Name: in vitro model of human cutaneous hypertrophic scarring using macromolecular crowding
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这项工作得到了新加坡科学、技术和研究机构"SPF 2013/004：皮肤生物学基础研究"和"热带病护理创新"IAF-PP/2017 （HBMS） H17/01/a0/009 的资助。作者感激地感谢保拉·本尼博士和迈克尔·拉古纳博士的建议和帮助。

1. 细胞培养
<ol><li>在Dulbecco的改良鹰介质（DMEM）中，在含有10%胎儿小牛血清（FCS）和1%v青霉素/链霉素溶液（P/S）的培养箱中，在37°C下，在5%CO2/95%空气中，保持从非病理组织（hNSFs）衍生的hHSF和正常皮肤2纤维细胞。</li>
	<li>从相应的公司购买菲科尔70，菲科尔400，抗坏血酸。</li>
</ol>2. 构建MMC肥大疤痕模型
<ol><li>将 hHSF 或 hNSF（50，000/well）放入 24 孔板中，每个井中含?...

<ol>
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该协议旨在优化和验证人类皮骨疤组织改进的"疤痕在一罐"体外模型。此前的研究曾报告将MMC技术应用于人类肺成纤维细胞12个，人类骨髓中皮干细胞23个，以及人类皮肤纤维细胞23个，使用dextran12、Ficoll12和PVP23作为23人群。在此报道的研究中，先前公布的肥大疤痕衍生人类皮肤成纤维细胞?...

疤痕组织代表组织修复的终点。然而，在许多个人，特别是那些遭受烧伤或创伤1，疤痕可能是过度的，并施加不良的影响，愈合的皮肤的形态和功能。虽然病理（肥大疤痕和酮）疤痕形成的确切机制尚未完全理解，但胶原蛋白在伤口愈合过程中的过度沉积已被证明是一个必不可少的贡献2。
众所周知，转化生长因子β 1 （TGF-+1） 和 α 平滑肌肉作用素 （#SMA） 在增肥疤痕的形成中起着关键作用。有证据表明，升高的TGF-β1通过调节SMAD信号通路3直接刺激胶原蛋白的过度沉积。此外，在伤口愈合过程中，通过调节细胞收缩和重皮化，发现αSMA有助于肥大疤痕的形成。缺乏适当的体外和体内模型是开发和评估疤痕补救干预措施和疗法的主要障碍。本研究的目的是利用现有的MMC技术，构建一个"体内类似"肥肥疤痕模型，适合评估新的和新兴的疤痕相关干预。
多年来，在科学界，在身体外复制活组织一直是一个目标。二十世纪初体外技术的发展部分实现了这一目标。目前的体外技术从Roux最初的证明中略有进化，即胚胎细胞可以在温暖的盐水5中存活几天。然而，体外方法大多限于在二维培养的单细胞类型，并且不能准确地重述体内的细胞组织。虽然可用于检查细胞生物化学、生理学和遗传学，但原生组织是三维组织，并包含多种细胞类型。简单的二维体外系统将哺乳动物细胞置于高度人工的环境中，其中原生组织特定的结构在6中丢失。反过来，这影响细胞内和细胞外事件，导致异常细胞形态，生理和行为7。
该协议背后的兴趣在于肥肥疤痕和酮类物的开发和临床管理。虽然众所周知，皮肤纤维细胞在很大程度上是导致疤痕组织中存在胶原蛋白的大量生产的原因，但使用二维体外系统培育皮肤纤维细胞不能重现体内8中观察到的胶原蛋白的周转。当代体外方法基本上仍然使用"暖盐水"，这种环境与活体组织中的环境完全不同。体内组织非常拥挤，蛋白质、核酸、核糖蛋白和多糖，占总体积的5%-40%。由于没有两个分子可以同时占据相同的空间，因此几乎没有可用的可用空间，而且几乎完全没有自由水9。
MMC 技术施加了影响细胞溶质和间质流体热力学特性的限制。分子相互作用、受体-配体信号复合物、酶和细胞器被限制和限制自由相互作用。近细胞环境中的相互作用（即间室）也受到限制。近期证据证实，高浓度惰性大分子在拥挤的溶液扰动扩散，物理关联，粘度和水动力特性10。
有趣的是，几种流行的挤水剂（即菲科尔、dextran、聚乙烯丙酮[PVP]和钠4苯乙烯硫化酸[PSS]）在应用于不同的细胞类型和不同设置时不等效。在以前的一项研究中，菲科尔与PVP相比，中质干细胞的细胞毒性较小。这些结果被解释为其中性电荷和相对较小的水动力半径11的结果。与此相反，第二项研究发现，与Ficoll12相比，dextran在刺激人肺成纤维细胞的胶原蛋白I沉积方面更有效。来自我们自身研究的数据表明，Ficoll通过肥大疤痕衍生成纤维细胞增强胶原蛋白沉积，而PVP对这些细胞13有毒。
实践证明，在高度拥挤的体内环境中，原胶原蛋白转化为胶原蛋白的速度更快，而生物反应的速度在稀释的二维培养系统15中延迟。我们在这里优化了体外方案，纳入了MMC，以表明皮肤纤维成纤维细胞的培养，作为皮肤纤维化和疤痕形成更"体内"的模型。与常见的二维培养系统相比，用MMC培养hHSF能显著刺激胶原蛋白的生物合成和沉积。值得注意的是，纤维化的其他特性（即，增加基质金属蛋白酶[MP]和亲炎细胞因子的表达）也在这个优化的MMC协议13下明显。使用这种方法培养时，结果表明，皮肤细胞重述了体内测量的生理、生化和功能参数。
优化的MMC体外方案已用于评估胶原蛋白和其他ECM蛋白的表达，通过从肥大疤痕真皮分离的皮肤成纤维细胞和未介入的相邻真皮。在体外MMC环境中培养时，观察到hHSFs具有某些特性（即mRNA、生物化学、生理学和表型），类似于体内皮肤增生性疤痕组织。证据表明，物理和化学特性是选择挤出体和优化MMC条件进行体外培养的重要考虑因素。
为了证明原理，MMC协议在这里应用，以定性和定量评估Shikonin及其模拟诱导凋亡的能力。这允许评估这些天然中药（TCM）化合物在管理皮肤疤痕方面的潜在应用。尽管如此，这项体外MMC协议的简单性、成本效益和及时性也满足了欧盟指令2010/63/EU和美国环境保护署（EPA）最近关于消除哺乳动物实验的规定。

