Name: hydrogel arrays enable increased throughput for screening effects of matrix components and therapeutics in 3d tumor models
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Description: Watch this Scientific Journal Video about Hydrogel Arrays Enable Increased Throughput for Screening Effects of Matrix Components and Therapeutics in 3D Tumor Models at JoVE.com

作者要特别感谢Carolyn Kim，Amelia Lao，Ryan Stoutamore和Itay Solomon对光凝胶化方案早期迭代的贡献。细胞系GS122和GS304由David Nathanson慷慨提供。所有数字都是用 BioRender.com 创建的。加州大学洛杉矶分校的核心设施，分子筛选共享资源以及纳米和Pico表征实验室对这项工作起到了重要作用。Chen Chia-Chun得到了加州大学洛杉矶分校Eli和Edythe Broad再生医学和干细胞研究培训计划中心的支持。Grigor Varuzhanyan得到了肿瘤细胞生物学培训计划NIH拨款（T32 CA 009056）的支持。

患者来源的GBM细胞系（GS122和GS304）由David Nathanson教授（我们的合作者）提供，他根据加州大学洛杉矶分校机构审查委员会批准的方案（IRB# 10-000655）开发了这些细胞系。细胞被提供去识别化，以便细胞系不能与个体患者联系起来。
1. 水凝胶溶液的制备
<ol><li>通过将 HEPES 粉末以 20 mM 溶解在 Hank 的平衡盐溶液 （HBSS） 中来制备 HEPES 缓冲溶液。完全溶剂化后将pH值?...

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目前的工作提出了在基于HA的基质硬度和可用于整合素参与的肽中生成3D小型化培养物的方法。该技术能够系统地研究基质参数如何影响细胞表型（例如，暴露于化疗的癌细胞的活力），并提高通量。先前的方法，包括本文提出的方法，已经通过改变最终配方中总聚合物的百分比来调整水凝胶刚度，其中较硬的水凝胶具有更高的聚合物含量21，31。然而，...

