Kinoshita Shigeru, Koizumi Noriko, Nakamura Takahiro
Department of Ophthalmology, Kyoto Prefectural University of Medicine, Kyoto, Japan.
Exp Eye Res. 2004 Mar;78(3):483-91. doi: 10.1016/j.exer.2003.09.004.
Ocular surface reconstruction by tissue engineering using somatic stem cells is a second-generation therapeutic modality. In view of future treatment of bilaterally affected, severe ocular surface disorders, two types of transplantable cultivated mucosal epithelial sheets can be used for reconstruction. One is an allogeneic corneal epithelial stem cell sheet, and the other is an autologous oral mucosal epithelial cell sheet. We first investigated the feasibility of amniotic membrane as an epithelial carrier, and found that denuded amniotic membrane was the most appropriate substrate for this purpose. Thus, cultivated corneal epithelial stem cell sheets were created by co-culturing with 3T3 fibroblast and air-lifting on amniotic membrane. These epithelial sheets demonstrated positive keratin 3 and 12 specific to in vivo corneal epithelium, light junction related proteins and proliferative activity. The transplanted allogeneic human corneal epithelial sheets existed successfully on the corneal surface, and were quite effective in achieving ocular surface stability in severe ocular surface disorders. A few cases, however, developed immunological reactions or opportunistic infections, etc. Secondly, we established transplantable autologous cultivated oral mucosal epithelial sheets in rabbits. The in vitro oral mucosal epithelial sheets showed histological characteristics similar to those of in vivo corneal epithelial sheets; for example, positive keratin 3 expression. Based on the fact that, the transplanted autologous oral mucosal epithelial sheets resembled corneal epithelium and that we achieved the recovery of corneal transparency in rabbits, we propose that cultivated oral mucosal epithelium may become the substitute for corneal epithelium in ocular surface reconstruction.
利用体干细胞通过组织工程进行眼表重建是一种第二代治疗方式。鉴于未来对双侧受累的严重眼表疾病的治疗，两种可移植的培养黏膜上皮片可用于重建。一种是同种异体角膜上皮干细胞片，另一种是自体口腔黏膜上皮细胞片。我们首先研究了羊膜作为上皮载体的可行性，发现去上皮羊膜是最适合此目的的基质。因此，通过与3T3成纤维细胞共培养并在羊膜上进行气提培养，制备了培养的角膜上皮干细胞片。这些上皮片显示出体内角膜上皮特有的角蛋白3和12呈阳性、与紧密连接相关的蛋白以及增殖活性。移植的同种异体人角膜上皮片成功地存在于角膜表面，并且在严重眼表疾病中实现眼表稳定方面非常有效。然而，少数病例出现了免疫反应或机会性感染等情况。其次，我们在兔体内建立了可移植的自体培养口腔黏膜上皮片。体外口腔黏膜上皮片显示出与体内角膜上皮片相似的组织学特征；例如，角蛋白3表达呈阳性。基于移植的自体口腔黏膜上皮片类似于角膜上皮且我们在兔体内实现了角膜透明度恢复这一事实，我们提出培养的口腔黏膜上皮可能成为眼表重建中角膜上皮的替代物。
