Centre for the Cellular Microenvironment, Institute of Molecular, Cell & Systems Biology, MVLS, University of Glasgow, Joseph Black Building, Glasgow, G12 8QQ, UK.
Department of Pure and Applied Chemistry, Technology and Innovation Centre, University of Strathclyde, Glasgow, G1 1RD, UK.
Sci Rep. 2022 May 17;12(1):8165. doi: 10.1038/s41598-022-12057-z.
Skeletal stem cells (SSCs, or mesenchymal stromal cells typically referred to as mesenchymal stem cells from the bone marrow) are a dynamic progenitor population that can enter quiescence, self-renew or differentiate depending on regenerative demand and cues from their niche environment. However, ex vivo, in culture, they are grown typically on hard polystyrene surfaces, and this leads to rapid loss of the SSC phenotype. While materials are being developed that can control SSC growth and differentiation, very few examples of dynamic interfaces that reflect the plastic nature of the stem cells have, to date, been developed. Achieving such interfaces is challenging because of competing needs: growing SSCs require lower cell adhesion and intracellular tension while differentiation to, for example, bone-forming osteoblasts requires increased adhesion and intracellular tension. We previously reported a dynamic interface where the cell adhesion tripeptide arginine-glycine-aspartic acid (RGD) was presented to the cells upon activation by user-added elastase that cleaved a bulky blocking group hiding RGD from the cells. This allowed for a growth phase while the blocking group was in place and the cells could only form smaller adhesions, followed by an osteoblast differentiation phase that was induced after elastase was added which triggered exposure of RGD and subsequent cell adhesion and contraction. Here, we aimed to develop an autonomous system where the surface is activated according to the need of the cell by using matrix metalloprotease (MMP) cleavable peptide sequences to remove the blocking group with the hypothesis that the SSCs would produce higher levels of MMP as the cells reached confluence. The current studies demonstrate that SSCs produce active MMP-2 that can cleave functional groups on a surface. We also demonstrate that SSCs can grow on the uncleaved surface and, with time, produce osteogenic marker proteins on the MMP-responsive surface. These studies demonstrate the concept for cell-controlled surfaces that can modulate adhesion and phenotype with significant implications for stem cell phenotype modulation.
骨骼干细胞（SSC，或通常被称为骨髓间充质基质细胞的间充质基质细胞）是一种动态祖细胞群体，它可以根据再生需求和龛位环境中的信号进入静止、自我更新或分化。然而，在体外培养时，它们通常生长在坚硬的聚苯乙烯表面上，这导致 SSC 表型迅速丧失。虽然正在开发可以控制 SSC 生长和分化的材料，但迄今为止，很少有反映干细胞可塑性的动态界面的例子被开发出来。实现这样的界面是具有挑战性的，因为存在竞争需求：生长 SSC 需要较低的细胞黏附和细胞内张力，而向例如成骨细胞的骨形成分化需要增加的黏附和细胞内张力。我们之前报道了一种动态界面，其中细胞黏附三肽精氨酸-甘氨酸-天冬氨酸（RGD）在用户添加的弹性蛋白酶激活时呈现给细胞，该弹性蛋白酶切割一个隐藏 RGD 以阻止细胞黏附的大体积阻断基团。这允许在阻断基团在位时进行生长阶段，并且细胞只能形成较小的黏附，然后在添加弹性蛋白酶后诱导成骨细胞分化阶段，这会触发 RGD 的暴露以及随后的细胞黏附和收缩。在这里，我们旨在开发一种自主系统，其中根据细胞的需要通过使用基质金属蛋白酶（MMP）可切割肽序列激活表面，假设当细胞达到汇合时，SSC 会产生更高水平的 MMP。目前的研究表明，SSC 会产生可切割表面上功能性基团的活性 MMP-2。我们还证明 SSC 可以在未切割的表面上生长，并且随着时间的推移，在 MMP 响应表面上产生成骨标记蛋白。这些研究证明了用于细胞控制表面的概念，该表面可以调节黏附和表型，对干细胞表型调节具有重要意义。