开发新药的预期成本在过去十年中稳步上升，目前估计超过10亿美元1。这笔费用的一部分是进入临床试验的药物的高失败率。大约12%的候选药物最终在2019年获得美国食品和药物管理局（FDA）的批准。许多药物由于意想不到的毒性2而在I期失败，而其他通过安全性试验的药物可能由于缺乏疗效而失败3。由于无效而导致的这种损耗可以部分解释为，在药物开发过程中使用的癌症模型对临床疗效是出了名的不预测4。
体外和体内模型之间的功能差异可归因于从其天然微环境中去除癌细胞，包括非肿瘤细胞和物理ECM5，6。通常，研究小组使用市售的培养基质，例如Matrigel（一种来自小鼠肉瘤的蛋白质基底膜基质）为培养的肿瘤细胞提供3D基质微环境。与2D培养相比，膜基质中的3D培养提高了体外结果的临床相关性7，8。然而，来自去细胞化组织（包括膜基质）的培养生物材料通常表现出批次间的变异性，这可能影响重现性9。此外，衍生自具有不同组织起源的肿瘤的基质可能与所研究的那些基质10可能不提供适当的生理线索。最后，具有高度肿瘤内异质性的癌症具有微环境特征，这些特征在亚微米级尺度上变化，并且膜基质无法调整以重述11。
胶质母细胞瘤（GBM）是一种致死性均值的脑肿瘤，中位生存时间约为15个月，是一种治疗开发特别困难的癌症12，13。目前GBM的治疗标准包括原发性肿瘤切除术，然后放疗，然后使用替莫唑胺（TMZ）进行化疗14。然而，超过一半的临床GBM肿瘤通过各种机制表现出治疗耐药性15，16，17。预测治疗方案对个体患者的疗效是极其困难的。用于预测个体结果的标准临床前模型包括将患者来源的肿瘤细胞异位移植到免疫功能低下的小鼠中。虽然患者来源的异种移植物可以概括临床GBM肿瘤的许多方面，并且对于临床前模型18很有价值，但它们本质上是昂贵的，低通量的，耗时的，并且涉及伦理问题19。在2D塑料表面上或作为球体培养患者来源的细胞，大多避免了这些问题。虽然患者来源的细胞保留了遗传畸变，但它们在2D或悬浮球体中的培养物在很大程度上在啮齿动物和原始患者肿瘤中患者来源的异种移植物的代表性较差20。以前，我们和其他人已经证明，在模仿脑组织的机械和生化特性的3D ECM中培养的GBM细胞可以保留耐药表型10，21，22，23。
透明质酸（HA）是一种在脑ECM中丰富的多糖，在GBM肿瘤中过表达，其CD44受体调节体外耐药性的获得21，24，25，26，27之间的相互作用。例如，在柔软的3D培养物中加入HA增加了患者来源的GBM细胞获得治疗耐药性的能力。这种机械反应性取决于HA与GBM细胞21上的CD44受体的结合。此外，整合素与含RGD肽结合，并入3D培养基质中，以刚度依赖性方式扩增CD44介导的化学抗性21。除HA外，几种ECM蛋白的表达，其中许多含有RGD区域，在正常脑和GBM肿瘤28之间变化。例如，一项研究报告说，28种不同的ECM蛋白在GBM肿瘤中上调29。在这种复杂的肿瘤基质微环境中，癌细胞整合机械和生化线索以产生特定的抗性表型，这取决于Young的整合素结合肽的模量或密度28，29，30的相对较小的差异（例如，小于一个数量级）。
本方案描述了肿瘤细胞如何解释基质线索的独特组合，并鉴定促进治疗耐药性的复杂，患者特异性基质微环境（图1A）。用于生成用于3D培养的小型化，精确调谐的基质的光化学方法提供了一个大的正交可变空间。由微控制器运行的定制LED阵列被整合到384孔板格式中的光交联水凝胶中，以提高自动化和可重复性。暴露强度在井间变化，以改变所得水凝胶的微机械性质，如使用原子力显微镜（AFM）评估的那样。虽然本手稿并不侧重于构建照明阵列本身，但提供了电路图（图1B）和零件清单（材料表）作为器件再现的辅助工具。
该报告展示了在独特的3D微环境中培养的GBM细胞阵列的快速生成，其中Young的模量（单个数量级上的四个水平）和整合素结合肽含量（来自四种不同的ECM蛋白）正交变化。然后，该方法用于研究水凝胶力学和ECM特异性整合素参与对患者来源的GBM细胞的活力和增殖的相对贡献，因为它们对替莫唑胺（TMZ）化疗产生耐药性。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>1.1 kOhm resistors, 6 W</td>
			<td>Digikey</td>
			<td>35601k1ft</td>
			<td></td>
		</tr>
		<tr>
			<td>1.7 mL microcentrifuge tube</td>
			<td>Genesse Scientific</td>
			<td>21-108</td>
			<td></td>
		</tr>
		<tr>
			<td>15 mL conical tube</td>
			<td>Fisher Scientific</td>
			<td>14-959-70C</td>
			<td></td>
		</tr>
		<tr>
			<td>365 nm LED</td>
			<td>Digikey</td>
			<td>ltpl-c034uvh365</td>
			<td></td>
		</tr>
		<tr>
			<td>384 well plate</td>
			<td>Bio Greiner One</td>
			<td>781090</td>
			<td></td>
		</tr>
		<tr>
			<td>40 &#181;m cell strainer</td>
			<td>MTC bio</td>
			<td>C4040</td>
			<td></td>
		</tr>
		<tr>
			<td>4-Armed thiol terminated polyethlene glycol (20 kDa)</td>
			<td>Laysan Bio</td>
			<td>4arm-PEG-SH-20K-1g</td>
			<td></td>
		</tr>
		<tr>
			<td>6 NPN BJTs</td>
			<td>Digikey</td>
			<td>2n5550ta</td>
			<td></td>
		</tr>
		<tr>
			<td>80 Ohm resistors, 0.125 W</td>
			<td>Digikey</td>
			<td>erjj-6enf80r6v</td>
			<td></td>
		</tr>
		<tr>
			<td>8-Armed norbornene terminated polyethylene glycol (20 kDa)</td>
			<td>Jenkem Technology</td>
			<td>A7025-1</td>
			<td></td>
		</tr>
		<tr>
			<td>Accutase</td>
			<td>Innovative Cell Technologies</td>
			<td>AT104500</td>