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			<td height="21" style="height:21px;width:487px;">0.2 &#956;m filter</td>
			<td style="width:195px;">Sartorius</td>
			<td style="width:119px;">16534</td>
			<td style="width:537px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">2-Mercaptoethanol</td>
			<td style="width:195px;">Sigma-Aldrich</td>
			<td style="width:119px;">M6250</td>
			<td></td>
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			<td height="21" style="height:21px;width:487px;">4&#8217;,6-diamidino-2-phenylindole (DAPI)</td>
			<td style="width:195px;">Thermo Fisher Scientific</td>
			<td style="width:119px;">P36962</td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Alexa Fluor 680</td>
			<td style="width:195px;">Thermo Fisher Scientific</td>
			<td style="width:119px;">A-21076</td>
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			<td height="21" style="height:21px;width:487px;">Alexa Fluor 800</td>
			<td style="width:195px;">Thermo Fisher Scientific</td>
			<td style="width:119px;">A-11371</td>
			<td></td>
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			<td height="21" style="height:21px;width:487px;">alpha smooth muscle actin (&#945;SMA) primary antibody</td>
			<td style="width:195px;">Abcam</td>
			<td style="width:119px;">ab5694</td>
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			<td height="21" style="height:21px;width:487px;">Applied Biosystems 7500 Fast Real-Time PCR System (thermal cycler )</td>
			<td style="width:195px;">Thermo Fisher Scientific</td>
			<td style="width:119px;">4351106</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Ascorbic acid</td>
			<td style="width:195px;">Wako</td>
			<td style="width:119px;">#013-12061</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Bovine serum albumin</td>
			<td style="width:195px;">Sigma-Aldrich</td>
			<td style="width:119px;">#A2153</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Bradford protein assay</td>
			<td style="width:195px;">Bio-Rad</td>
			<td style="width:119px;">500-0006</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Collagen I primary antibody (for immunostaining)</td>
			<td style="width:195px;">Abcam</td>
			<td style="width:119px;">6308</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Collagen I primary antibody (for western blot)</td>
			<td style="width:195px;">Abcam</td>
			<td style="width:119px;">ab21286</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Collagen III primary antibody</td>
			<td style="width:195px;">Abcam</td>
			<td style="width:119px;">ab7778</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Collagen IV primary antibody</td>
			<td style="width:195px;">Abcam</td>
			<td style="width:119px;">ab6586</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Direct Red 80</td>
			<td style="width:195px;">Sigma-Aldrich</td>
			<td>2610108</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Dulbecco&#39;s Modified Eagle&#39;s Medium (DMEM)</td>
			<td style="width:195px;">Life Technologies</td>
			<td style="width:119px;">11996-065</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Fetal calf serum (FCS)</td>
			<td style="width:195px;">Life Technologies</td>
			<td style="width:119px;">6000-044</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Ficoll 400</td>
			<td style="width:195px;">GE HealthCare</td>
			<td style="width:119px;">#17-0300-10</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Ficoll 70</td>
			<td style="width:195px;">GE HealthCare</td>
			<td style="width:119px;">#17-0310-10</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">GAPDH primary antibody</td>
			<td style="width:195px;">Sigma-Aldrich</td>
			<td style="width:119px;">G8795</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Goat Anti-Rabbit secondary antibody</td>
			<td style="width:195px;">Abcam</td>
			<td style="width:119px;">ab97050</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Human hypertrophic scar/normal fibroblasts (hHSF/hNSF)</td>
