Department of Bioengineering, University of Pennsylvania, 210 S, 33rd Street, Philadelphia, PA 19104, USA.
Integr Biol (Camb). 2012 Oct;4(10):1164-74. doi: 10.1039/c2ib20059g.
Encapsulations of cells in type-I collagen matrices are widely used three-dimensional (3D) in vitro models of wound healing and tissue morphogenesis and are common constructs for drug delivery and for in vivo implantation. As cells remodel the exogenous collagen scaffold, they also assemble a dense fibronectin (Fn) matrix that aids in tissue compaction; however, the spatio-temporal (re)organization of Fn and collagen in this setting has yet to be quantitatively investigated. Here, we utilized microfabricated tissue gauges (μTUGs) to guide the contraction of microscale encapsulations of fibroblasts within collagen gels. We combined this system with a Foerster Radius Energy Transfer (FRET) labeled biosensor of Fn conformation to probe the organization, conformation and remodeling of both the exogenous collagen and the cell-assembled Fn matrices. We show that within hours, compact Fn from culture media adsorbed to the collagen scaffold. Over the course of tissue remodeling, this Fn-coated collagen scaffold was compacted into a thin, sparsely populated core around which cells assembled a dense fibrillar Fn shell that was rich in both cell and plasma derived Fn. This resulted in two separate Fn populations with different conformations (compact/adsorbed and extended/fibrillar) in microtissues. Cell contractility and microtissue geometry cooperated to remodel these two populations, resulting in spatial gradients in Fn conformation. Together, these results highlight an important spatio-temporal interplay between two prominent extracellular matrix (ECM) molecules (Fn and collagen) and cellular traction forces, and will have implications for future studies of the force-mediated remodeling events that occur within collagen scaffolds either in 3D in vitro models or within surgical implants in vivo.
细胞在 I 型胶原基质中的包封广泛用于伤口愈合和组织形态发生的三维（3D）体外模型，也是药物输送和体内植入的常见构建体。随着细胞重塑外源性胶原支架，它们还组装了密集的纤维连接蛋白（Fn）基质，有助于组织压实；然而，在这种情况下，Fn 和胶原的时空（再）组织尚未被定量研究。在这里，我们利用微制造组织量规（μTUG）引导成纤维细胞在胶原凝胶中的微封装收缩。我们将该系统与纤维连接蛋白构象的 Foerster 半径能量转移（FRET）标记生物传感器结合使用，以探测外源性胶原和细胞组装的 Fn 基质的组织、构象和重塑。我们表明，在几小时内，来自培养基的紧凑 Fn 吸附到胶原支架上。在组织重塑过程中，这种涂有 Fn 的胶原支架被压缩成一个薄而稀疏的核心，细胞围绕着这个核心组装了一个密集的纤维状 Fn 壳，其中富含细胞和血浆衍生的 Fn。这导致微组织中存在两种具有不同构象（紧凑/吸附和伸展/纤维状）的不同 Fn 群体。细胞收缩性和微组织几何形状共同重塑了这两种群体，导致 Fn 构象的空间梯度。总之，这些结果强调了两种突出的细胞外基质（ECM）分子（Fn 和胶原）与细胞牵引力之间的重要时空相互作用，并将对未来研究在三维体外模型中或在体内手术植入物内发生的胶原支架内的力介导重塑事件产生影响。