			<td>&#160;cell dissociation&#160; reagent</td>
		</tr>
		<tr>
			<td>AFM Probes</td>
			<td>Novascan</td>
			<td></td>
			<td>0.01 N/m Nominal spring constant, 2.5 &#181;m SiO2 particle</td>
		</tr>
		<tr>
			<td>Arduino IDE</td>
			<td>Arduino</td>
			<td>1.8.19</td>
			<td></td>
		</tr>
		<tr>
			<td>Arduino Nano</td>
			<td>Makerfire</td>
			<td>Mini Nano V3.0 ATmega328P Microcontroller Board</td>
			<td></td>
		</tr>
		<tr>
			<td>bFGF</td>
			<td>Peprotech</td>
			<td>100-18B</td>
			<td>20 ng/mL</td>
		</tr>
		<tr>
			<td>CCK8</td>
			<td>Abcam</td>
			<td>ab228554</td>
			<td></td>
		</tr>
		<tr>
			<td>Centrifuge</td>
			<td>Thermoscientific</td>
			<td>sorvall legend xtr</td>
			<td></td>
		</tr>
		<tr>
			<td>CP100ST</td>
			<td>Gilson</td>
			<td>F148415</td>
			<td>Pipette tips for positive displacement pipette</td>
		</tr>
		<tr>
			<td>Cubis Semi-Micro Balance</td>
			<td>Sartorius</td>
			<td>MSA225S100DI</td>
			<td></td>
		</tr>
		<tr>
			<td>DMEM - F12 (50-50)</td>
			<td>Life Technologies</td>
			<td>11330057</td>
			<td>1x</td>
		</tr>
		<tr>
			<td>DMSO</td>
			<td>Fisher Scientific</td>
			<td>BP231-100</td>
			<td></td>
		</tr>
		<tr>
			<td>DPBS Ca (-) Mg (-)</td>
			<td>Genesse Scientific</td>
			<td>25-508</td>
			<td></td>
		</tr>
		<tr>
			<td>EGF</td>
			<td>Peprotech</td>
			<td>AF100-15</td>
			<td>50 ng/mL</td>
		</tr>
		<tr>
			<td>Ethanol, Anhydrous</td>
			<td>Fisher Scientific</td>
			<td>A405P</td>
			<td>Add DI water to dilute to 70%</td>
		</tr>
		<tr>
			<td>Fisherbrand Class B Amber Glass threaded vials</td>
			<td>Fisher Scientific</td>
			<td>03-339-23C</td>
			<td></td>
		</tr>
		<tr>
			<td>Fisherbrand Weighing Paper</td>
			<td>Fisher Scientific</td>
			<td>09-898-12B</td>
			<td></td>
		</tr>
		<tr>
			<td>G21 Supplement</td>
			<td>Gemini Bio</td>
			<td>400-160</td>
			<td>50x</td>
		</tr>
		<tr>
			<td>Hanks Balanced Salt Solution</td>
			<td>Thermo Fisher Scientific</td>
			<td>14175095</td>
			<td></td>
		</tr>
		<tr>
			<td>HCl, ACS, 12M</td>
			<td>Sigma Aldrich</td>
			<td>S25838A</td>
			<td>Add DI water to dilute to 1 M</td>
		</tr>
		<tr>
			<td>Heparin sodium salt from porcine intestinal mucosa</td>
			<td>Sigma Aldrich</td>
			<td>H3149-100Ku</td>
			<td>25 &#181;g/mL</td>
		</tr>
		<tr>
			<td>HEPES</td>
			<td>Sigma Aldrich</td>
			<td>H7006-100G</td>
			<td></td>
		</tr>
		<tr>
			<td>Hot Air Gun</td>
			<td>Wagner</td>
			<td>HT1000</td>
			<td></td>
		</tr>
		<tr>
			<td>Integrin-binding sialoprotein (IBSP) peptide</td>
			<td>Genscript</td>
			<td>Custom Order</td>
			<td>GCGYGGGGNGEPRGDTYRAY</td>
		</tr>
		<tr>
			<td>Lithium phenyl-2,4,6 trimethylbenzoylphosphinate (LAP) , &#62;95%</td>
			<td>Sigma Aldrich</td>
			<td>900889-1G</td>
			<td></td>
		</tr>
		<tr>
			<td>Magnetic stir plate</td>
			<td>Thermo Scientific</td>
			<td>SP194715</td>
			<td></td>
		</tr>
		<tr>
			<td>Microcentrifuge</td>
			<td>Thermo Scientific</td>
			<td>Sorvall legend micro 21R</td>
