Name: synthesis of biocompatible liquid crystal elastomer foams as cell scaffolds for 3d spatial cell cultures
Uploaded: 2017-04-11T15:00:00
Description: Watch this Scientific Journal Video about Synthesis of Biocompatible Liquid Crystal Elastomer Foams as Cell Scaffolds for 3D Spatial Cell Cultures at JoVE.com

Bu projenin mali destek için - Yazarlar Kent State University (ReMedIKS Kent State Rejeneratif Tıp Girişimi ortak araştırma hibe ve destek) teşekkür etmek istiyorum.

Not: 3-kollu yıldız blok kopolimeri kullanılarak 3D LCE köpük benzeri hazırlanması için, aşağıdaki aşamaları, Şekil 1 &#39;de gösterilmiştir. Nükleer manyetik rezonans (NMR) karakterizasyonu için, spektrumları bir Bruker DMX 400 MHz aleti üzerinde oda sıcaklığında dötoryumlanmış kloroform (CDCI3) kaydedilir ve içten 7.26 kalıntı tepeleri referans. Fourier zayıflatılmış toplam yansıma modu kullanılarak bir Bruker Vektör 33 FT-IR spektrometresi...

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Sıvı kristal elastomerler son zamanlarda nedeniyle uyaranlar tepki için biyouyumlu hücre yapı iskeleti olarak incelenmiştir. Bunlar hücre yapı iskeleti olarak ideal bir platform olduğu kanıtlanmıştır. Ancak, hazırlama ve yeni bir LCE iskele tasarlarken akılda tutulması gereken önemli bir faktördür gözeneklilik olduğunu. Sızabilir katı 23 ya da gazların dahil, her zaman homojen gözenekli ya da tamamen birbirine gözenekler ile sonuçlanmaz. kazınmış edilebilir bir metal...

