Eye Institute and Affiliated Xiamen Eye Center, Xiamen University, Xiamen, Fujian, China.
Stem Cells. 2009 Oct;27(10):2516-26. doi: 10.1002/stem.174.
Importin13 (IPO13), the newest member of importin-beta family discovered recently, is a unique nucleus-cytoplasm bidirectional transport receptor protein. In this study, IPO13 expression in human corneal tissue, limbal epithelial primary explant and clonal culture was evaluated by immunostaining and reverse-transcription polymerase chain reasgon. IPO13 function was evaluated in the corneal epithelial culture treated with IPO13 inhibitor, or fetal bovine serum (FBS)-containing Dulbecco's modified Eagle's medium (DMEM) medium by colony-forming efficiency, clone growth capacity, MTT, immunostaining, and Western blotting assay. IPO13 protein was expressed mainly in nuclei of limbal epithelial basal cells, but not in the other cell layers of limbus and full thickness of corneal epithelia. IPO13 was expressed in the majority of epithelial cells in early-stage clones and in the margin of late-stage clones. IPO13 was positively expressed in mouse TKE2 progenitor cells cultured in keratinocyte serum-free defined medium, while it became negative in FBS-containing DMEM, which promoted TKE2 cell differentiation. In the presence of IPO13 inhibitor, IPO13 expression and the proliferative capacity decreased in human limbal epithelial clones and mouse TKE2 cells, which were accompanied with the cell differentiation. In conclusion, our findings demonstrate for the first time that IPO13 is uniquely expressed by human limbal basal epithelial cells, and plays an important role in maintaining the phenotype, high proliferative potential, and less differentiation of corneal epithelial progenitor cells, suggesting that IPO13 could serve as a novel potential marker for corneal epithelial progenitor cells.
Importin13（IPO13），最近发现的 importin-β 家族的最新成员，是一种独特的核质双向转运受体蛋白。在本研究中，通过免疫染色和逆转录聚合酶链反应评估了 IPO13 在人角膜组织、角膜缘上皮原代外植体和克隆培养物中的表达。通过集落形成效率、克隆生长能力、MTT、免疫染色和 Western blot 分析，评估了用 IPO13 抑制剂或含胎牛血清（FBS）的 Dulbecco 改良 Eagle 培养基（DMEM）处理的角膜上皮培养物中的 IPO13 功能。IPO13 蛋白主要表达在角膜缘上皮基底层细胞的核内，但不在角膜缘的其他细胞层和全层角膜上皮内表达。IPO13 在早期克隆的大多数上皮细胞中和晚期克隆的边缘表达。在无血清角质形成细胞定义培养基中培养的小鼠 TKE2 祖细胞中，IPO13 呈阳性表达，而在含 FBS 的 DMEM 中则呈阴性表达，后者促进了 TKE2 细胞分化。在存在 IPO13 抑制剂的情况下，人角膜缘上皮克隆和小鼠 TKE2 细胞中的 IPO13 表达和增殖能力下降，同时伴随着细胞分化。总之，我们的研究结果首次表明，IPO13 由人角膜缘基底上皮细胞特异性表达，在维持角膜上皮祖细胞的表型、高增殖潜能和较少分化方面发挥重要作用，提示 IPO13 可作为角膜上皮祖细胞的一种新的潜在标志物。
