Netherlands Institute for Innovative Ocular Surgery, Rotterdam, The Netherlands.
Melles Cornea Clinic Rotterdam, Rotterdam, The Netherlands.
PLoS One. 2019 Nov 20;14(11):e0225462. doi: 10.1371/journal.pone.0225462. eCollection 2019.
Endothelial cell migration plays a crucial role in achieving corneal clearance after corneal transplantation when using smaller-sized endothelial grafts to increase the donor pool. In this study we investigated how different strategies of Quarter-Descemet Membrane Endothelial Keratoplasty (Quarter-DMEK) limbal graft edge modification influence peripheral endothelial cell migration in an in vitro culture environment. For this study, 15 Quarter-DMEK grafts, prepared from 7 corneas deemed ineligible for transplantation but with intact and viable endothelial cells, were embedded in a cooled biocompatible, thermoresponsive matrix for culture. The limbal edge of ten Quarter-DMEK grafts were modified, either by using a small diameter punch or by peripheral radial cuts. All Quarter-DMEK grafts showed substantial collective endothelial cell migration from the radial cut graft edges, as observed by light microscopy at standardized time intervals. Grafts were retrieved from the polymer matrix after the two-week culture for immunohistochemistry analyses of the newly formed cell monolayers; this demonstrated the presence of tightly packed and viable cells that showed higher migratory ability at the leading edge. Peripheral endothelial cell migration, however, was not triggered by increasing cell exposure to free space through surgical modifications of the far periphery. Our data suggest that alterations in the far peripheral area of Quarter-DMEK grafts were insufficient to triggering cell migration from the limbal graft edge. This may be due to transient-amplifying cells that reside in the far periphery and which lack cytokinetic directional cues. Understanding the migration capacity of the peripheral endothelium could unlock cells' therapeutic potential which are, at present, routinely discarded from transplantation. Encouraging peripheral cell migration may also improve clinical outcomes from Quarter-DMEK, but a more effective solution is required prior to clinical implementation of modified grafts.
内皮细胞迁移在使用较小尺寸的内皮移植物增加供体库以实现角膜移植后角膜清除中起着至关重要的作用。在这项研究中，我们研究了不同的 Quarter-Descemet 膜内皮角膜移植术（Quarter-DMEK）角膜缘移植物边缘修饰策略如何影响体外培养环境中周边内皮细胞的迁移。在这项研究中，将 7 个不适合移植但具有完整和存活的内皮细胞的角膜制备的 15 个 Quarter-DMEK 移植物嵌入冷却的生物相容性热响应基质中进行培养。十个 Quarter-DMEK 移植物的角膜缘边缘通过使用小直径冲头或通过周边放射状切割进行修饰。在标准时间间隔通过光镜观察，所有 Quarter-DMEK 移植物均显示出从放射状切割移植物边缘大量的集体内皮细胞迁移。在两周的培养后，将移植物从聚合物基质中取出，用于新形成的细胞单层的免疫组织化学分析；这表明存在紧密堆积和存活的细胞，在前沿表现出更高的迁移能力。然而，通过手术修饰远周边缘增加细胞暴露于自由空间，并没有引发周边内皮细胞的迁移。我们的数据表明，Quarter-DMEK 移植物的远周边缘区域的改变不足以触发细胞从角膜缘移植物边缘迁移。这可能是由于位于远周边缘的暂态扩增细胞缺乏细胞分裂的定向线索。了解周边内皮细胞的迁移能力可以释放细胞的治疗潜力，目前这些细胞通常被排除在移植之外。鼓励周边细胞迁移也可能改善 Quarter-DMEK 的临床结果，但在临床实施修饰移植物之前，需要找到更有效的解决方案。
