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Abstract：
Contradictory behavior of microvascular retinal endothelial cells (REC) - a reliable in vitro model to study retinal diseases - have recently been reported which might result from cultivating the cells in standard DMEM not optimized for this cell type. Therefore, we studied DMEM\'s effects on phenotype and behavior of immortalized bovine REC. Cells were cultivated in endothelial cell growth medium (ECGM) until a confluent monolayer was reached and then further kept for 1-4 days in ECGM, DMEM, or mixes thereof all supplemented with 5% fetal bovine serum, endothelial cell growth supplement, 90 μg/ml heparin, and 100 nM hydrocortisone. Within hours of cultivation in DMEM, the cell index - measured to assess the cell layer\'s barrier function - dropped to ~5% of the initial value and only slowly recovered, not only accompanied by stronger expression of HSP70 mRNA and secretion of interleukin-6, but also by lower expressions of tight junction proteins claudin-5, claudin-1 or of the marker of cell type conversion caveolin-1. Altered subcellular localizations of EC-typic claudin-5, vascular endothelial cadherin and von Willebrand factor were also observed. Taken together, all experiments with (retinal) EC cultivated in common DMEM need to be interpreted very cautiously and should at least include phenotypic validation.
摘要：
微血管视网膜内皮细胞（REC）的矛盾行为-研究视网膜疾病的可靠体外模型-最近有报道，这可能是由于在未针对这种细胞类型进行优化的标准DMEM中培养细胞所致。因此,我们研究了DMEM对永生化牛REC表型和行为的影响。将细胞培养在内皮细胞生长培养基（ECGM）中，直到达到汇合的单层，然后在ECGM中进一步保持1-4天，DMEM,或其混合物均补充有5%胎牛血清，内皮细胞生长补充剂，90μg/ml肝素，和100nM氢化可的松。在DMEM中培养数小时内，细胞指数-测量以评估细胞层的屏障功能-下降到初始值的〜5％，并且仅缓慢恢复，不仅伴随着HSP70mRNA的更强表达和白细胞介素6的分泌，而且还伴随着紧密连接蛋白claudin-5，claudin-1或细胞类型转换标记物caveolin-1的表达降低。还观察到EC型claudin-5，血管内皮钙粘蛋白和vonWillebrand因子的亚细胞定位改变。一起来看,在普通DMEM中培养的(视网膜)EC的所有实验都需要非常谨慎地解释,并且至少应包括表型验证.