			<td></td>
		</tr>
		<tr>
			<td>Microman E single Channel Pipettor</td>
			<td>Gilson</td>
			<td>FD10004</td>
			<td>Positive displacement pipette</td>
		</tr>
		<tr>
			<td>Micropipette Tips</td>
			<td>Various Manufacturs</td>
			<td></td>
			<td>Various sizes</td>
		</tr>
		<tr>
			<td>mLine micropipette</td>
			<td>Sartorious</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>N-acetyl Cysteine</td>
			<td>Sigma Aldrich</td>
			<td>A7250-10G</td>
			<td></td>
		</tr>
		<tr>
			<td>Nanowizard 4</td>
			<td>Bruker</td>
			<td></td>
			<td>AFM microscope</td>
		</tr>
		<tr>
			<td>NaOH</td>
			<td>Fisher Scientific</td>
			<td>ss255-1</td>
			<td>Add DI water to dilute to 1 M</td>
		</tr>
		<tr>
			<td>Normoicin</td>
			<td>Invivogen</td>
			<td>ant-nr-1</td>
			<td>500x</td>
		</tr>
		<tr>
			<td>Osteopontin Peptide</td>
			<td>Genscript</td>
			<td>Custom Order</td>
			<td>GCGYGTVDVPDGRGDSLAYG</td>
		</tr>
		<tr>
			<td>Pipet Aid</td>
			<td>Drummond</td>
			<td>4000102</td>
			<td></td>
		</tr>
		<tr>
			<td>Plain Microscope Slides</td>
			<td>Globe Scientific</td>
			<td>1301</td>
			<td></td>
		</tr>
		<tr>
			<td>Press-To-Seal silicone Isolator, 12-4.5mm diam x 2mm deep</td>
			<td>Grace Bio Labs</td>
			<td>664201-A</td>
			<td>Cut so that 8 individual molds are made from a single sheet</td>
		</tr>
		<tr>
			<td>Processing</td>
			<td>Processing</td>
			<td>3.5.4</td>
			<td></td>
		</tr>
		<tr>
			<td>Repeater M4</td>
			<td>Eppendorf</td>
			<td>4982000322</td>
			<td></td>
		</tr>
		<tr>
			<td>Repeater Pipette Tips</td>
			<td>Sartorious</td>
			<td>30089430</td>
			<td>1 mL sizes</td>
		</tr>
		<tr>
			<td>RGD Peptide</td>
			<td>Genscript</td>
			<td></td>
			<td>GCGYGRGDSPG</td>
		</tr>
		<tr>
			<td>Scoth Tape</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Serological Pipettes</td>
			<td>Genesse Scientific</td>
			<td>12-102,12-104</td>
			<td>5,10 mL Pipettes</td>
		</tr>
		<tr>
			<td>Solder Paste</td>
			<td>Digikey</td>
			<td>315-NC191LT15T5-ND</td>
			<td></td>
		</tr>
		<tr>
			<td>Solder Wire</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Straight dissecting forceps</td>
			<td>VWR Scientific</td>
			<td>82027-408</td>
			<td></td>
		</tr>
		<tr>
			<td>Synergy H1 Plate Reader</td>
			<td>Biotek</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>T-75 Cell Culture Treated Flask</td>
			<td>Genesee Scientific</td>
			<td>25-209</td>
			<td></td>
		</tr>
		<tr>
			<td>Temozolomide</td>
			<td>Sigma Aldrich</td>
			<td>T2577</td>
			<td>Typically used from 10 &#181;M to 100 &#181;M</td>
		</tr>
		<tr>
			<td>Tenascin-C Peptide</td>
			<td>Genscript</td>
			<td></td>
			<td>GCGYGRSTDLPGLKAATHYTITIR 
			GV</td>
		</tr>
		<tr>
			<td>Thiolated Hyaluronic Acid (700 kDa), 6-8% modified</td>
			<td>Lifecore Biomedical</td>
			<td>HA700K5</td>
			<td></td>
		</tr>
		<tr>
			<td>VWR Spinbar, Flea Micro</td>
			<td>VWR</td>
			<td>58948-375</td>
			<td></td>
		</tr>
	</tbody>
</table>

