Micropatterned Biphasic Nanocomposite Platform for Maintaining Chondrocyte Morphology
Micropatterned Biphasic Nanocomposite Platform for Maintaining Chondrocyte Morphology
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DOI：
10.1021/acsami.9b22596
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发表时间：
2020-04-01
影响因子：
9.5
通讯作者：
Wood, Scott T.
中科院分区：
文献类型：
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作者：
Saraswat, Ram;Ratnayake, Ishara;Wood, Scott T.
阻碍体外骨关节炎研究转化为临床疾病修饰疗法的一个主要限制是软骨细胞在标准单层条件下快速扩散和去分化。目前在培养中保持软骨细胞圆形形态的策略要么不自然地限制粘附并将软骨细胞置于过度僵硬的机械环境中，要么在许多应用中使用是不切实际的。为了解决现有技术的局限性，我们开发了一种独特的复合薄膜细胞培养平台CellWell，以模拟关节软骨，该平台利用微图案化的半球形威尔斯孔，其尺寸精确到适合单个细胞（12-18 μ m直径），以促进生理球形软骨细胞形态，同时保持与标准细胞培养和分析技术的兼容性。CellWells由15 μ m厚的5%琼脂糖膜构建，其中包埋有电纺聚乙烯醇（PVA）纳米纤维。PVA纳米纤维的透射电子显微镜（TEM）图像显示平均直径为60.9 +/- 24 nm，与观察到的人踝关节II型胶原纤维的平均直径53.8 +/- 29 nm密切匹配。使用AFM纳米压痕，CellWells被发现在15 μ m/s压痕时具有158 +/- 0.60 kPa的压缩模量，与天然细胞周基质的已发表刚度值紧密匹配。将取自踝关节软骨的原代人关节软骨细胞接种在CellWells中，并在24小时进行评估。软骨细胞在CellWells中保持其圆形形态（平均纵横比为0.87 +/- 0.1 vs三维（3D）对照[0.86 +/- 0.11）比在标准条件下接种的那些（0.65 +/- 0.3）更有效，平均存活率> 85%。CellWell的设计在生理硬度的薄膜基底中具有开放的半球形威尔斯孔，结合了二维（2D）培养系统的实用优势和3D系统的生理优势。通过其易用性和保持软骨细胞生理形态的能力，我们预计CellWell将提高使用该培养平台进行的未来研究的临床可翻译性。
One major limitation hindering the translation of in vitro osteoarthritis research into clinical disease-modifying therapies is that chondrocytes rapidly spread and dedifferentiate under standard monolayer conditions. Current strategies to maintain rounded morphologies of chondrocytes in culture either unnaturally restrict adhesion and place chondrocytes in an excessively stiff mechanical environment or are impractical for use in many applications. To address the limitations of current techniques, we have developed a unique composite thin-film cell culture platform, the CellWell, to model articular cartilage that utilizes micropattemed hemispheroidal wells, precisely sized to fit individual cells (12-18 pm diameters), to promote physiologically spheroidal chondrocyte morphologies while maintaining compatibility with standard cell culture and analytical techniques. CellWells were constructed of 15-mu m-thick 5% agarose films embedded with electrospun poly(vinyl alcohol) (PVA) nanofibers. Transmission electron microscope (TEM) images of PVA nanofibers revealed a mean diameter of 60.9 +/- 24 nm, closely matching the observed 53.8 +/- 29 nm mean diameter of human ankle collagen II fibers. Using AFM nanoindentation, CellWells were found to have compressive moduli of 158 +/- 0.60 kPa at 15 mu m/s indentation, closely matching published stiffness values of the native pericellular matrix. Primary human articular chondrocytes taken from ankle cartilage were seeded in CellWells and assessed at 24 h. Chondrocytes maintained their rounded morphology in CellWells (mean aspect ratio of 0.87 +/- 0.1 vs three-dimensional (3D) control [0.86 +/- 0.11) more effectively than those seeded under standard conditions (0.65 +/- 0.3), with average viability of >85%. The CellWell's design, with open, hemispheroidal wells in a thin film substrate of physiological stiffness, combines the practical advantages of two-dimensional (2D) culture systems with the physiological advantages of 3D systems. Through its ease of use and ability to maintain the physiological morphology of chondrocytes, we expect that the CellWell will enhance the dinical translatability of future studies conducted using this culture platform.