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The influence of preparation and storage time on endothelial cells in Quarter-Descemet membrane endothelial keratoplasty (Quarter-DMEK) grafts in vitro.
Cell and Tissue Banking ( IF 1.5 ) Pub Date : 2020-08-14 , DOI: 10.1007/s10561-020-09854-z
Daniele Spinozzi 1 , Alina Miron 1 , Jessica Lie 1, 2 , Silke Oellerich 1 , Sorcha Ni Dhubhghaill 1, 3 , Gerrit R J Melles 1, 2, 3
Affiliation  

Quarter–Descemet membrane endothelial keratoplasty (Quarter-DMEK) has been introduced as a modification of the standard DMEK technique to increase the pool of endothelial grafts. In this study, we evaluated in vitro changes in endothelial cell distribution, viability and morphology of Quarter–DMEK grafts when stored in organ-culture medium. Quarter–DMEK grafts were prepared from 5 corneas and stored in organ-culture medium for 4, 7 and 11 days. Endothelial cell re-distribution was investigated by light microscopy, cell viability by a Calcein-AM assay, and expression of endothelial and non-endothelial markers by immunohistochemistry. Three standard DMEK-grafts were used as controls. After preparation, all Quarter–DMEK grafts showed a band with no viable endothelial cells along the radial cut graft edges [average width 190 (± 20) µm]. Endothelial cell density in the central graft area decreased by 12%, 23% and 26% after 4, 7, and 11 days of storage, respectively. At the same time, empty bands along the cut edges were re-populated and some cells migrated to the stromal side of the Descemet membrane (DM). These cells showed an altered phenotype, as indicated by expression of migration marker CD73 and fibroblast marker αSMA. Majority of migration occurred within the first 4 days of storage. Our data suggest that endothelial cells on Quarter–DMEK grafts re-distribute during organ-culture storage to re-populate preparation-induced empty bands and after re-distribution, cells may show further migration to the stromal DM side during storage.



中文翻译:

制备和储存时间对体外 Quarter-Descemet 膜内皮角膜移植术 (Quarter-DMEK) 移植物中内皮细胞的影响。

Quarter-Descemet 膜内皮角膜移植术 (Quarter-DMEK) 已被引入作为标准 DMEK 技术的改进,以增加内皮移植物的池。在这项研究中,我们评估了 Quarter-DMEK 移植物储存在器官培养基中时内皮细胞分布、活力和形态的体外变化。从 5 个角膜制备四分之一-DMEK 移植物,并在器官培养基中储存 4、7 和 11 天。通过光学显微镜研究内皮细胞的重新分布,通过钙黄绿素-AM 测定法研究细胞活力,并通过免疫组织化学法研究内皮和非内皮标记物的表达。三个标准 DMEK 移植物用作对照。制备后,所有 Quarter-DMEK 移植物在径向切割移植物边缘均显示一条带,没有活的内皮细胞 [平均宽度 190 (± 20) µm]。在储存 4、7 和 11 天后,中央移植区的内皮细胞密度分别降低了 12%、23% 和 26%。同时,沿着切割边缘的空带被重新填充,一些细胞迁移到后旋膜 (DM) 的基质侧。这些细胞显示出改变的表型,如迁移标记 CD73 和成纤维细胞标记 αSMA 的表达所示。大多数迁移发生在存储的前 4 天内。我们的数据表明 Quarter-DMEK 移植物上的内皮细胞在器官培养储存过程中重新分布以重新填充制备诱导的空条带,并且在重新分布后,细胞可能在储存过程中进一步迁移到基质 DM 侧。沿着切割边缘的空带被重新填充,一些细胞迁移到 Descemet 膜 (DM) 的基质侧。这些细胞显示出改变的表型,如迁移标记 CD73 和成纤维细胞标记 αSMA 的表达所示。大多数迁移发生在存储的前 4 天内。我们的数据表明 Quarter-DMEK 移植物上的内皮细胞在器官培养储存过程中重新分布以重新填充制备诱导的空条带,并且在重新分布后,细胞可能在储存过程中进一步迁移到基质 DM 侧。沿着切割边缘的空带被重新填充,一些细胞迁移到 Descemet 膜 (DM) 的基质侧。这些细胞显示出改变的表型,如迁移标记 CD73 和成纤维细胞标记 αSMA 的表达所示。大多数迁移发生在存储的前 4 天内。我们的数据表明 Quarter-DMEK 移植物上的内皮细胞在器官培养储存过程中重新分布以重新填充制备诱导的空条带,并且在重新分布后,细胞可能在储存过程中进一步迁移到基质 DM 侧。大多数迁移发生在存储的前 4 天内。我们的数据表明 Quarter-DMEK 移植物上的内皮细胞在器官培养储存过程中重新分布以重新填充制备诱导的空条带,并且在重新分布后,细胞可能在储存过程中进一步迁移到基质 DM 侧。大多数迁移发生在存储的前 4 天内。我们的数据表明 Quarter-DMEK 移植物上的内皮细胞在器官培养储存过程中重新分布以重新填充制备诱导的空条带,并且在重新分布后,细胞可能在储存过程中进一步迁移到基质 DM 侧。

更新日期:2020-08-14
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