>
			<td style="width:173px;">G3582</td>
			<td>CellTiter 96&#174; AQueous&#160;One Solution Cell Proliferation Assay&#160;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">Live/Dead viability cytotoxicity kit</td>
			<td>Molecular Probes,ThermoFisher Scientific</td>
			<td>L3224</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Trypsin 0.25/EDTA 1X</td>
			<td>Gibco</td>
			<td>25200-072</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">Corning 96-well plate</td>
			<td style="width:213px;">Corning</td>
			<td style="width:173px;">3595</td>
			<td>Clear Flat Bottom Polystyrene TC-Treated Microplate, Individually Wrapped, with Low Evaporation Lid, Sterile</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">Autoclave Tuttnauer</td>
			<td style="width:213px;">Heidolph Brinkmann</td>
			<td style="width:173px;">N/A</td>
			<td>Heidolph Tuttnauer 2540E Autoclave Sterilizer Electronic Model with 4 Stainless Steel Trays, 23L Capacity</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">Trypan blue</td>
			<td style="width:213px;">Invitrogen&#160;</td>
			<td style="width:173px;">T10282</td>
			<td>0.4% solution</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">Ethanol</td>
			<td style="width:213px;">Commercial Alcohols</td>
			<td style="width:173px;">P016EA95</td>
			<td>Greenfield Speciality Alcohols</td>
		</tr>
		<tr height="25">
			<td height="25" style="height:25px;width:372px;">CO2 Incubator</td>
			<td style="width:213px;">Panasonic</td>
			<td style="width:173px;">N/A</td>
			<td>MCO 19AIC-PA</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">Lyophilizer&#160;</td>
			<td style="width:213px;">SP Scientific</td>
			<td style="width:173px;">N/A</td>
			<td>Virtis Sentry 2.0</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">SolidWorks</td>
			<td style="width:213px;">Dassault Systems</td>
			<td style="width:173px;">N/A</td>
			<td>A CAD software used to build demostrative propeller-like model</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:372px;">MATLAB</td>
			<td style="width:213px;">The MathWorks</td>
			<td style="width:173px;">N/A</td>
			<td>A programming software used to generate G-code for BioScaffolder 3.1</td>
		</tr>
	</tbody>
</table>

bioprintable alginate/gelatin hydrogel 3d in vitro model systems induce cell spheroid formation

De cellulaire, biochemische en biofysische heterogeniteit van de oorspronkelijke tumor communicatie is niet gerecapituleerd door groeiende vereeuwigd kanker cellijnen met behulp van conventionele celcultuur tweedimensionale (2D). Deze uitdagingen kunnen worden overwonnen met behulp van bioprinting technieken om te bouwen van heterogene driedimensionale (3D) tumor modellen waarbij verschillende soorten cellen zijn ingesloten. Alginaat en gelatine zijn twee van de meest voorkomende biomaterialen werkzaam in bioprinting als gevolg van hun biocompatibiliteit, biomimicry en mechanische eigenschappen. Door het combineren van de twee polymeren, bereikt we een hydrogel bioprintable composiet met overeenkomsten met de microscopische architectuur van een inheemse tumor stroma. We onderzocht de bedrukbaarheid van de samengestelde hydrogel via reologie en verkregen van het optimale afdrukken venster. Borstkankercellen en fibroblasten werden ingesloten in de hydrogels en afgedrukt om te vormen van een 3D-model na te bootsen van de communicatie in vivo . De heterogene model van bioprinted bereikt een hoge levensvatbaarheid voor langetermijnkweek van cel (> 30 dagen) en bevordert de zelf-assemblage van borstkankercellen in meercellige tumor spheroïden (MCTS). We hebben vastgesteld dat de migratie en de interactie van de cellen van de fibroblast kanker-geassocieerde (IGFA) met de MCTS in dit model. Met behulp van bioprinted cel cultuur platformen als co cultuur systemen, biedt het een uniek instrument om te bestuderen van de afhankelijkheid van tumorvorming van de stroma-samenstelling. Deze techniek beschikt over een hoge-doorvoer, lage kosten en hoge reproduceerbaarheid kan, en het ook een alternatief model voor conventionele enkelgelaagde celculturen en dierlijke tumor modellen om te studeren Kankerbiologie.

De temperatuur sweep toont een duidelijk verschil van de voorloper van de A3G7 bij 25 ° C en 37 ° C. De voorloper is vloeibare bij 37 ° C en heeft een complexe viscositeit van 1938.1 ± 84.0 mPa x s, die is gevalideerd door een grotere G "over G'. Als de temperatuur daalt, ondergaat de voorloper fysieke gelering toe te schrijven aan de spontane fysieke verstrengeling van de gelatine moleculen in een tri-helix vorming29,30. Zowe...

