Notara Maria, Shortt Alex J, Galatowicz Grazyna, Calder Virginia, Daniels Julie T
Department of Ocular Biology and Therapeutics, UCL Institute of Ophthalmology, London, UK.
Stem Cell Res. 2010 Nov;5(3):188-200. doi: 10.1016/j.scr.2010.07.002. Epub 2010 Aug 3.
The corneal epithelium is maintained by the limbal epithelial stem cells (LESCs). In this study, an in vitro model is proposed for the investigation of cell-cell interactions involving LESC maintenance. Imaging of the limbal niche demonstrated close spatial arrangement between basal limbal epithelial cells within putative LESC niche structures and fibroblasts in the stroma. Interactions of the human limbal epithelial (HLE) cells and mitotically active human limbal fibroblasts (HLF) were studied for the first time in a serum-free in vitro model that simulated aspects of the limbal niche microenvironment. HLE cocultured in a ratio 3:1 with HLF exhibited enhanced expression of the putative stem cell markers ABCG2 and p63α and holoclones were preserved as shown by colony-forming efficiency assays, clonal analysis, and colony characterisation. Interleukin 6 (IL6) was found to be up-regulated in the 3.1SF system when compared to the HLE culture with growth-arrested fibroblasts and serum (gold standard system, GS). IL6 caused a time-dependent phosphorylation of STAT3 in HLE cells. STAT3 and IL6 inhibition in 3.1SF cultures significantly reduced HLE colony-forming efficiency, suggesting a previously undetected STAT3-mediated involvement of IL6 in the maintenance of HLE cells in a progenitor-like state.
角膜上皮由角膜缘上皮干细胞（LESCs）维持。在本研究中，提出了一种体外模型，用于研究涉及LESC维持的细胞间相互作用。角膜缘生态位成像显示，在假定的LESC生态位结构内的基底角膜缘上皮细胞与基质中的成纤维细胞之间存在紧密的空间排列。在一种模拟角膜缘生态位微环境的无血清体外模型中，首次研究了人角膜缘上皮（HLE）细胞与有丝分裂活跃的人角膜缘成纤维细胞（HLF）之间的相互作用。与HLF以3:1比例共培养的HLE表现出假定干细胞标志物ABCG2和p63α的表达增强，并且通过集落形成效率测定、克隆分析和集落表征表明全克隆得以保留。与具有生长停滞的成纤维细胞和血清的HLE培养物（金标准系统，GS）相比，发现白细胞介素6（IL6）在3.1SF系统中上调。IL6导致HLE细胞中STAT3的时间依赖性磷酸化。在3.1SF培养物中抑制STAT3和IL6显著降低了HLE集落形成效率，表明以前未检测到的STAT3介导的IL6参与将HLE细胞维持在祖细胞样状态。