			<td style="width:195px;">Cell Research Corporation</td>
			<td style="width:119px;">106, 107, 108</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">iScript cDNA Synthesis Kit</td>
			<td style="width:195px;">Bio-Rad</td>
			<td style="width:119px;">#1708890</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">MMP-1 primary antibody</td>
			<td style="width:195px;">Abcam</td>
			<td style="width:119px;">ab38929</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">MMP-13 primary antibody</td>
			<td style="width:195px;">Abcam</td>
			<td style="width:119px;">ab39012</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">MMP-2 primary antibody</td>
			<td style="width:195px;">Abcam</td>
			<td style="width:119px;">ab37150</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">MMP-9 primary antibody</td>
			<td style="width:195px;">Abcam</td>
			<td style="width:119px;">ab38898</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">NanoDrop Microvolume Spectrophotometers</td>
			<td style="width:195px;">Thermo Fisher Scientific</td>
			<td style="width:119px;">N/A</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Nitrocellulose membrane</td>
			<td style="width:195px;">Bio-Rad</td>
			<td style="width:119px;">10484060</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">NuPAGE 4-12% Bis-Tris Protein Gels</td>
			<td style="width:195px;">Thermo Fisher Scientific</td>
			<td style="width:119px;">NP0321BOX</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Odyssey blocking buffer</td>
			<td style="width:195px;">LI-COR Biosciences</td>
			<td style="width:119px;">927&#8211;40000</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Odyssey Fc Imaging System</td>
			<td style="width:195px;">LI-COR Biosciences</td>
			<td style="width:119px;">N/A</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Olympus IX-81 HCS microscope (for immunostaining)</td>
			<td style="width:195px;">Olympus</td>
			<td style="width:119px;">N/A</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Penicillin/streptomycin solution (P/S)</td>
			<td style="width:195px;">Life Technologies</td>
			<td style="width:119px;">15140-122</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">PrimePCR Assays</td>
			<td style="width:195px;">Bio-Rad</td>
			<td style="width:119px;"></td>
			<td>Customized primers pre-coated in 96-well plates based on requirement</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Protease inhibitor cocktail (PIC)</td>
			<td style="width:195px;">Sigma-Aldrich</td>
			<td style="width:119px;">11697498001</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">PVP 360</td>
			<td style="width:195px;">Sigma</td>
			<td style="width:119px;">#PVP360</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">PVP 40</td>
			<td style="width:195px;">Sigma</td>
			<td style="width:119px;">#PVP40</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">RIPA buffer</td>
			<td style="width:195px;">Merck</td>
			<td style="width:119px;">R0278</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">RNeasy Plus Mini Kit</td>
			<td style="width:195px;">QIAGEN</td>
			<td style="width:119px;">#74134</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Sodium vanadate</td>
			<td style="width:195px;">Sigma-Aldrich</td>
			<td style="width:119px;">450022</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Sodium vanadate</td>
			<td style="width:195px;">Sigma-Aldrich</td>
			<td style="width:119px;">450243</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">SpectraMax M5 Multi-Mode microplate reader</td>
			<td style="width:195px;">Molecular Devices</td>
			<td style="width:119px;">N/A</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">SsoAdvanced universal SYBR green supermix</td>
			<td style="width:195px;">Bio-Rad</td>
			<td style="width:119px;">#172-5270</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:487px;">Tween 20</td>
			<td style="width:195px;">Sigma-Aldrich</td>
			<td style="width:119px;">P9416</td>
			<td></td>
		</tr>
	</tbody>
</table>