Hücre çalışmalarında ve hücre bağlanması ve yayılması, 1, 2, 3, 4, 5 amaçlayan doku rejenerasyonu için bir uygulama için tasarlanmış biyolojik ve biyolojik olarak uyumlu sentetik malzemelerin çeşitli örnekler vardır. 6, 7 sipariş moleküler anizotropik harici uyaranlara yanıt olabilir sıvı kristal elastomerler (LCEs) olarak bilinen biyolojik olarak uyumlu malzemelerden birkaç örnek, olmuştur. LCEs optik özelliklerin ve sıvı kristal 8 moleküler sipariş, 9 elastomerlerin mekanik ve elastik özellikler kombine uyaranlara yanıt veren malzemelerdir. LCEs dış stimülasyonu yanıt olarak şekil değişiklikleri, mekanik deformasyon, elastik davranış ve optik özellikleri karşılaşabiliruli (yani., ısı, gerilim, ışık, vs.), 10, 11, 12, 13, 14, 15, 16. Daha önce yapılan çalışmalar, sıvı kristaller (LCS) hücre 4, 17 büyümesini ve yönünü duyu göstermiştir. LCEs hücre iskele ve hizalama gibi biyolojik ve tıbbi açıdan alakalı uygulamalar için uygun olabileceğini varsaymak mümkündür. Daha önce, bir &quot;Swiss-peynir türü&quot; gözenekli morfolojiyi 6, 18 sahip smektik biyolojik olarak uyumlu, biyolojik olarak parçalanabilen, döküm kalıplı ve ince LCEs filmlerin hazırlanmasını bildirmektedir. Ayrıca, hücre büyümesi 19 &lt;için iskeleler globüler morfolojisi olan nematik biyo-uyumlu LCEs hazırlanansup&gt;, 20. Çalışmalarımız ilgi 21&#39;in dokusunun içine alacak şekilde malzemelerin mekanik özelliklerinin ayar hedeflendi. Ayrıca, bu çalışmalar elastomer hücre etkileşimleri yanı sıra elastomerler dış uyaranlara maruz hücresel yanıtı anlamak odaklanır. temel zorluklar elastomer matrisi yoluyla hücre bağlanması ve nüfuz imkan vermek için LCEs gözenekliliğini uygun hale kısmen ve daha iyi kütle transportuna yönelik idi. Bu ince filmler 6 gözenekliliği matrisin kütlesi boyunca hücre geçirgenliği için izin verilir, ancak tüm gözenekler tamamen birbirine bağlandığı veya gözenek (homojen) daha düzenli boyutu vardı. Sonra küresel morfolojileri ile biyouyumlu nematik LCE elastomerler bildirildi. Bu nematik elastomerler bağlanma ve hücre proliferasyonu için izin verilir, ancak gözenek boyutu önlenir 10-30 um, sadece değişmektedir veya bunların kullanımı sınırlıdırhücre çizgileri 19, 20 daha geniş bir çeşitliliği ile elastomerler. Kung ve arkadaşları tarafından önceki iş. Bir &quot;gözden çıkarılabilir&quot; Metal şablonu kullanılarak grafin köpüklerin oluşumuna ilişkin elde edilen grafin köpük seçilen metal şablon 22 tarafından dikte çok düzenli bir gözenekli morfolojiyi sahip olduğunu göstermiştir. Bu metodoloji gözeneklilik ve gözenek boyutu tam kontrol sunuyor. Farklı şablon oluşumu, köpük hazırlama öncesinde şekiller aynı zamanda, bir metal şablon yumuşaklık ve esneklik sağlar. Bu malzeme süzme 23, gaz çiftleşmiş 24 veya elektro-eğrilmiş elyaflar 25 gibi diğer teknikler, 26 aynı zamanda gözenekli malzemelerin hazırlanması için bir potansiyel sunmaktadır, ancak daha fazla zaman, bazı durumlarda, gözenek boyutu ile sınırlıdır, alıcı ve vardır sadece birkaç mikrometre. Köpük-benzeri 3D LCEs metal şablonları daha yüksek bir hücre yükü için izin kullanılarak hazırlandı; geliştirilmiş bir proliferasyon hızı; eş-kültürleme; ve, son olarak değil, daha iyi toplu taşıma yönetimi (yani besin, gazlar ve atık) tam doku gelişimini 27 sağlamak için. Köpük gibi 3B LCEs hücre uyumu geliştirmek için görünür; Bu hücre büyümesi ve hücre yönünü algılayacak LC kolye göre çok muhtemeldir. LCE içinde LC kısımların varlığı LCE iskele içinde hücre konumu ile ilgili olarak, hücre hizalama artırmaktadır. Dikmeler, birlikte (birleşme) 27 birleştiği yerde açık bir yönlendirme gözlenirken hücreler, LCE direklerinden içinde hizalamak. Genel olarak, bir hücre destek ortamı olarak LCE hücre iskeleti platformu ayarlamak için bir elastomer morfolojisi ve elastik özellikleri ve özellikle de o sıralı, uzamsal düzenlemeleri oluşturmak için (bireysel) hücre tipleri hizalanmasını yönlendirmek için fırsatlar sunaryaşayan sistemlere benzer f hücreleri. Yanı sıra sürdürülmesi ve uzun süreli hücre büyümesini ve üremesini yönlendirebilen bir iskele sağlamasının, LCEs hücre yönlendirme ve etkileşimler anında değiştirilebilir dinamik deneyler, izin verir.

