Prechondrogenic ATDC5 Cell Attachment and Differentiation on Graphene Foam; Modulation by Surface Functionalization with Fibronectin
Prechondrogenic ATDC5 Cell Attachment and Differentiation on Graphene Foam; Modulation by Surface Functionalization with Fibronectin
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DOI：
10.1021/acsami.9b14670
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发表时间：
2019-11-13
影响因子：
9.5
通讯作者：
Oxford, Julia Thom
中科院分区：
文献类型：
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作者：
Frahs, Stephanie M.;Reeck, Jonathon C.;Oxford, Julia Thom
石墨烯泡沫为组织工程应用带来了希望。在这项研究中，石墨烯泡沫被用作三维支架来评估细胞附着、细胞形态和早期分化的分子标记。本研究的目的是确定在建立三维细胞培养物之前，是否可以通过用纤连蛋白（一种细胞粘附良好的细胞外基质蛋白）对石墨烯泡沫进行功能化来调节细胞附着和细胞外基质的形成。分子动力学模拟表明，纤连蛋白-石墨烯相互作用主要通过石墨烯和蛋白质精氨酸侧链之间的相互作用来稳定。准静态和动态机械测试表明，石墨烯的纤连蛋白功能化改变了石墨烯泡沫的机械性能。由于纤连蛋白，支架的弹性强度增加，但粘弹性机械行为保持不变。当石墨烯泡沫涂有纤连蛋白并与细胞一起培养 28 天时，观察到机械刚度的附加效应。通过荧光显微镜评估的细胞骨架组织表明肌动蛋白细胞骨架的纤连蛋白依赖性重组和肌动蛋白应力纤维的增加。通过定量实时聚合酶链反应评估编码细胞附着蛋白的 9 个基因（Cd44、Ctnna1、Ctnnb1、Itga3、Itga5、Itgav、Itgb1、Ncam1、Sgce）、编码细胞外基质蛋白的 16 个基因（Col1a1、Col2a1、Col3a1、Col5a1、Col6a1、Ecm1、Emilin1、Fn1、 Hapin1、Lamb3、Postn、Sparc、Spp1、Thbs1、Thbs2、Tnc）和 9 个编码重塑调节剂的基因（Adamts1、Adamts2、Ctgf、Mmp14、Mmp2、Tgfbi、Timp1、Timp2、Timp3）表明石墨烯泡沫提供了有利于早期软骨形成中重要基因表达的微环境。石墨烯泡沫与纤连蛋白的功能化改变了细胞对石墨烯泡沫的反应，这通过相对基因表达水平的降低得到证明。这些发现说明了在设计用于组织工程应用的三维石墨烯支架时需要考虑的微米级材料特性和纳米级分子特征的组合因素。
Graphene foam holds promise for tissue engineering applications. In this study, graphene foam was used as a three -dimension scaffold to evaluate cell attachment, cell morphology, and molecular markers of early differentiation. The aim of this study was to determine if cell attachment and elaboration of an extracellular matrix would be modulated by functionalization of graphene foam with fibronectin, an extracellular matrix protein that cells adhere well to, prior to the establishment of three-dimensional cell culture. The molecular dynamic simulation demonstrated that the fibronectin-graphene interaction was stabilized predominantly through interaction between the graphene and arginine side chains of the protein. Quasi-static and dynamic mechanical testing indicated that fibronectin functionalization of graphene altered the mechanical properties of graphene foam. The elastic strength of the scaffold increased due to fibronectin, but the viscoelastic mechanical behavior remained unchanged. An additive effect was observed in the mechanical stiffness when the graphene foam was both coated with fibronectin and cultured with cells for 28 days. Cytoskeletal organization assessed by fluorescence microscopy demonstrated a fibronectin-dependent reorganization of the actin cytoskeleton and an increase in actin stress fibers. Gene expression assessed by quantitative real-time polymerase chain reaction of 9 genes encoding cell attachment proteins (Cd44, Ctnna1, Ctnnb1, Itga3, Itga5, Itgav, Itgb1, Ncam1, Sgce), 16 genes encoding extracellular matrix proteins (Col1a1, Col2a1, Col3a1, Col5a1, Col6a1, Ecm1, Emilin1, Fn1, Hapin1, Lamb3, Postn, Sparc, Spp1, Thbs1, Thbs2, Tnc), and 9 genes encoding modulators of remodeling (Adamts1, Adamts2, Ctgf, Mmp14, Mmp2, Tgfbi, Timp1, Timp2, Timp3) indicated that graphene foam provided a microenvironment conducive to expression of genes that are important in early chondrogenesis. Functionalization of graphene foam with fibronectin modified the cellular response to graphene foam, demonstrated by decreases in relative gene expression levels. These findings illustrate the combinatorial factors of microscale materials properties and nanoscale molecular features to consider in the design of three-dimensional graphene scaffolds for tissue engineering applications.