Evaluation of potential organ culture media for eye banking using a human corneal endothelial cell growth assay
Evaluation of potential organ culture media for eye banking using a human corneal endothelial cell growth assay
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DOI：
10.1007/s004170100354
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发表时间：
2001-10-01
影响因子：
2.7
通讯作者：
Engelmann, K
中科院分区：
文献类型：
--
作者：
Moller-Pedersen, T;Hartmann, U;Engelmann, K
背景资料：评价不同市售细胞培养基诱导人角膜内皮细胞（HCEC）原代培养物增殖和形态学变化的能力。使用该筛选模型试图为开发用于人类供体角膜长期器官培养的定义明确的无血清保存培养基建立合理的基础。方法：对添加0%、2%、5%和10%胎牛血清（FCS）的11种不同培养基进行比较。试验培养基分为三组：第1组：基于最小必需培养基（MEM）的培养基，目前用于欧洲眼库的长期角膜器官培养;第2组：基于F99的培养基，在血清减少条件下富集角膜内皮细胞生长;第3组：设计用于特殊细胞类型生长或短期角膜器官培养的培养基。使用HCEC增殖试验定量每种试验培养基的促生长能力，而通过相差显微镜评价细胞形态的变化。结果如下：HCEC的形态特征在基于F99的培养基组中保持得最好，其在血清减少的条件下也诱导最高水平的细胞增殖。具体地，培养基F99-Sr（F99富含抗坏血酸、胰岛素、bFGF、转铁蛋白。硒和脂质）在0%和2%FCS下诱导的HCEC密度比所有其它测试培养基高2 - 3倍，并且它还保持了最内皮细胞样的形态。此外，在较高的血清浓度（5%和10%FCS）下，细胞生长在F99-Sr以及最初设计用于血管内皮细胞无血清生长的培养基SFM中最为显著。结论：这项研究表明，培养基F99-Sr和SFM应进一步测试和完善，作为潜在的新的存储解决方案，在生理温度下的长期角膜器官培养。
Background: To evaluate the ability of different commercially available cell Culture media to induce proliferation and morphological changes in primary cultures of human corneal endothelial cells (HCEC). This screening model was used in an attempt to establish a rational basis for the development of well-defined, serum-free preservation media for long-term organ culture of human donor corneas. Methods: A total of I I different culture media enriched with 0%, 2%, 5%, and 10% fetal calf serum (FCS) were compared. The test media were divided into three groups: Group 1: Media based on minimal essential medium (MEM), currently used for long-term corneal organ culture in European eye banks; Group 2: F99-based media, enriched for growth of corneal endothelial cells at serum-reduced conditions; and Group 3: Media designed for growth of special cell types or for short-term corneal organ culture. The growth-promoting capacity of each test medium was quantified using an HCEC proliferation assay, whereas changes in cell morphology were evaluated by phase-contrast microscopy. Results: The morphological characteristics of HCEC were best maintained in the group of F99-based media, which also induced the highest level of cell proliferation under serum-reduced conditions. Specifically, the medium F99-Sr (F99 enriched with ascorbic acid, insulin, bFGF, transferrin. selenium, and lipids) induced a two- to three-fold higher HCEC density at both 0% and 2% FCS when compared to all other test media, and it also maintained the most endothelial cell-like morphology. Also, at higher serum concentrations (5% and 10% FCS), the cell growth was most prominent in F99-Sr, as well as in the medium SFM that originally was designed for serum-free growth of vascular endothelial cells. Conclusion: This study suggests that the media F99-Sr and SFM should be further tested and refined as potential new storage solutions for long-term corneal organ culture at physiological temperatures.