in vitro model of human cutaneous hypertrophic scarring using macromolecular crowding

研究表明，体内组织中蛋白质、核酸、核糖蛋白、多糖等非常拥挤。以下协议应用大分子挤拥 （MMC） 技术，通过在体外添加中性聚合物（挤出体）来模拟这种生理挤拥。以前使用Ficoll或dextran作为人群的研究表明，使用MMC技术，在WI38和WS-1细胞系中胶原蛋白I和纤维素的表达得到显著增强。然而，这种技术尚未在原发肥疤源人类皮肤成纤维细胞（hHSFs）中得到验证。由于胶原蛋白的过度沉积会产生肥大疤痕，该协议旨在通过将MMC技术与hHSF一起应用，构建富含胶原蛋白的体外增肥疤痕模型。与传统二维（2-D）细胞培养系统相比，这种优化的MMC模型与体内疤痕组织具有更多的相似性。此外，与动物模型相比，它具有成本效益、时间高效且合乎道德。因此，此处报告的优化模型为肥肥疤痕相关研究提供了先进的"体内类似"模型。

在每个实验中进行三重样本，每个实验使用三个个体患者的细胞重复3倍。数据以控制组的百分比表示。应用单向 ANOVA 和 Tukey 的事后测试来分析统计差异（=p = 0.05）。
MMC使用Ficoll在9%FVO（分数体积占用）提高胶原蛋白和胶原蛋白I沉积在hHSF13的总量。如图1A所示，与使用PVP的对照和MMC相比，使用Ficoll在培养后，hHSF的细胞密度显著?...

Watch this Scientific Journal Video about In Vitro Model of Human Cutaneous Hypertrophic Scarring using Macromolecular Crowding at JoVE.com

In Vitro Model of Human Cutaneous Hypertrophic Scarring using Macromolecular Crowding

该协议描述了使用大分子挤拥创建体外人类肥肥疤痕组织模型，类似于在体内条件。在拥挤的大分子环境中培养时，人类皮肤成纤维细胞表现出类似疤痕组织的表型、生物化学、生理学和功能特性。

利用大分子挤出的人类食性增大疤痕的体外模型

It has been shown that in vivo tissues are highly crowded by proteins, nucleic acids, ribonucleoproteins, polysaccharides, etc. The following protocol ...

