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Effect of three different media on serum free culture of donor corneas and isolated human corneal endothelial cells

机译：三种不同培养基对供体角膜和离体人角膜内皮细胞无血清培养的影响

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BACKGROUND—Removal of bovine serum from organ culture medium is necessary because of the variability in serum composition and the potential risk of infection. Two specific endothelial cell media (F99 and Endothelial-SFM) were compared with the routinely used medium MEM for their use in serum free cultivation of human corneal endothelial cells (HCEC) and donor corneas.
METHODS—HCEC were incubated in three test media with or without increasing serum content and a growth assay was performed. Seven pairs of donor corneas were cultured in each of three media for 3 weeks, one cornea with serum supplementation and one without. Endothelial cell density was determined once each week. Trypan blue staining of the endothelium and vital staining of keratocytes was performed after 3 weeks.
RESULTS—All three media promoted proliferation of cultured HCEC when supplemented with serum. Endothelial cell density of donor corneas was comparable after 3 weeks of cultivation in the different media. Only corneas cultured in medium MEM without serum exhibited a higher endothelial cell loss. Trypan blue staining of the endothelium after cultivation revealed the lowest number of damaged cells on corneas cultured in the medium Endothelial-SFM. The highest densities of keratocytes were found in corneas cultured in Endothelial-SFM and the lowest densities occurred after culture in MEM.
CONCLUSION—After incubation in Endothelial-SFM even under serum free conditions corneas were found to be of higher quality with respect to endothelial cell survival, cell membrane integrity, and keratocyte density. This medium may replace MEM, which is routinely used in European eye banks but requires supplementation with serum.


机译：背景技术由于血清成分的可变性和潜在的感染风险，必须从器官培养基中去除牛血清。将两种特定的内皮细胞培养基（F99和内皮-SFM）与常规使用的培养基MEM进行比较，比较它们在无血清培养人角膜内皮细胞（HCEC）和供体角膜中的用途。方法— HCEC在有或没有增加血清含量的三种测试培养基中孵育，并进行生长测定。在三种培养基中的每一种中分别培养7对供体角膜3周，其中一种补充血清，另一种不补充血清。每周一次测定内皮细胞密度。 3周后进行内皮锥虫蓝染色和角膜细胞活力染色。结果—当补充血清时，所有三种培养基均促进培养的HCEC的增殖。在不同培养基中培养3周后，供体角膜的内皮细胞密度相当。仅在无血清的MEM培养基中培养的角膜表现出较高的内皮细胞损失。培养后内皮的锥虫蓝染色显示在内皮-SFM培养基中培养的角膜上受损细胞的数量最少。在内皮-SFM中培养的角膜中发现最高的角膜细胞密度，而在MEM中培养后的最低密度发生在角膜中。结论—在内皮-SFM中孵育后，即使在无血清条件下，在内皮细胞存活率，细胞膜完整性和角膜细胞密度方面，角膜的质量也更高。这种培养基可以代替MEM，后者通常在欧洲眼库中使用，但需要补充血清。

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