hydrogel arrays enable increased throughput for screening effects of matrix components and therapeutics in 3d tumor models

细胞基质相互作用通过生化、机械和几何线索介导复杂的生理过程，影响病理变化和治疗反应。在药物开发管道的早期考虑基质效应预计将增加新疗法临床成功的可能性。基于生物材料的策略在3D细胞培养中重述特定的组织微环境，但将这些与主要用于药物筛选的2D培养方法相结合一直具有挑战性。因此，这里介绍的方案详细介绍了在多孔板格式的小型化生物材料基质中进行3D培养的方法开发，以促进与现有药物筛选管道和常规细胞活力测定的集成。由于对于在培养细胞中保存临床相关表型至关重要的基质特征具有高度的组织和疾病特异性，因此有必要对基质参数进行组合筛选，以确定特定应用的适当条件。这里描述的方法使用小型化培养形式来评估癌细胞对基质力学和配体呈递的正交变化的反应。具体而言，这项研究展示了使用该平台来研究基质参数对患者来源的胶质母细胞瘤（GBM）细胞对化疗反应的影响。

AFM测量结果证实，使用定制的Arduino控制LED阵列，在光交联过程中，水凝胶力学的精确控制是紫外辐照度（mW/cm2）的函数（图2A）。该方案中使用的水凝胶制剂可以在表2中找到。提供的模板上LED的间距与384孔板的所有其他孔的间距相匹配，从而允许在板内形成凝胶（图2B）。对单个水凝胶表面微米级区域的AFM询问表明，平均杨氏模量...

Watch this Scientific Journal Video about Hydrogel Arrays Enable Increased Throughput for Screening Effects of Matrix Components and Therapeutics in 3D Tumor Models at JoVE.com

Hydrogel Arrays Enable Increased Throughput for Screening Effects of Matrix Components and Therapeutics in 3D Tumor Models

本方案描述了一个实验平台，用于评估机械和生化线索对3D基质模拟培养物中患者来源的胶质母细胞瘤细胞的化疗反应的影响，使用定制的UV照明装置促进具有可调机械特征的水凝胶的高通量光交联。

水凝胶阵列可提高 3D 肿瘤模型中基质成分和治疗药物筛选效果的通量

Cell-matrix interactions mediate complex physiological processes through biochemical, mechanical, and geometrical cues, influencing pathological changes ...