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			<td height="21" style="height:21px;width:291px;">1H, 1H, 2H, 2H-perfluorooctyltriethoxysilane</td>
			<td style="width:148px;">Alfa Aesar</td>
			<td style="width:132px;">L16606</td>
			<td style="width:153px;">Silanizing agent</td>
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		<tr height="21">
			<td height="21" style="height:21px;">2-bis(4-hydroxy-cyclohexyl)propane</td>
			<td>TCI</td>
			<td><a href="http://www.tcichemicals.com/eshop/en/us/commodity/B0928/">B0928</a></td>
			<td>Reagent</td>
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			<td height="21" style="height:21px;">2-chlorohexanone&#160;</td>
			<td>Alfa Aesar</td>
			<td>A18613</td>
			<td>Reagent</td>
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			<td height="21" style="height:21px;">2-heptanone&#160;</td>
			<td>Sigma Aldrich</td>
			<td>W254401</td>
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			<td height="21" style="height:21px;">2-propanol&#160;</td>
			<td>Sigma Aldrich</td>
			<td>278475</td>
			<td>Solvent</td>
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			<td height="21" style="height:21px;">3-chloroperbenzoic acid, m-CPBA</td>
			<td>Sigma Aldrich</td>
			<td>273031</td>
			<td>Reagent</td>
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			<td height="21" style="height:21px;">4-dimethylaminopyridine</td>
			<td>Alfa Aesar</td>
			<td>A13016</td>
			<td>Reagent</td>
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			<td height="21" style="height:21px;">4&#39;,6-diamidino-2-phenylindole, DAPI&#160;</td>
			<td>Invitrogen</td>
			<td>D1306</td>
			<td>Nuclear Stain</td>
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			<td height="21" style="height:21px;">5-hexynoic acid&#160;</td>
			<td>Alfa Aesar</td>
			<td>B25132-06</td>
			<td>Reagent</td>
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			<td height="21" style="height:21px;">Acetic acid</td>
			<td>VWR</td>
			<td>36289</td>
			<td>Solvent</td>
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			<td height="21" style="height:21px;">Acetone</td>
			<td>Sigma Aldrich</td>
			<td>34850</td>
			<td>Solvent</td>
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		<tr height="21">
			<td height="21" style="height:21px;">Alcohol 200 proof ACS Grade&#160;</td>
			<td>VWR</td>
			<td>71001-866</td>
			<td>Reagent</td>
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			<td height="21" style="height:21px;">Benzene</td>
			<td>Alfa Aesar</td>
			<td>AA33290</td>
			<td>Solvent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">&#949;-caprolactone&#160;</td>
			<td>Alfa Aesar</td>
			<td>A10299-0E</td>
			<td>Reagent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Chloroform</td>
			<td>VWR</td>
			<td>BDH1109</td>
			<td>Solvent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Cholesterol</td>
			<td>Sigma Aldrich</td>
			<td>C8503</td>
			<td>Reagent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Chromium(VI) oxide</td>
			<td>Sigma Aldrich</td>
			<td>232653</td>
			<td>Reagent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Copper (I) iodide</td>
			<td>Strem Chemicals</td>
			<td>100211-060</td>
			<td>Reagent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">D,L-Lactide&#160;</td>
			<td>Alfa Aesar</td>
			<td>L09026</td>
			<td>Reagent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Dichloromethane</td>
			<td>Sigma Aldrich</td>
			<td>320269</td>
			<td>Solvent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Diethyl ether&#160;</td>
			<td>Emd Millipore</td>
			<td>EX0190</td>
			<td>Solvent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">N,N-Dimethylformamide</td>
			<td>Sigma Aldrich</td>
			<td>270547</td>
			<td>Solvent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Dulbecco&#8217;s modified Eagle medium, DEME&#160;</td>
			<td>CORNING Cellgo</td>
			<td>10-013</td>
			<td>Cell Media</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Ethanol</td>
			<td>Alfa Aesar</td>
			<td>33361</td>
			<td>Solvent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Formaldehyde&#160;</td>
			<td>SIGMA Life Science</td>
			<td>F8775</td>
			<td>Fixative</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Fetal bovine serum, FBS&#160;</td>
			<td>HyClone</td>
			<td>SH30071.01</td>
			<td>Media Component</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Filter paper, Grade 415, qualitative, crepe</td>
			<td>VWR</td>
			<td>28320</td>
			<td>Filtration</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Glycerol</td>
			<td>Sigma Aldrich</td>
			<td>G5516</td>
			<td>Central node (3-arm)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Hexamethylene diisocyanate, HDI</td>
			<td>Sigma Aldrich</td>
			<td>52649</td>
			<td>Crosslinker</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Iron(III) chloride&#160;</td>
			<td>Alfa Aesar</td>
			<td>12357</td>
			<td>Etching agent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Isopropyl alcohol</td>
			<td>VWR</td>
			<td>BDH1133</td>
			<td>Solvent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Methanol</td>
			<td>Alfa Aesar</td>
			<td>L13255</td>
			<td>Solvent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">N,N&#39;-dicyclohexylcarbodiimide</td>
			<td>Aldrich</td>
			<td>D80002</td>
			<td>Solvent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">N,N-Dimethylformamide</td>
			<td>Sigma Aldrich</td>
			<td>270547</td>
			<td>Solvent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Nickel metal template</td>
			<td>American Elements</td>
			<td>Ni-860</td>
			<td>Foam template</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Neuroblastomas cells (SH-SY5Y)</td>
			<td>ATCC</td>
			<td>CRL-2266</td>
			<td>Cell line</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Penicillin streptomycin&#160;</td>
			<td>Thermo SCIENTIFIC</td>
			<td>15140122</td>
			<td>Antibiotics</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Polyethylene glycol 2000, PEG</td>
			<td>Alfa Aesar</td>
			<td>B22181</td>
			<td>Reagent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Sodium azide&#160;</td>
			<td>VWR</td>
			<td>97064-646</td>
			<td>Reagent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Sodium bicarbonate</td>
			<td>AMRESCO</td>
			<td>865</td>
			<td>Drying salt</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Sodium chloride</td>
			<td>BDH</td>
			<td>BDH9286</td>
			<td>Drying salt</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Sodium phosphate dibasic heptahydrate</td>
			<td>Fisher Scientific</td>
			<td>S-374</td>
			<td>Drying salt</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Sodium phosphate monobasic monohydrate</td>
			<td>Sigma Aldrich</td>
			<td>S9638</td>
			<td>Drying salt</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Sodium sulfate</td>
			<td>Sigma Aldrich</td>
			<td>239313</td>
			<td>Drying salt</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Tetrahydrofuran</td>
			<td>Alfa Aesar</td>
			<td>41819</td>
			<td>Solvent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Thiosulfate de sodium</td>
			<td>AMRESCO</td>
			<td>393</td>
			<td>Drying salt</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Tin(II) 2-ethylhexanoate</td>
			<td>Aldrich</td>
			<td>S3252</td>
			<td>Reagent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Toluene&#160;</td>
			<td>Alfa Aesar</td>
			<td>22903</td>
			<td>Solvent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Triethylamine</td>
			<td>Sigma Aldrich</td>
			<td>471283</td>
			<td>Reagent</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Trypsin&#160;</td>
			<td>HyClone</td>
			<td>SH30042.01</td>
			<td>Cell Detachment</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Olympus FV1000</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
	</tbody>
</table>

