Engelmann K, Böhnke M, Friedl P
Department of Cytogenetics, GBF-Gesellschaft für Biotechnologische Forschung mbH, Braunschweig, Federal Republic of Germany.
Invest Ophthalmol Vis Sci. 1988 Nov;29(11):1656-62.
Human corneal endothelial cells (HCEC) were isolated by means of enzymatic treatment of excised corneas. The corneas were incubated for 1.5 hr together with a high concentration of collagenase (0.5%), followed by a long-term incubation (up to 16 hr) using a low concentration of the enzyme (0.04%). Endothelial cells were enriched against contaminating fibroblasts by using a selective L-valine-free medium which inhibited fibroblast growth during the first passages. Subcultures of HCEC were passaged for more than 20 generations without showing signs of senescence. Laminin and chondroitin sulfate functioned as a substrate for HCEC, promoting proliferation and allowing the cells to grow in monolayer formation. The inclusion of fibroblast growth factor (FGF) as well as chondroitin sulfate in the medium led to an additional increase in the rate of proliferation.
人角膜内皮细胞（HCEC）通过对切除的角膜进行酶处理来分离。将角膜与高浓度胶原酶（0.5%）一起孵育1.5小时，然后使用低浓度酶（0.04%）进行长期孵育（长达16小时）。通过使用选择性无L - 缬氨酸培养基来富集内皮细胞以对抗污染的成纤维细胞，该培养基在最初几代培养过程中抑制成纤维细胞生长。HCEC的传代培养超过20代而未显示衰老迹象。层粘连蛋白和硫酸软骨素作为HCEC的底物，促进增殖并使细胞以单层形式生长。培养基中加入成纤维细胞生长因子（FGF）以及硫酸软骨素导致增殖速率进一步增加。
