Skottman H, Muranen J, Lähdekorpi H, Pajula E, Mäkelä K, Koivusalo L, Koistinen A, Uusitalo H, Kaarniranta K, Juuti-Uusitalo K
Faculty of Medicine and Life Sciences, BioMediTech, University of Tampere, Tampere, Finland.
Department of Obstetrics and Gynecology, Tampere University Hospital, Finland.
Exp Cell Res. 2017 Oct 1;359(1):101-111. doi: 10.1016/j.yexcr.2017.08.004. Epub 2017 Aug 8.
Here we evaluated the effects of human retinal microvascular endothelial cells (hREC) on mature human embryonic stem cell (hESC) derived retinal pigment epithelial (RPE) cells. The hESC-RPE cells (Regea08/017, Regea08/023 or Regea11/013) and hREC (ACBRI 181) were co-cultured on opposite sides of transparent membranes for up to six weeks. Thereafter barrier function, small molecule permeability, localization of RPE and endothelial cell marker proteins, cellular fine structure, and growth factor secretion of were evaluated. After co-culture, the RPE specific CRALBP and endothelial cell specific von Willebrand factor were appropriately localized. In addition, the general morphology, pigmentation, and fine structure of hESC-RPE cells were unaffected. Co-culture increased the barrier function of hESC-RPE cells, detected both with TEER measurements and cumulative permeability of FD4 - although the differences varied among the cell lines. Co-culturing significantly altered VEGF and PEDF secretion, but again the differences were cell line specific. The results of this study showed that co-culture with hREC affects hESC-RPE functionality. In addition, co-culture revealed drastic cell line specific differences, most notably in growth factor secretion. This model has the potential to be used as an in vitro outer blood-retinal barrier model for drug permeability testing.
在此，我们评估了人视网膜微血管内皮细胞（hREC）对成熟的人胚胎干细胞（hESC）来源的视网膜色素上皮（RPE）细胞的影响。将hESC-RPE细胞（Regea08/017、Regea08/023或Regea11/013）和hREC（ACBRI 181）在透明膜的相对两侧共培养长达六周。此后，评估了屏障功能、小分子通透性、RPE和内皮细胞标记蛋白的定位、细胞精细结构以及生长因子分泌。共培养后，RPE特异性的CRALBP和内皮细胞特异性的血管性血友病因子定位适当。此外，hESC-RPE细胞的总体形态、色素沉着和精细结构未受影响。共培养增加了hESC-RPE细胞的屏障功能，通过跨上皮电阻测量和FD4的累积通透性均能检测到——尽管不同细胞系之间存在差异。共培养显著改变了VEGF和PEDF的分泌，但同样，这些差异具有细胞系特异性。本研究结果表明，与hREC共培养会影响hESC-RPE的功能。此外，共培养揭示了显著的细胞系特异性差异，最明显的是在生长因子分泌方面。该模型有潜力用作药物通透性测试的体外血视网膜外屏障模型。
