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角蛋白14在发育中的人类角膜上皮祖细胞中的表达

Keratin 14 Expression in Epithelial Progenitor Cells of the Developing Human Cornea.

作者信息

Eghtedari Yas, Richardson Alexander, Mai Kelly, Heng Benjamin, Guillemin Gilles J, Wakefield Denis, Di Girolamo Nick

机构信息

1 Ocular Diseases Research Group, School of Medical Sciences, University of New South Wales , Sydney, Australia .

2 Neuroinflammation Group, Faculty of Medicine & Health Sciences, Macquarie University , Sydney, Australia .

出版信息

Stem Cells Dev. 2016 May 1;25(9):699-711. doi: 10.1089/scd.2016.0039. Epub 2016 Apr 12.

Abstract

A healthy and transparent cornea is essential for exquisite vision. During adulthood, its epithelium is constantly replenished through the activity of its stem cells (SCs). Precisely when these cells develop and their distribution across the ocular surface remain incompletely characterized in man. We postulated that the human fetal cornea harbors SCs that can be identified with keratin (K) 14 and αv-integrin, two markers we and others previously used to identify their adult counterparts. Immunofluorescence, cell culture, quantitative real-time polymerase chain reaction (qRT-PCR), and colony-forming assays were performed on fetal and adult biomaterial to locate progenitors and establish their phenotypic and functional properties. K14 was used to map the spatiotemporal distribution of precursor cell activity across the developing cornea, divulging a dynamic pattern of vertical and horizontal consolidated expression with increasing gestational age. K14 was coexpressed with αv-integrin in fetal and adult corneas and cultured corneolimbal epithelium, and colony-forming efficiency (an indicator of SC activity) was similar in cells from both sources. Finally, fetal cells were adherent, grew well, and maintained a K14 phenotype on contact lenses, a substrate we previously used to deliver cells to patients with blinding corneal disease. This study provides valuable insights into the development of the cornea, including the formation of the SC repository, the distribution of these cells across the ocular surface, and a preliminary attempt at harnessing, phenotyping, and functionally characterizing these cells. Future studies will focus on isolating fetal SCs to determine their utility as an alternative cell therapy for patients suffering from corneal blindness.

摘要

健康且透明的角膜对于敏锐的视力至关重要。在成年期,其上皮细胞通过干细胞(SCs)的活动不断得到补充。在人类中,这些细胞何时发育以及它们在眼表的分布仍未完全明确。我们推测人类胎儿角膜中存在可通过角蛋白(K)14和αv整合素识别的干细胞,这两种标志物是我们和其他人先前用于识别其成年对应物的。对胎儿和成人生物材料进行了免疫荧光、细胞培养、定量实时聚合酶链反应(qRT-PCR)和集落形成测定,以定位祖细胞并确定其表型和功能特性。K14用于绘制发育中角膜前体细胞活动的时空分布,揭示了随着胎龄增加垂直和水平整合表达的动态模式。K14在胎儿和成人角膜以及培养的角膜缘上皮中与αv整合素共表达,并且两种来源细胞的集落形成效率(干细胞活性指标)相似。最后,胎儿细胞具有粘附性,生长良好,并在隐形眼镜(我们先前用于将细胞递送给患有致盲性角膜疾病患者的基质)上维持K14表型。本研究为角膜的发育提供了有价值的见解,包括干细胞库的形成、这些细胞在眼表的分布,以及对这些细胞进行利用、表型分析和功能表征的初步尝试。未来的研究将集中于分离胎儿干细胞,以确定其作为角膜盲患者替代细胞疗法的效用。

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