Wang Ji, Nain Amrinder S
Department of Mechanical Engineering and †Department of Biomedical Engineering and Mechanics, Virginia Tech , Blacksburg, Virginia 24061, United States.
Langmuir. 2014 Nov 18;30(45):13641-9. doi: 10.1021/la503011u. Epub 2014 Nov 6.
Extracellular matrix (ECM) is a fibrous natural cell environment, possessing complicated micro- and nanoarchitectures, which provide extracellular signaling cues and influence cell behaviors. Mimicking this three-dimensional microenvironment in vitro is a challenge in developmental and disease biology. Here, suspended multilayer hierarchical nanofiber assemblies (diameter from micrometers to less than 100 nm) with accurately controlled fiber orientation and spacing are demonstrated as biological scaffolds fabricated using the non-electrospinning STEP (Spinneret based Tunable Engineered Parameter) fiber manufacturing technique. Micro/nanofiber arrays were manufactured with high parallelism (relative angles between fibers were maintained less than 6°) and well controlled interfiber spacing (<15%). Using these controls, we demonstrate a bottom up hierarchical assembly of suspended six layer structures of progressively reduced diameters and spacing from several polymer systems. We then demonstrate use of STEP scaffolds to study single and multicell arrangement at high magnifications. Specifically, using double layer divergent (0°-90°) suspended nanofibers assemblies, we show precise quantitative control of cell geometry (change in shape index from 0.15 to 0.57 at similar cell areas), and through design of scaffold porosity (80 × 80 μm(2) to 5 × 5 μm(2)) quadruple the cell attachment density. Furthermore, using unidirectional or crisscross patterns of sparse and dense fiber arrays, we are able to control the cell spread area from ∼400 to ∼700 μm(2), while the nucleus shape index increases from 0.75 to 0.99 with cells nearly doubling their focal adhesion cluster lengths (∼15 μm) on widely spaced nanofiber arrays. The platform developed in this study allows a wide parametric investigation of biophysical cues which influence cell behaviors with implications in tissue engineering, developmental biology, and disease biology.
细胞外基质（ECM）是一种纤维状的天然细胞环境，具有复杂的微米和纳米结构，能提供细胞外信号线索并影响细胞行为。在体外模拟这种三维微环境是发育生物学和疾病生物学面临的一项挑战。在此，展示了具有精确控制的纤维取向和间距的悬浮多层分级纳米纤维组件（直径从微米到小于100纳米），作为使用非静电纺丝STEP（基于喷丝头的可调工程参数）纤维制造技术制造的生物支架。微/纳米纤维阵列以高平行度制造（纤维之间的相对角度保持小于6°）且纤维间距控制良好（<15%）。通过这些控制，我们展示了由几种聚合物系统构成的直径和间距逐渐减小的悬浮六层结构的自下而上分级组装。然后，我们展示了使用STEP支架在高倍放大下研究单细胞和多细胞排列。具体而言，使用双层发散（0°-90°）悬浮纳米纤维组件，我们展示了对细胞几何形状的精确定量控制（在相似细胞面积下形状指数从0.15变为0.57），并且通过设计支架孔隙率（从80×80μm²到5×5μm²）使细胞附着密度增加四倍。此外，使用稀疏和密集纤维阵列的单向或交叉图案，我们能够将细胞铺展面积控制在约400至约700μm²之间，而细胞核形状指数从0.75增加到0.99，细胞在宽间距纳米纤维阵列上的粘着斑簇长度几乎翻倍（约15μm）。本研究中开发的平台允许对影响细胞行为的生物物理线索进行广泛的参数研究，这对组织工程、发育生物学和疾病生物学具有重要意义。