我们创建了一个个体的胶原蛋白丰富的模型，以模拟细胞使用大分子挤挤技术在体内体验的环境。皮肤是一个拥挤的环境，不像我们已经用于在体外生长细胞的液体环境。与传统的单层细胞培养系统相比，该模型再现了细胞在体内经历的拥挤的分子环境，特别是在大量细胞外基质取代液态水的疤痕组织中。除了肥大性疤痕，这种大分子挤塞技术可用于细胞外基质丰富的研究，如肺纤维化。根据手稿方向生长疤痕衍生和正常的成纤维细胞。然后将它们播种到24个井板中，每孔5万个细胞在一毫升的介质中。在37摄氏度和5%的二氧化碳下孵育板过夜。通过将菲科尔 70、菲科尔 400 和抗坏血酸混合到 10%FCS DMEM 中，准备大分子拥挤或 MMC 介质。在水浴中孵育介质，将人群分散到溶液中。然后用0.2微米过滤器进行消毒。从成纤维细胞中吸出已花的介质，代之以新制作的 MMC 介质。在37摄氏度下孵育细胞6天，每三天更换一次MMC介质。在200毫升蒸馏去ion化水中溶解0.2克直接红80粉末，用一毫升醋酸溶解，准备天狼星红溶液。从细胞中吸出 MMC 介质，并在每井中加入 300 微升的天狼星红溶液。然后将细胞返回孵化器90分钟。孵育后，用自来水轻轻冲洗天狼星红溶液，让板在一夜之间风干。第二天，将200微升0.1摩尔氢氧化钠加入到每井中，并摇动盘子5至10分钟，提取天狼星红。将抽取的污渍的 100 微升转移到透明 96 井板中，并使用微板读取器测量 620 纳米的吸光度。用200微升PBS清洗每口井，用4摄氏度的甲醇将细胞固定10分钟。然后向井中加入3%BSA，在室温下孵育板30分钟。吸气阻断溶液，并添加200微升的抗胶原蛋白一个抗体到每一个井。孵育板90分钟。然后吸吸抗体，然后用PBS洗井三次，每次洗涤5分钟。在每个井中加入200微升山羊抗兔-FITC二级抗体和DAPI。用铝箔盖住板，在室温下孵育30分钟。丢弃二次抗体和DAPI，用PBS重复洗涤。然后在显微镜下直接可视化荧光。使用 PBS 洗涤细胞两次后，将 40 微升的解液缓冲液添加到每个井中，然后用移液器尖端刮擦细胞层。然后将蛋白质酸盐转移到微离心管中，以测量蛋白质浓度。将 10 微克蛋白质加载到 4 至 12% 的 Bis-Tris 蛋白凝胶的每个井中，并在 200 伏特下执行 SDS-PAGE 30 分钟。完成后，通过运行90伏特的西印90分钟，将蛋白质转移到硝基纤维素膜上，注意避免凝胶和膜之间的气泡形成。用 10 毫升的阻滞缓冲液阻断膜。然后在四摄氏度下孵育原抗体过夜。第二天，用0.1%的TBS Tween 20洗五次膜，每次洗涤5分钟。然后在室温下用适合物种的二级抗体孵育膜一小时。使用成像系统重复洗涤并可视化荧光。使用商用RNA提取试剂盒纯化总RNA后，使用微体积分光光度计测量RNA浓度。然后使用cDNA合成试剂盒进行第一链DNA合成。将cDNA与10微升的SYBR绿色超级混合混合，将样品转移到定制96井板预涂的寡核苷酸底剂。将总体积调整为每井 20 微升水，并根据手稿指示运行 RT-PCR。与使用 PVP 或对照组相比，使用 Ficoll 培养后，肥大疤痕衍生的人类皮肤成纤维细胞的细胞密度显著增加。与其他配方相比，与Fifiol的巨量拥挤在9%的分量占用率显著地增加了胶原蛋白的沉积，包括胶原蛋白一。定量分析进一步证明了这一点。发现在MMC环境中培养的疤痕衍生成纤维细胞和正常皮肤成纤维细胞，除了胶原蛋白外，还调节ECM物种的表达。值得注意的是，与本地组织相比，基质金属蛋白蛋白或MMP的升高表达在肥大疤痕组织中积累。据观察，MMP2、9和13的表达在大分子拥挤条件下培养的两种细胞培养中均明显上调节。MMP在伤口愈合和疤痕形成、调节 ECM 装配和改造过程中发挥重要作用。白细胞间六和血管内皮生长因子的合成在西方的印迹中是检测不到的。但RT-PCR分析表明，白细胞间六的表达明显上调节，而血管内皮生长因子的表达在MMC条件下培养的成纤维细胞中则受到低调节。在尝试此协议时，选择很少使用 CO 病原体的皮质成纤维细胞非常重要。我们还必须记住，抗坏血酸是MMC介质中的基本成分。我们特别感兴趣的是进一步检测该 MMC 模型中的胶原蛋白沉积和对齐方式。我们计划使用先进的微观技术来比较这种体外模型与体内的原生疤痕组织。我们鼓励其他人使用这种大分子挤占技术，以提高辅助模态组织的体外真实性。当然，大分子拥挤也可以用来重新计算体外疾病和病理学的模型。

