Engelmann Katrin, Bednarz Jürgen, Valtink Monika
University Eye Hospital, University Hospital Carl Gustav Carus at the TU Dresden, Fetscherstrasse 74, Dresden 01307, Germany.
Exp Eye Res. 2004 Mar;78(3):573-8. doi: 10.1016/s0014-4835(03)00209-4.
The human corneal endothelium has a limited proliferative capacity in vivo. Until now it has only been possible to replace damaged endothelium by transplantation of a donor cornea. After establishing methods for the isolation and in vitro cultivation of human corneal endothelial cells (HCEC), transplantation of these cells may be an alternative therapeutic option.
In this review methods for the in vitro cultivation of HCEC and their transplantation onto the Descemet membrane of donor corneas are described.
In vitro proliferation of human adult corneal endothelial cells was achieved by the development of defined cell culture conditions, including supplementation of culture medium with specified growth factors. Dependent on the culture conditions, in vitro cultured endothelial cells showed phenotypic changes and different proliferative behaviour. The propagation of corneal endothelial cells in vitro offered the possibility of their transplantation onto donor corneas in an in vitro model. After transplantation, these cells formed a monolayer whose morphology and cell density depended on the differentiation status of the cells in vitro. Highest cell numbers up to 3000 cells/mm2 were achieved using a SV40-transformed HCEC-cell line. Monolayer integrity could be demonstrated by positive staining for integrins and light junction proteins, and pump function of the newly established endothelium was proven by perfusion studies.
Methods to transplant HCEC onto human denuded corneas have been successfully established to reconstruct human corneas. Recent developments in genetic manipulation of cells and tissue engineering will be of great help in constructing suitable corneas for keratoplasty. Thus corneal endothelial cell transplantation is one of the promising future possibilities to provide corneas of high quality for patients. Furthermore, improvement of the transplantation technique may lead to a method to directly manipulate the diseased endothelium of patients with corneal endothelial dystrophies.
人角膜内皮细胞在体内的增殖能力有限。到目前为止，只能通过移植供体角膜来替换受损的内皮细胞。在建立了人角膜内皮细胞（HCEC）的分离和体外培养方法后，移植这些细胞可能是一种替代治疗选择。
本综述描述了HCEC的体外培养方法及其移植到供体角膜后弹力层的方法。
通过开发特定的细胞培养条件，包括在培养基中添加特定生长因子，实现了人成年角膜内皮细胞的体外增殖。根据培养条件，体外培养的内皮细胞表现出表型变化和不同的增殖行为。角膜内皮细胞在体外的增殖为在体外模型中将其移植到供体角膜上提供了可能性。移植后，这些细胞形成单层，其形态和细胞密度取决于体外细胞的分化状态。使用SV40转化的HCEC细胞系可实现高达3000个细胞/mm2的最高细胞数量。通过整合素和紧密连接蛋白的阳性染色可证明单层的完整性，通过灌注研究证实了新建立的内皮细胞的泵功能。
已成功建立将HCEC移植到人类裸露角膜上以重建人类角膜的方法。细胞基因操作和组织工程的最新进展将极大地有助于构建适合角膜移植的角膜。因此，角膜内皮细胞移植是未来为患者提供高质量角膜的有前景的可能性之一。此外，移植技术的改进可能会导致一种直接处理角膜内皮营养不良患者患病内皮的方法。