synthesis of biocompatible liquid crystal elastomer foams as cell scaffolds for 3d spatial cell cultures

Burada, bir 3D, biyolojik olarak parçalanabilen, köpük-benzeri hücre iskelesi bir adım adım hazırlanmasını sunmaktadır. Bu iskeleleri Smektik-A (SmA), sıvı kristal elastomerler (LCEs) &#39;de elde edilen, yan zincire asılı gruplar halinde kolesterol birimleri içeren yıldız blok ko-polimerler çapraz bağlama ile hazırlanmıştır. Köpük gibi iskeleleri, 3D hücre kültür yapı iskeleti olarak kullanılmasını uygun hale metal şablonları kullanılarak hazırlandı birbirine mikrokanallar bulunmaktadır. Metal köpük ve geleneksel gözenekli şablonu filmler, ancak kitlesel taşıma aynı zamanda daha iyi yönetimi (örneğin, besin maddeleri, gaz, atık ile karşılaştırıldığında yalnızca yüksek hücre çoğalmasını teşvik eden bir 3D cep iskele elastomer sonuç normal yapının birleşik özellikleri vb.) Metal şablon doğası köpük şekillerde (örneğin, rulo veya film) kolayca müdahale edilebilmesini sağlar ve farklı hücre çalışmaları için farklı gözenek boyutlarının ıskelesının interconnec koruyarakşablonun ted gözenekli doğası. Dağlama işlemi, bunların biyolojik olarak uyumlu ve biyolojik olarak parçalanabilir doğa koruma, elastomerlerin kimyası etkilemez. Biz hücrelerin büyümesini ve çoğalmasını teşvik ederken bu smektik LCEs, geniş süreler boyunca büyüyünce klinik açıdan ve karmaşık doku yapıları çalışma sağlar göstermektedir.

Bu rapor, bir nikel şablonu kullanılarak hücre kültürü için bir iskele olarak gözenekli bir 3D LCE hazırlanması yöntemini göstermektedir. Elde edilen 3D LCE kolay bir hücre infiltrasyonu sağlar kompleks birbirine kanal ağı, hem de daha uygun bir kütle transferini 27 göstermektedir. Bu hücreler, tam olarak birbirine bağlanmış kanal ağ nüfuz edebilmek ve aynı zamanda LCE içinde hizalamak için mümkün olduğu bulunmuştur. Burada, bir meta...