Watch this Scientific Journal Video about Bioprintable Alginate/Gelatin Hydrogel 3D In Vitro Model Systems Induce Cell Spheroid Formation at JoVE.com

Bioprintable Alginate/Gelatin Hydrogel 3D In Vitro Model Systems Induce Cell Spheroid Formation

Wij ontwikkeld een heterogene borst kanker model bestaande uit vereeuwigd tumor en fibroblast cellen ingebed in een bioprintable alginaat/gelatine bioink. Het model recapituleert de tumor in vivo communicatie en vergemakkelijkt de vorming van meercellige tumor spheroïden, inzicht in de mechanismen van tumorvorming rijden oplevert.

Bioprintable alginaat/gelatine Hydrogel 3D In Vitro modelsystemen induceren sferoïde celvorming

The cellular, biochemical, and biophysical heterogeneity of the native tumor microenvironment is not recapitulated by growing immortalized cancer cell ...

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Bioengineering

Bioprintable Alginate/Gelatin Hydrogel 3D In Vitro Model Systems Induce Cell Spheroid Formation

Eye Irritation Test (EIT) for Hazard Identification of Eye Irritating Chemicals using Reconstructed Human Cornea-like Epithelial (RhCE) Tissue Model

Eye Irritation Test EIT for Hazard Identification of Eye Irritating Chemicals using Reconstructed Human Cornea-like Epithelial RhCE Tissue Model

Test op oogirritatie (EIT) voor Risico oog irriterende stoffen met gereconstrueerde humane Cornea-achtige Epithelial (RHCE) Tissue Model

To comply with the Seventh Amendment to the EU Cosmetics Directive and EU REACH legislation, validated non-animal alternative methods for reliable and ...

Modeling Ovarian Cancer Multicellular Spheroid Behavior in a Dynamic 3D Peritoneal Microdevice

Modelleren van eierstokkanker Meercellige Sferoïde Gedrag in een dynamische 3D peritoneale microdevice

Ovarian cancer is characterized by extensive peritoneal metastasis, with tumor spheres commonly found in the malignant ascites. This is associated with ...

An In Vitro Organ Culture Model of the Murine Intervertebral Disc

An In Vitro Organ Culture Model of the Murine Intervertebral Disc

Een<em&gt; In Vitro</em&gt; Organ Culture Model van het Muizen tussenwervelschijf

Intervertebral disc (IVD) degeneration is a significant contributor to low back pain. The IVD is a fibrocartilaginous joint that serves to transmit and ...

Improved 3D Hydrogel Cultures of Primary Glial Cells for In Vitro Modelling of Neuroinflammation

Improved 3D Hydrogel Cultures of Primary Glial Cells for In Vitro Modelling of Neuroinflammation

Verbeterd 3D Hydrogel culturen van primaire gliacellen voor In Vitro modellering van Neuroinflammation

In the central nervous system, numerous acute injuries and neurodegenerative disorders, as well as implanted devices or biomaterials engineered to enhance ...

3D Printed Porous Cellulose Nanocomposite Hydrogel Scaffolds

3D afgedrukt poreuze Cellulose nanocomposietmaterialen Hydrogel steigers

This work demonstrates the use of three-dimensional (3D) printing to produce porous cubic scaffolds using cellulose nanocomposite hydrogel ink, with ...

A Human 3D Extracellular Matrix-Adipocyte Culture Model for Studying Matrix-Cell Metabolic Crosstalk

Een menselijke 3D-extracellulaire matrix-Adipocyte cultuur model voor het bestuderen van matrix-cel metabole Overspraak

The extracellular matrix (ECM) plays a central role in regulating tissue homeostasis, engaging in crosstalk with cells and regulating multiple aspects of ...

Protocols of 3D Bioprinting of Gelatin Methacryloyl Hydrogel Based Bioinks

Protocollen van 3D Bioprinting van gelatine Methacryloyl hydrogel gebaseerde Bioinks

Gelatin methacryloyl (GelMA) has become a popular biomaterial in the field of bioprinting. The derivation of this material is gelatin, which is hydrolyzed ...

Using Multilayered Hydrogel Bioink in Three-Dimensional Bioprinting for Homogeneous Cell Distribution

Met behulp van meerlaagse Hydrogel Bioink in driedimensionale bioprinting voor homogene celdistributie

During the extrusion-based three-dimensional bioprinting process, liquid-like bioinks with low viscosity can protect cells from membrane damage induced by ...

Advanced 3D Liver Models for In vitro Genotoxicity Testing Following Long-Term Nanomaterial Exposure

Geavanceerde 3D-levermodellen voor in vitro genotoxiciteitstests na langdurige blootstelling aan nanomaterialen

Due to the rapid development and implementation of a diverse array of engineered nanomaterials (ENM), exposure to ENM is inevitable and the development of ...

3D Printing of In Vitro Hydrogel Microcarriers by Alternating Viscous-Inertial Force Jetting

3D Printing of In Vitro Hydrogel Microcarriers by Alternating Viscous-Inertial Force Jetting

3D-printen van in vitro hydrogel microdragers door afwisselend viskeuze-traagheidskracht jetting

Microcarriers are beads with a diameter of 60-250 &#181;m and a large specific surface area, which are commonly used as carriers for large-scale cell ...