Research

JoVE Journal

Bioengineering

Visualizing Proteins and Macromolecular Complexes by Negative Stain EM: from Grid Preparation to Image Acquisition

负染色的EM可视化蛋白质和大分子复合物：从电网的制备图像采集

Single particle electron microscopy (EM), of both negative stained or frozen hydrated biological samples, has become a versatile tool in structural ...

科研

生物工程

Anatomical Reconstructions of the Human Cardiac Venous System using Contrast-computed Tomography of Perfusion-fixed Specimens

人类心脏静脉系统使用对比计算机断层扫描灌注固定的标本解剖重建

A detailed  understanding of the complexity and relative variability within the human  cardiac venous system is crucial for the development of cardiac ...

Cell Co-culture Patterning Using Aqueous Two-phase Systems

利用双水相系统的细胞共培养图案

Cell patterning  technologies that are fast, easy to use and affordable will be required for the  future development of high throughput cell assays, ...

Tissue Engineering of a Human 3D in vitro Tumor Test System

Tissue Engineering of a Human 3D in vitro Tumor Test System

一个人的3D组织工程<em&gt;在体外</em&gt;肿瘤测试系统

Cancer is one of the leading causes of death worldwide.  Current therapeutic strategies are predominantly developed in 2D culture  systems, which ...

In vitro Cell Culture Model for Toxic Inhaled Chemical Testing

In vitro Cell Culture Model for Toxic Inhaled Chemical Testing

对于中吸入有毒化学测试体外细胞培养模型

Cell cultures are indispensable to develop and study efficacy of therapeutic agents, prior to their use in animal models. We have the unique ability to ...

Production, Characterization and Potential Uses of a 3D Tissue-engineered Human Esophageal Mucosal Model

三维组织工程人食管黏膜型号生产，表征和潜在用途

The incidence of both esophageal adenocarcinoma and its precursor, Barrett&#8217;s Metaplasia, are rising rapidly in the western world. Furthermore ...

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

眼刺激试验（EIT）使用重构型人角膜上皮样（RHCE）组织模型眼刺激性化学品危险性鉴定

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

Improving 2D and 3D Skin In Vitro Models Using Macromolecular Crowding

Improving 2D and 3D Skin In Vitro Models Using Macromolecular Crowding

提高2D和3D皮肤<em&gt;体外</em&gt;模型使用大分子拥挤

The glycoprotein family of collagens represents the main structural proteins in the human body, and are key components of biomaterials used in modern ...

Microfluidic Flow Chambers Using Reconstituted Blood to Model Hemostasis and Platelet Transfusion In Vitro

Microfluidic Flow Chambers Using Reconstituted Blood to Model Hemostasis and Platelet Transfusion In Vitro

微流体流钱伯斯再造使用血止血建模和血小板输注<em&gt;体外</em

Blood platelets prepared for transfusion gradually lose hemostatic function during storage. Platelet function can be investigated using a variety of ...

An In Vitro Organ Culture Model of the Murine Intervertebral Disc

An In Vitro Organ Culture Model of the Murine Intervertebral Disc

一个<em&gt;体外</em鼠类的椎间盘&gt;器官培养模型

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