Watch this Scientific Journal Video about Synthesis of Biocompatible Liquid Crystal Elastomer Foams as Cell Scaffolds for 3D Spatial Cell Cultures at JoVE.com

Synthesis of Biocompatible Liquid Crystal Elastomer Foams as Cell Scaffolds for 3D Spatial Cell Cultures

Bu çalışma, 3D, biyo-uyumlu yan zincir sıvı kristal elastomerler (LCEs) göre biyolojik olarak parçalanabilen, köpük-benzeri hücre iskeleleri hazırlanması için bir yöntem sunar. Konfokal mikroskopi deneyleri köpüğümsü LCEs hücre eki, proliferasyon ve C2C12s miyoblastların kendiliğinden uyum sağlamak olduğunu göstermektedir.

3D Mekansal Hücre Kültürleri için hücre yapı iskeleleri olarak biyo-uyumlu sıvı kristal Elastomer Köpüklerin sentezi

Here, we present a step-by-step preparation of a 3D, biodegradable, foam-like cell scaffold. These scaffolds were prepared by cross-linking star block ...

The goal of this procedure is to prepare biodegradable, three dimensional foam-like cell scaffolds based on biocompatible side chain liquid crystal elastomers. This method can help answer key questions in the liquid crystal and biomedical fields, such as the effects of liquid crystal elastomer properties on cell proliferation and alignment. The main advantage of this method of two dimensional cell scaffolds is it permits the study of spatial cell-cell interactions which is rarely possible in 2D macro environments.Generally, those new to this method will struggle with the tactile compression tests. We first had the idea for this method after having to change the media in hundreds of Petri dishes. We wondered if liquid crystal elastomers could support muscle cells on a 3D network, to eliminate the use of so many Petri dishes.Visual demonstration of this method is critical, as some steps require careful manipulation of chemicals. Further, shaping the metal foam template may be challenging. First, fill a 20 milliliter ampoule with a 2%by volume solution of PFOTES in toluene.Stir the solution in the ampoule for 24 hours to silanize the ampoule interior. Rinse the silanized ampoule with isopropyl alcohol and dry it at 140 degrees celsius for 30 minutes. Place 3.64 grams of distilled epsilon-caprolactone, 0.5 grams of alpha-chloro-epsilon-caprolactone, and 0.25 milliliters of glycerol in the dry ampoule.Vortex the mixture for one minute. Then, add 4.90 grams of DL-lactide to the mixture, and purge the ampoule atmosphere with nitrogen gas for one minute. Cover the ampoule opening with aluminum foil, and heat the mixture at 120 degrees Celsius for about two hours to melt the DL-lactide.Vortex the mixture to disrupt any unmelted solids, and then, add 66 microliters of 10-2-ethylhexanoate to the ampoule, and vortex again. Close the ampoule with aluminum foil and heat the mixture at 120 degrees Celsius for 10 minutes to remelt the DL-lactide. Once the DL-lactide has melted again, vigorously vortex the mixture, and purge the ampoule atmosphere with nitrogen gas.Seal the ampoule with a rubber septum. Insert a needle connected to a vacuum line through the septum, and start the vacuum. Flame-seal the neck of the ampoule, being careful not to melt the rubber stopper.Once the neck is sealed, heat the reaction mixture at 140 degrees Celsius for 48 hours. Then, allow the mixture to cool to room temperature. Break open the ampoule and dissolve the viscous reaction mixture in 10 milliliters of dichloromethane.Transfer the mixture to a separatory funnel. Place a flask containing 100 milliliters of methanol in a dry ice and acetone bath to cool down the methanol to approximately minus 78 degrees Celsius. Once the methanol has cooled down, secure the separatory funnel over the flask.Add the reaction mixture to the cold methanol at a rate of two drops every second. Collect the resulting white precipitate on filter paper. Dry the precipitate in a vacuum oven between 50 and 60 degrees Celsius to obtain the three-arm alpha-chloro SBC product.To prepare the alpha cholesteryl three-arm SBC, the pendant chlorine atom is replaced by an azide. A click reaction with cholesteryl 5-hexynoate results in a pendant cholesteryl as the liquid crystal moiety. To begin preparing the liquid crystal elastomer scaffold, combine 0.75 grams of alpha-cholesteryl three-arm SBC with 0.25 milliliters of HDI and 0.24 milliliters of distilled epsilon-caprolactone.Add to this 60 microliters of 10-2-ethylhexanoate and vortex the mixture. Then, cut out a one centimeter by four centimeter piece of nickel metal foam. Roll the nickel foam into a cylinder one centimeter in diameter and one centimeter tall to form a template for the liquid crystal elastomer foam scaffold.Place the template in a glass vial or aluminum foil enclosure, and pour the liquid crystal elastomer mixture over the template until it is completely covered. Allow the template to sit in the liquid crystal elastomer mixture for two minutes, and then, remove the excess with a pipette. Heat the mixture and template at 80 degrees Celsius overnight.Then, peel away the aluminum foil, or break the glass. Use a razor blade to remove excess liquid crystal elastomer to expose the nickel metal template. Place the foam in a flask and add 70 milliliters of a saturated aqueous solution of iron three chloride.Stir the foam in the solution for three days at room temperature to dissolve the nickel template. Every 24 hours, stir the foam in deionized water for 30 minutes, and