Micropattern width dependent sarcomere development in human ESC-derived cardiomyocytes.
Micropattern width dependent sarcomere development in human ESC-derived cardiomyocytes.
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DOI：
10.1016/j.biomaterials.2014.02.001
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发表时间：
2014-05
期刊：
影响因子：
14
通讯作者：
Crone, Wendy C.
中科院分区：
文献类型：
--
作者：
Salick, Max R.;Napiwocki, Brett N.;Sha, Jin;Knight, Gavin T.;Chindhy, Shahzad A.;Kamp, Timothy J.;Ashton, Randolph S.;Crone, Wendy C.
关键词：
在这项研究中，人类胚胎干细胞来源的心肌细胞被种植到可控的二维微图案化特征上，并在特定特征几何形状中观察到肌节形成和细胞排列的改善。利用高分辨率光刻技术和微接触印刷来产生各种矩形几何形状的特征，面积从2,500μm2到160,000μm2不等。采用微接触印刷法，在镀金玻片上形成聚乙二醇非粘附区的图案。Matrigel和纤维连接蛋白细胞外基质(ECM)蛋白被层叠在镀金玻璃片上，为细胞黏附提供了受控的几何形状。我们使用基于小分子的分化和抗生素纯化步骤，从H9人胚胎干细胞中分离出纯净的未成熟心肌细胞。然后，我们将这一纯人类心肌细胞群体播种到不同大小的微图案化特征上，并观察心肌细胞如何根据特征几何形状重塑其肌丝结构。免疫荧光检测α-肌动蛋白的表达，鬼臼蛋白染色检测肌动蛋白在图案化细胞中的存在。核排列分析也被用来确定细胞方向是如何受到这些特征的影响的。播种的细胞显示出与特征的清晰对齐，这取决于特征的宽度而不是整体特征的纵横比。结果表明，宽度在30μm和80μm之间的特征促进了高度排列的心肌细胞，肌节相对于图案长轴的排列显著增加。这种具有强大肌节结构的高度排列的细胞聚集体的建立在推进基于细胞的药理学研究方面具有巨大的潜力，并将帮助研究人员了解ECM几何形状如何影响hESC来源的心肌细胞的肌丝结构和成熟。
In this study, human embryonic stem cell-derived cardiomyocytes were seeded onto controlled two-dimensional micropatterned features, and an improvement in sarcomere formation and cell alignment was observed in specific feature geometries. High-resolution photolithography techniques and microcontact printing were utilized to produce features of various rectangular geometries, with areas ranging from 2,500μm2 to 160,000μm2. The microcontact printing method was used to pattern non-adherent poly(ethylene glycol) regions on gold coated glass slides. Matrigel and fibronectin extracellular matrix (ECM) proteins were layered onto the gold-coated glass slides, providing a controlled geometry for cell adhesion. We used small molecule-based differentiation and an antibiotic purification step to produce a pure population of immature cardiomyocytes from H9 human embryonic stem cells. We then seeded this pure population of human cardiomyocytes onto the micropatterned features of various sizes and observed how the cardiomyocytes remodeled their myofilament structure in response to the feature geometries. Immunofluorescence was used to measure α-actinin expression, and phalloidin stains were used to detect actin presence in the patterned cells. Analysis of nuclear alignment was also used to determine how cell direction was influenced by the features. The seeded cells showed clear alignment with the features, dependent on the width rather than the overall aspect ratio of the features. It was determined that features with widths between 30μm and 80μm promoted highly aligned cardiomyocytes with a dramatic increase in sarcomere alignment relative to the long axis of the pattern. This creation of highly-aligned cell aggregates with robust sarcomere structures holds great potential in advancing cell-based pharmacological studies, and will help researchers to understand the means by which ECM geometries can affect myofilament structure and maturation in hESC-derived cardiomyocytes.
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影响因子：
--
作者：
Bray, Mark-Anthony;Sheehy, Sean P.;Parker, Kevin Kit
通讯作者：
Parker, Kevin Kit
影响因子：
14
作者：
Feinberg, Adam W.;Alford, Patrick W.;Jin, Hongwei;Ripplinger, Crystal M.;Werdich, Andreas A.;Sheehy, Sean P.;Grosberg, Anna;Parker, Kevin Kit
通讯作者：
Parker, Kevin Kit
影响因子：
2.8
作者：
Cimetta, Elisa;Pizzato, Sara;Elvassore, Nicola
通讯作者：
Elvassore, Nicola
DOI：
10.1073/pnas.0903269107
发表时间：
2010-03-16
影响因子：
11.1
作者：
Kilian, Kristopher A.;Bugarija, Branimir;Mrksich, Milan
通讯作者：
Mrksich, Milan
影响因子：
--
作者：
FERNANDEZTERAN, MA;HURLE, JM
通讯作者：
HURLE, JM