Surgical Rehabilitation of Ocular Surface : Epithelial Transplantation

我们的目标是开发治疗原发性眼表疾病的有效方法。为此，本课题主要研究控制角膜上皮细胞增殖和分化的方法。首先，我们建立了兔角膜上皮细胞冷冻和长期传代培养的高贵细胞培养体系。该培养基含很低的钙离子浓度和较高的氨基酸含量。其次，建立了永生化兔角膜上皮细胞系。为此，用重组SV40-腺病毒载体感染原代培养的兔角膜上皮细胞，并进行克隆。该细胞系具有与正常角膜上皮细胞一致的特性。虽然这些细胞目前还不能作为上皮细胞移植，但我们在体外处理角膜上皮细胞方面取得了显著的进展。第三，我们研究了几种细胞生长和/或调节因子的生物学作用。例如，主要来自泪腺的表皮生长因子促进了角膜上皮细胞的生长。在角质细胞生长因子中也发现了类似的现象，但这只在角膜基质细胞中产生。另一方面，转化生长因子-β确实抑制了角膜上皮细胞的生长。我们还研究了三种转化生长因子-β亚型在角膜中的部分分布，发现转化生长因子-β2主要是转化生长因子-β2。在体内同种异体上皮移植排斥反应的研究中，我们正在研究环孢素治疗的患者角膜上皮移植后泪液中IL-2的水平。

We aimed at developing effective treatment for primary ocular surface disorders. For this, in this project, we mainly focused on the methods controlling proliferation and differentiation of corneal epithelial cells. First, we established a noble cell culture system for freeze and long-term subculture of rabbit corneal epithelial cells. This culture media is composed of very low Ca^<2+> concentration with high aminoacid contents. Second, we established an immortalized rabbit corneal epithelial cell line. For this, primary cultured rabbit corneal epithelial cells were infected with a recombinant SV40-adenovirus vector and were cloned. This cell line shares properties consistent with normal corneal epithelial cells. Although these cells were not transferable as a epithelial transplantation at this point, our success has made a marked progress in handling corneal epithelial cells in vitro. Third, we investigated biological roles of several cell growth and/or regulatory factors. For instance, epidermal growth factor, which mainly comes from lacrimal gland, promoted corneal epithelial cell growth. Similar phenomenon was found in keratinocyte growth factor, but this was produced only in the corneal stromal cells. On the other hand, transforming growth factor -beta did inhibit corneal epithelial cell growth. We also studied spartial distribution of three TGF-beta isoforms in the cornea, and found TGF- beta 2 predominantly.In the study of epithelial allograft rejection in vivo, we are now investigating IL-2 levels in tears in patients with cyclosporin treatment after corneal epithelial transplantation.