then, resume stirring in a fresh iron three chloride solution. After the second day of stirring, perform a tactile compression test on the liquid crystal elastomer foam.Resistance to tactile compression indicates that the nickel template is still present in the foam. After the third day, all nickel template has been eliminated, and the resulting foam appears very soft and easy to fully compress. The nickel template must be completely eliminated.It is important to thoroughly rinse the liquid crystal elastomer foam in iron chloride until it's soft to the touch when a tactile compression test is performed. The liquid crystal elastomer foam scaffold is now ready to be characterized and used. Once the liquid crystal elastomer foam is soft, rinse it with 70%ethanol to sterilize it.To begin the seeding procedure, wash again the liquid crystal elastomer foam scaffolds twice in one milliliter of 70%ethanol to sterilize the elastomer surfaces. Then, irradiate the scaffolds with UV light for 10 minutes. Wash the scaffolds with another one milliliter portion of 70%ethanol.Rinse the scaffolds with one milliliter each of sterile water and phosphate-buffered saline. Load the sterilized liquid crystal elastomer scaffolds into 24-well culture plates. Prepare and count a suspension of the cells of interest in the appropriate cell growth medium with penicillin and streptomycin.Dilute the cell suspension to 1.5 times 10 to the fifth cells per 100 microliters using growth medium. Deposit a drop of diluted cell suspension on top of each liquid crystal elastomer scaffold. Incubate the seeded liquid crystal elastomer scaffolds at 37 degrees Celsius in a 5%CO2 atmosphere for two hours.Add another 0.5 milliliters of growth medium to each scaffold, and continue incubation. Every 48 hours, wash the seeded scaffolds with one milliliter of PBS, and add fresh growth medium. Continue incubating the scaffolds until the cells are ready for microscopy.These smetic LCs enable the study of complex tissue constructs while promoting cell growth and proliferation. In order to maintain cell viability, foams must be sterilized and cleanliness of cultures maintained at all times. To prepare for microscopy, fix the cells on the scaffolds with a solution of 4%paraformaldehyde in PBS for 15 minutes.Soak the samples three times in three milliliters of PBS for five minutes. Place one scaffold sample in an Eppendorf tube. Stain the sample with a solution of 0.1%DAPI in 500 microliters of PBS for 10 minutes.Soak the sample twice in one milliliter of PBS for five minutes. And then, immediately image the sample with confocal microscopy. Acquire image stacks spanning the sample and analyze the data in an image processing program.A three-arm SBC with liquid crystal moieties was cast on a nickel foam template with HDI as the cross-linker. The nickel template was removed by etching to obtain the liquid crystal elastomer foam. Compression deformation testing was periodically performed to monitor the etching process.Once the nickel had been completely dissolved, the compression deformation testing showed a 70%reduction in size when compressed. Upon releasing compression, the liquid crystal elastomer foam consistently recovered its original size and shape. This behavior is attributed to the liquid crystal moieties, as similar elastomer foams lacking the cholesterol epsilon-caprolactone moiety did not recover from compression.SCM of the internal foam morphology showed an interconnected network of hollow foam struts. The regular morphology was attributed to the structure of the nickel foam, indicating that pore size and overall shape can be controlled by selection of an appropriate metal template. The foam was seeded with neuroblastoma cells, which attached to the walls of the network within two days.30 days after seeding, confocal microscopy revealed that the cells extended over the liquid crystal elastomer network, and had formed multiple layers dispersed throughout the elastomer foam. Elongation of cell nuclei was observed, and was correlated to cell alignment. Once mastered, this technique can be done within three to four weeks if performed properly.After watching this video, you should have a good understanding of how to design and prepare liquid crystal elastomer cell scaffolds with specific pore sizes and morphologies. While attempting this procedure, it's important to fully characterize each product and to monitor cell viability and expansion. Further, proper staining technique is important to easily acquire confocal microscopy images.Following this procedure, liquid crystal elastomers can also be exposed to external stimuli, such as stress, or magnetic and electric fields, to answer additional questions about how the anisotropic molecular ordering of the liquid crystal elastomers affects cell response. The implications of this technique apply to the study of disease processes, as it provides a platform to dynamically study the effects of external agents on simulated disease activity, and subsequent repair processes. Though this smectic liquid crystal elastomer foam can provide insight into cell-cell interactions, they can also be used with other materials, such as semiconductor or metal nanostructures.After its development, this technique paved the way for researchers in biomedical fields to design long-term experimental platforms mirroring living systems.