这种方法使研究人员能够系统地探测基质线索如何影响3D培养物中细胞的表型。我们使用GBM细胞培养物演示该过程。这种技术的主要优点是高通量实现，这使我们能够筛选许多矩阵条件。该技术可用于开发组织模拟基质，以保留细胞培养模型中的特定功能，例如肿瘤细胞的耐药性，并可能鉴定新的治疗方法。来自Seidlits实验室的本科研究人员Mary Epperson和Kelly Tamura将展示该程序。首先通过在Hanks平衡盐溶液中以20毫摩尔溶解HEPES粉末来制备HEPES缓冲溶液。完全吸附后将pH值调节至7。在HEPES缓冲溶液中，溶解硫醇化HA，使每种葡萄糖醛酸上的6~8%羧酸残基在缓冲溶液中以每毫升10毫克的浓度用硫醇修饰。在室温下使用磁力搅拌板以低于每分钟1， 000次旋转的速度搅拌，直到完全溶解，通常约为45分钟。当HA溶解时，在微量离心管中制备每毫升100毫克8臂PEG降冰片烯，每毫升100毫克4臂PEG硫醇，半胱氨酸或含半胱氨酸肽的四毫摩尔和每毫升LAP的四毫克溶液。此时，准备所有肽的四毫摩尔溶液以拴在单个水凝胶中。混合HA，PEG降冰片烯，PEG硫醇和含半胱氨酸硫醇肽的单个溶液，以达到最终水凝胶基质的最终浓度。在磁性楼梯板上以低于每分钟1， 000圈的速度搅拌至少30分钟，以充分混合。将样品与照明设备对齐，每隔一个LED在硅胶模具或384孔板的单列中对齐。打开 UV LED 参数并输入强度和照明值。然后单击“完成”开始照明。照明后，当放置在一个角时，将孔板移动到下一个角并重复。要照亮板另一半上的孔，请将板从支架中取出并旋转180度。要产生具有不同力学原理的水凝胶，首先使用胶带清洁载玻片和硅胶模具以清除碎屑。将硅胶模具粘在载玻片上，向下按压以确保良好的密封性，并置换任何气泡。接下来，将80微升的水凝胶前驱体溶液移液到载玻片上的每个硅胶模具中。将载玻片放在照明设备上，该设备与单列中的所有其他LED对齐。将水凝胶前体暴露在紫外线下15秒以进行照片交联。照明停止后，取回载玻片，并通过用细尖描摹模具的内圆周将凝胶从模具中松开。用镊子或镊子取下硅胶模具。在12孔板中填充两毫升DPBS。通过润湿刮刀并将其从载玻片上轻轻推开，将交联水凝胶移动到单个孔板中。将所需细胞制备为单细胞溶液。从T-75烧瓶悬浮培养物中收集直径约150微米的胶质母细胞瘤球体到15毫升锥形管中。用五毫升DPBS冲洗培养瓶以除去任何残留的细胞和培养基，并将该体积加入锥形管中。在室温下将含有细胞的锥形管以200倍G离心五分钟。离心后，用五毫升血清移液管除去上清液，注意不要干扰细胞沉淀并重悬于五毫升DPBS中。在室温下以200倍G离心5分钟以洗涤细胞。用五毫升血清学移液管吸出上清液，注意不要干扰细胞沉淀，然后将细胞重悬于两毫升细胞解离试剂中。在室温下孵育10至15分钟。加入三毫升完整的培养基，轻轻移液三到五次，将球体分解成单细胞悬浮液。将单细胞悬浮液以400倍G离心5分钟，以在室温下沉淀细胞。用五毫升血清移液管吸出上清液，注意不要干扰细胞沉淀。将细胞重悬于一毫升完整的培养基中。使用血细胞计数器取出一部分细胞进行计数。用台盼蓝将这部分稀释两倍，其渗透细胞活力受损。只计算活的无色细胞。确定封装所需的电池数量。将含有所需细胞总数的培养基体积转移到无菌的1.7毫升微量离心管中。在室温下以400倍G旋转五分钟。用微量移液管吸出上清液，注意不要干扰细胞沉淀。将细胞沉淀重悬于水凝胶前体溶液中，通过用1， 000微升微量移液管上下移液四到五次来充分混合。将细胞加载到重复的移液器中，以分配10微升。为避免气泡和不均匀的分配，通过在废物容器中再分配一到两次来预处理重复移液器。在384孔板的每个孔中，从重复移液器中分配10微升悬浮在水凝胶溶液中的细胞。使用LED阵列，照亮每个含有细胞的孔15秒钟，以达到所需的机械性能。向每个含有细胞的孔中加入40微升的完整培养基。向凝胶周围的非实验性干井中加入50微升DPBS，以最大限度地减少蒸发造成的损失。对于胶质母细胞瘤细胞，加入40微升含培养基的药物以达到最终所需的浓度，在包封后三天开始。向每个含有细胞的孔中加入10微升CCK8试剂。根据制造商的说明孵育一到四个小时。孵育后所有孔的450纳米处读取吸光度。AFM对单个水凝胶表面微米级区域的询问表明，平均杨氏模量较软的水凝胶的模量范围也比较硬的水凝胶小。与以每毫升500， 000个细胞的密度接种的GS122细胞相比，在培养7天后评估时，以每毫升2， 500， 000个细胞的密度接种的GS122细胞的3D培养物表现出显着更高的活力。当骨桥蛋白衍生的肽包含在基质中时，GS122细胞在8千帕斯卡条件下显示出存活率增加，而整合素结合的sialop蛋白或tenascin C衍生肽的掺入提供了最小的生存益处，例如与生姜RGD肽的培养和基质。相反，没有肽在0.8千帕斯卡培养条件下获得生存率。GS122和GS304细胞在软或硬水凝胶基质（包括含RGD的肽）中培养时扩散。GS122细胞在骨桥蛋白的0.8和8千帕斯卡中都显示出类似的缺乏扩散。然而，GS122仅在8千帕斯卡条件下显示出对替莫唑胺的增强抵抗力。最后，这种小型化的3D培养平台可用于从其他肿瘤类型中培养人类细胞，包括终末分化的神经内分泌前列腺癌细胞的活体类器官。在此程序之后，可以使用诸如单细胞RNA测序之类的进一步技术来进一步表征细胞表型。

