Conditional reprogrammed human limbal epithelial cells represent a novel in vitro cell model for drug responses.
Conditional reprogrammed human limbal epithelial cells represent a novel in vitro cell model for drug responses.
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DOI：
10.1016/j.bbrc.2018.03.168
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发表时间：
2018-05
影响因子：
3.1
通讯作者：
Ling Wang;Lin Ye;Gaobin Wei;Yu Chen;Lina Ye;Xiaoting Wu;Zhihong Zeng;Y. Wang;Guogan Yin;Xiang Long;Hui Li
中科院分区：
文献类型：
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作者：
Ling Wang;Lin Ye;Gaobin Wei;Yu Chen;Lina Ye;Xiaoting Wu;Zhihong Zeng;Y. Wang;Guogan Yin;Xiang Long;Hui Li
在本研究中，我们利用条件重编程(CR)技术从正常角膜缘组织中建立了人角膜缘上皮细胞(CR-LEC细胞)。我们已经成功地建立了三个人供体的CR-LEC细胞株(3个)，以及正常兔(2个)和猪(1个)的细胞株。在Matrigel 3D培养中，CR-LEC细胞保持持续稳定的增殖状态，核型正常，对DNA损伤的反应正常，结构清晰的球体。细胞对干扰素α2b、更昔洛韦和5-氟尿嘧啶的反应不同，提示这些药物对这些细胞的毒性与预期的不同。更重要的是，早期和晚期传代的CR-LEC细胞对药物的反应没有显著差异(p>；0.05)，表明CR-LEC细胞可以服务于稳定的正常人体细胞模型进行毒性评估。通过F-肌动蛋白和DSG-1的表达来检测单层培养的CR-LEC细胞的毒性。三种药物在LD_(50)浓度下的毒性作用使单层破坏逐渐增加，这与三种药物对人角膜上皮的刺激程度一致。因此，CR-LEC细胞为评价角膜毒性提供了一种新颖、可靠的体外生理学细胞模型。
In this study, we established human limbal epithelial cells (LECs) from normal limbal tissues by using Conditional Reprogramming (CR) technology (refer to CR-LEC cells in this study). We have successfully established CR-LEC cell strains from three human donors (3 out of 3), and normal rabbits (2 out of 2) and pig (1 out of 1) as well. CR-LEC cells sustained a continuous and stable proliferation status with a normal karyotype, normal response to DNA damage, well-defined structured spheres in matrigel 3D culture. Responses of CR-LEC cells to IFN α2b, Ganciclovir and 5-Fluorouracil were different, suggesting that these drugs had different toxicities to these cells as expected. More important, there was no significant difference of responses to drugs between early and late passages of CR-LEC cells (p>0.05), indicating CR-LEC cells can serve a stable normal human cell model for toxicity assessment. Toxicity tests with monolayer cultures of CR-LEC cells were measured by staining the F-actin and Dsg-1 expression. Toxicity of three drugs at LD50 concentration resulted in a gradually increased destruction of monolayer, which is, in accordance with the irritation grade of three drugs on human cornea epithelium. Therefore, CR-LEC cells provide a novel and reliable in vitro physiological cell model for corneal toxicity assessment.