Research

JoVE Journal

Bioengineering

Alginate Microcapsule as a 3D Platform for Propagation and Differentiation of Human Embryonic Stem Cells (hESC) to Different Lineages

Alginate Microcapsule as a 3D Platform for Propagation and Differentiation of Human Embryonic Stem Cells hESC to Different Lineages

Farklı soy İnsan Embriyonik Kök Hücreleri (hESC) Yayılması ve Farklılaşma için 3D Platformu olarak aljinat Mikrokapsül

Human embryonic stem cells (hESC) are emerging as an attractive alternative source for cell replacement therapy since they can be expanded in culture ...

Biyomühendislik

Preparation of 3D Fibrin Scaffolds for Stem Cell Culture Applications

3D Fibrin hazırlanması Kök Hücre Kültürü Uygulamaları için iskeleleri

Stem cells are found in naturally occurring 3D microenvironments in vivo, which are often referred to as the stem cell niche 1. Culturing stem cells ...

Self-reporting Scaffolds for 3-Dimensional Cell Culture

3-Boyutlu Hücre Kültürü için Self-raporlama iskeleleri

Culturing cells in 3D on appropriate scaffolds is thought to better mimic the in vivo microenvironment and increase cell-cell interactions. The resulting ...

Fabrication of Mechanically Tunable and Bioactive Metal Scaffolds for Biomedical Applications

Biyomedikal Uygulamalar için Mekanik ayarlanabilir ve Biyoaktif Metal iskelelerinin Fabrikasyon

Biometal systems have been widely used for biomedical applications, in particular, as load-bearing materials. However, major challenges are high stiffness ...

Alternating Magnetic Field-Responsive Hybrid Gelatin Microgels for Controlled Drug Release

Kontrollü İlaç Release için Manyetik Alan-Duyarlı Hibrid Jelatin mikroj Alternatif

Magnetically-responsive nano/micro-engineered biomaterials that enable a tightly controlled, on-demand drug delivery have been developed as new types of ...

Preparation of Monodomain Liquid Crystal Elastomers and Liquid Crystal Elastomer Nanocomposites

Monodomain Sıvı Kristal Elastomerler ve Likit Kristal Elastomer Nanokompozitlerinin Hazırlanması

LCEs are shape-responsive materials with fully reversible shape change and potential applications in medicine, tissue engineering, artificial muscles, and ...

Micropatterning and Assembly of 3D Microvessels

Micropatterning ve 3D mikrodamarlar Meclisi

In vitro platforms to study endothelial cells and vascular biology are largely limited to 2D endothelial cell culture, flow chambers with polymer or glass ...

Fabrication of Extracellular Matrix-derived Foams and Microcarriers as Tissue-specific Cell Culture and Delivery Platforms

Doku spesifik Hücre Kültürü ve Delivery Platform olarak Ekstrasellüler Matriks kaynaklı Köpükler ve mikro Fabrikasyon

Cell function is mediated by interactions with the extracellular matrix (ECM), which has complex tissue-specific composition and architecture. The focus ...

A Microcontroller Operated Device for the Generation of Liquid Extracts from Conventional Cigarette Smoke and Electronic Cigarette Aerosol

Sıvı özleri geleneksel sigara duman ve elektronik sigara Aerosol gelen nesil için aygıt mikroişlemci işletilen

Electronic cigarettes are the most popular tobacco product among middle and high schoolers and are the most popular alternative tobacco product among ...

3D Analysis of Multi-cellular Responses to Chemoattractant Gradients

Chemoattractant degradelere Multi-hücresel tepkiler 3D Analizi

Various limitations of 2D cell culture systems have sparked interest in 3D cell culture and analysis platforms, which would better mimic the spatial and ...