Research

JoVE Journal

Bioengineering

Longitudinal Measurement of Extracellular Matrix Rigidity in 3D Tumor Models Using Particle-tracking Microrheology

细胞外基质刚性三维肿瘤模型的纵向测量用粒子跟踪Microrheology

The mechanical microenvironment has been shown to act as a crucial regulator of tumor growth behavior and signaling, which is itself remodeled and ...

科研

生物工程

Porous Silicon Microparticles for Delivery of siRNA Therapeutics

多孔硅微粒进行交付的siRNA治疗的

Small interfering RNA (siRNA) can be used to suppress gene expression, thereby providing a new avenue for the treatment of various diseases. However, the ...

3D Hydrogel Scaffolds for Articular Chondrocyte Culture and Cartilage Generation

3D水凝胶支架的关节软骨细胞培养及软骨生成

Human articular cartilage is highly susceptible to damage and has limited self-repair and regeneration potential. Cell-based strategies to engineer ...

3D Microtissues for Injectable Regenerative Therapy and High-throughput Drug Screening

3D Microtissues 用于可注射再生疗法和高通量药物筛选

To upgrade traditional 2D cell culture to 3D cell culture, we have integrated microfabrication with cryogelation technology to produce macroporous ...

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

改良的3D 水凝胶培养的原发胶质细胞的体外Neuroinflammation 模型的建立

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

Fabricating a Kidney Cortex Extracellular Matrix-Derived Hydrogel

肾皮质细胞外基质衍生水凝胶的制备

Extracellular matrix (ECM) provides important biophysical and biochemical cues to maintain tissue homeostasis. Current synthetic hydrogels offer robust ...

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

用于研究基质细胞代谢串扰的人类3D细胞外基质-脂肪培养模型

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

Fabrication of Ti3C2 MXene Microelectrode Arrays for In Vivo Neural Recording

Fabrication of Ti3C2 MXene Microelectrode Arrays for In Vivo Neural Recording

用于Vivo神经记录的 Ti3C2 MXene 微电极阵列的制造

Implantable microelectrode technologies have been widely used to elucidate neural dynamics at the microscale to gain a deeper understanding of the neural ...

An Open Source Technology Platform to Manufacture Hydrogel-Based 3D Culture Models in an Automated and Standardized Fashion

以自动化和标准化方式制造基于水凝胶的3D培养模型的开源技术平台

Current mixing steps of viscous materials rely on repetitive and time-consuming tasks which are performed mainly manually in a low throughput mode. These ...

Photodegradable Hydrogel Interfaces for Bacteria Screening, Selection, and Isolation

用于细菌筛选、选择和分离的可光降解水凝胶接口

Biologists have long attempted to understand the relationship between phenotype and genotype. To better understand this connection, it is crucial to ...