He Y G, Alizadeh H, Kinoshita K, McCulley J P
Department of Ophthalmology, The University of Texas Southwestern Medical Center at Dallas, 75235-9057, USA.
Cornea. 1999 Sep;18(5):570-9.
Tissue-cultured corneal epithelial transplantation is a novel procedure that uses tissue-cultured epithelial cells to restore severely damaged ocular surfaces. In this study, we used tissue-cultured human limbal and amniotic epithelial cells as donor cells to investigate the feasibility of this procedure for reestablishment of a damaged ocular surface in experimental conditions.
Primary human limbal epithelial cultures were established from banked limbal tissue. Amniotic epithelial cells were isolated from serologically screened human placenta and maintained in a specialized nutrient medium. Suspended cells (5 x 10(5)/ml) were seeded onto the concave surface of collagen corneal shields and incubated at 37 degrees C for 2-3 days. These cell-covered shields were then placed on a denuded stromal surface in organ culture and on New Zealand albino rabbit ocular surfaces that had the native epithelium previously removed. Specimens were collected 24, 48, 72, and 96 h later from organ-cultured corneal buttons and recipient animals, processed, and evaluated histologically.
The cells grown on the collagen shield were spread uniformly and unpolarized after 48 h in culture. They were repolarized and tightly adhered to the recipient corneal stroma 24 h after transplantation, as demonstrated by formation of cell-substrate hemidesmosomes (HDs) and donor-specific antigen immunostaining. The donor cells were retained in six of 15 rabbits receiving limbal cells and four of 12 rabbits receiving amniotic cells for as long as 10 days after surgery.
Cultured human limbal and amniotic epithelial cells can be successfully transplanted onto a denuded corneal surface where they adhere tightly to underlying stroma by hemidesmosomes.
组织培养角膜上皮移植是一种利用组织培养上皮细胞修复严重受损眼表的新方法。在本研究中，我们使用组织培养的人角膜缘和羊膜上皮细胞作为供体细胞，在实验条件下研究该方法重建受损眼表的可行性。
从储存的角膜缘组织建立原代人角膜缘上皮培养物。从经血清学筛查的人胎盘中分离羊膜上皮细胞，并在特殊营养培养基中培养。将悬浮细胞（5×10⁵/ml）接种到胶原角膜罩的凹面上，于37℃孵育2 - 3天。然后将这些细胞覆盖的罩放置在器官培养中裸露的基质表面以及先前已去除天然上皮的新西兰白化兔眼表上。在24、48、72和96小时后从器官培养的角膜纽扣和受体动物中收集标本，进行处理并进行组织学评估。
培养48小时后，在胶原罩上生长的细胞均匀铺展且未极化。移植24小时后，它们重新极化并紧密粘附于受体角膜基质，细胞 - 基质半桥粒（HDs）的形成和供体特异性抗原免疫染色证明了这一点。在接受角膜缘细胞的15只兔子中的6只以及接受羊膜细胞的12只兔子中的4只中，供体细胞在手术后长达10天内得以保留。
培养的人角膜缘和羊膜上皮细胞可成功移植到裸露的角膜表面，在那里它们通过半桥粒紧密粘附于下方的基质。
