van der Putten Cas, van den Broek Daniëlle, Kurniawan Nicholas A
Department of Biomedical Engineering, Eindhoven University of Technology, Eindhoven, Netherlands.
Institute for Complex Molecular Systems, Eindhoven University of Technology, Eindhoven, Netherlands.
Front Cell Dev Biol. 2022 Jul 22;10:930373. doi: 10.3389/fcell.2022.930373. eCollection 2022.
Functional tissue repair after injury or disease is governed by the regenerative or fibrotic response by cells within the tissue. In the case of corneal damage, keratocytes are a key cell type that determine the outcome of the remodeling response by either adapting to a fibroblast or myofibroblast phenotype. Although a growing body of literature indicates that geometrical cues in the environment can influence Myo(fibroblast) phenotype, there is a lack of knowledge on whether and how differentiated keratocyte phenotype is affected by the curved tissue geometry in the cornea. To address this gap, in this study we characterized the phenotype of fibroblastic and transforming growth factor β (TGFβ)-induced myofibroblastic keratocytes and studied their migration behavior on curved culture substrates with varying curvatures. Immunofluorescence staining and quantification of cell morphological parameters showed that, generally, fibroblastic keratocytes were more likely to elongate, whereas myofibroblastic keratocytes expressed more pronounced α smooth muscle actin (α-SMA) and actin stress fibers as well as more mature focal adhesions. Interestingly, keratocyte adhesion on convex structures was weak and unstable, whereas they adhered normally on flat and concave structures. On concave cylinders, fibroblastic keratocytes migrated faster and with higher persistence along the longitudinal direction compared to myofibroblastic keratocytes. Moreover, this behavior became more pronounced on smaller cylinders (i.e., higher curvatures). Taken together, both keratocyte phenotypes can sense and respond to the sign and magnitude of substrate curvatures, however, myofibroblastic keratocytes exhibit weaker curvature sensing and slower migration on curved substrates compared to fibroblastic keratocytes. These findings provide fundamental insights into keratocyte phenotype after injury, but also exemplify the potential of tuning the physical cell environments in tissue engineering settings to steer towards a favorable regeneration response.
损伤或疾病后的功能性组织修复由组织内细胞的再生或纤维化反应所控制。在角膜损伤的情况下，角膜细胞是一种关键的细胞类型，它通过适应成纤维细胞或肌成纤维细胞表型来决定重塑反应的结果。尽管越来越多的文献表明环境中的几何线索可以影响肌（成纤维）细胞表型，但对于角膜中弯曲的组织几何形状是否以及如何影响分化的角膜细胞表型，人们还缺乏了解。为了填补这一空白，在本研究中，我们对成纤维细胞性和转化生长因子β（TGFβ）诱导的肌成纤维细胞性角膜细胞的表型进行了表征，并研究了它们在具有不同曲率的弯曲培养底物上的迁移行为。免疫荧光染色和细胞形态参数的定量分析表明，一般来说，成纤维细胞性角膜细胞更倾向于伸长，而肌成纤维细胞性角膜细胞则表达更明显的α平滑肌肌动蛋白（α-SMA）和肌动蛋白应力纤维以及更成熟的粘着斑。有趣的是，角膜细胞在凸形结构上的粘附较弱且不稳定，而它们在平坦和凹形结构上能正常粘附。在凹形圆柱体上，与肌成纤维细胞性角膜细胞相比，成纤维细胞性角膜细胞沿纵向迁移得更快且持续性更高。此外，这种行为在较小的圆柱体（即更高的曲率）上变得更加明显。综上所述，两种角膜细胞表型都能感知并响应底物曲率的正负和大小，然而，与成纤维细胞性角膜细胞相比，肌成纤维细胞性角膜细胞在弯曲底物上表现出较弱的曲率感知和较慢的迁移。这些发现为损伤后角膜细胞表型提供了基本见解，也例证了在组织工程环境中调整物理细胞环境以引导产生有利的再生反应的潜力。
