Evidence suggesting the existence of stem cells for the human corneal endothelium.
Evidence suggesting the existence of stem cells for the human corneal endothelium.
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发表时间：
2005-09
期刊：
影响因子：
2.2
通讯作者：
D. Whikehart;C. Parikh;Alexia V Vaughn;K. Mishler;H. Edelhauser
中科院分区：
文献类型：
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作者：
D. Whikehart;C. Parikh;Alexia V Vaughn;K. Mishler;H. Edelhauser
众所周知，人角膜内皮细胞（HCECs）不愿分裂，这一直引起研究者的兴趣。与此相关的是，在人角膜外周中内皮细胞群增加的发现促使了对内皮外周中存在干细胞样细胞的证据的研究。表明干细胞或瞬时扩增细胞可能存在于外周中，这可能解释HCEC的起源，并表明这些细胞在伤口修复中的来源。此外，这些细胞可能在培养或作为人工角膜合成的来源方面具有价值。方法从眼库中获取人角膜，并检测端粒酶活性和BrdU（溴脱氧核苷）掺入，以分别确定干细胞样标志物和复制活性的存在。在端粒酶活性的情况下，通过使用环钻将组织分为中心、中间和周围区域。BrdU染色（使用碱性磷酸酶结合的二抗）在内皮侧暴露于BrdU抗体的整个角膜加上巩膜缘上进行，而BrdU荧光（使用荧光素结合的二抗）通过相同的程序从这些组织的横切面获得。一些角膜受伤，以确定受伤的区域是否刺激BrdU（通过染色或荧光），然后合成转化生长因子β（TGF β）。后者通过定量ELISA测定。还测定了兔角膜的BrdU掺入，以将其细胞分裂的证据与人的细胞分裂的证据进行比较。结果将角膜分为中央、中间和周边三部分后，切取的内皮组织在周边和中间部分显示端粒酶活性阳性。在中央内皮组织或小梁网和Schwalbe线之间的利姆布斯中未观察到活性。BrdU染色与碱性磷酸酶偶尔观察到受伤地区的人角膜内皮细胞受伤后。当用荧光素对角膜横切面进行BrdU荧光测定时，荧光发生在小梁网及其附近的区域，但在角膜内皮未见荧光。伤后BrdU荧光延伸至角膜内皮。TGF-β水平在创伤后浸泡内皮的液体中增加，但增加滞后于创伤事件。结论：角膜内皮细胞与小梁网交界处可能存在干细胞样细胞。这些假定的干细胞可能为角膜内皮和小梁提供新的细胞。有证据表明，来自该区域的细胞迁移（可能作为瞬时扩增细胞）到内皮外周，并且可能在需要时迁移到角膜内皮的受伤区域。迁移可能不是恒定的，可能是年龄依赖性的。
PURPOSE The well-known reluctance of human corneal endothelial cells (HCECs) to divide has continually intrigued investigators. Related to this, the discovery of an increased endothelial cell population in the periphery of the human cornea has prompted an investigation for evidence of the existence of stem-like cells in the endothelial periphery. Showing that stem cells or transient amplifying cells may exist in the periphery might explain the origin of HCECs and indicate a source for these cells in wound repair. In addition, these cells might be of value in culturing or as a source for the synthesis of artificial corneas. METHODS Human corneas with attached scleral rims were obtained from eye banks and were assayed for telomerase activity and BrdU (bromodeoxyridine) incorporation to determine, respectively, the presence of a stem-like cell marker and replicative activity. In the case of telomerase activity, the tissues were divided into central, intermediate and peripheral areas by the use of trephines. BrdU staining (using alkaline phosphatase bound secondary antibody) was performed on whole corneas plus scleral rims exposed to BrdU antibodies on the endothelial side whereas BrdU fluorescence (using fluorescein bound secondary antibody) was obtained from transverse sections of the these tissues by the same procedure. Some corneas were wounded to determine whether the wounded areas stimulated BrdU (by staining or fluorescence) followed by the synthesis of transforming growth factor beta (TGFbeta). The latter was determined by quantitative ELISA. Rabbit corneas were also assayed for BrdU incorporation to compare their evidence of cell division with that of humans. RESULTS After dividing corneas into central, intermediate, and peripheral sections, the dissected endothelial tissues exhibited positive telomerase activity in the peripheral and intermediate sections. No activity was observed in the central endothelial tissues or the limbus between the trabecular meshwork and Schwalbe's line. BrdU staining with alkaline phosphatase was occasionally observed in the wounded area's human corneal endothelial cells after wounding. When BrdU fluorescence assays were made on corneal transverse sections with fluorescein, fluorescence occurred in an area just at and adjacent to the trabecular meshwork, but was not seen at the corneal endothelium. After wounding, BrdU fluorescence extended into the corneal endothelium. TGF-beta levels were increased in fluids bathing the endothelium following wounding, but the increases lagged behind the wounding event. CONCLUSIONS It is suggested stem-like cells may be sequestered in a niche at the junctional region where the corneal endothelial cells and the trabecular meshwork come together. These putative stem cells may supply new cells for both the corneal endothelium and the trabeculae. Evidence suggests that cells from this area migrate (perhaps as transient amplifying cells) to the endothelial periphery and, perhaps, to wounded areas of the corneal endothelium when needed. The migration may not be constant and may be age dependent.