摘要
目的:视网膜色素上皮细胞(RPE)功能障碍或死亡是各类视网膜退行性疾病的常见病因。最近的报道表明ROCK途径抑制剂以细胞类型依赖性方式调节细胞增殖或凋亡。在这里,我们的目标是在体外研究ROCK抑制剂Y-27632对人视网膜色素上皮细胞(RPE)的作用。 方法:分别采用CCK-8和流式细胞术分析细胞增殖和凋亡。通过免疫荧光和免疫印迹检测细胞增殖标志物。使用扫描电子显微镜评估细胞形态。使用原子力显微镜(AFM)评估活细胞的形态和生物力学特性。另外,通过western印迹和免疫荧光检测细胞骨架和上皮 - 间质转化(EMT)标记。 结果:30μMY-27632显着促进细胞增殖并降低细胞凋亡。与对照组相比,用30μMY-27632处理的人视网膜色素上皮细胞系ARPE-19细胞显示出显着降低的细胞膜粗糙度(Ra:41.04±1.63nm对24.41±0.75nm,P <0.01; Rq:51.56±2.03nm与30.81±0.95nm比较,P <0.01),弹性模量增加(16.66±0.83KPa比32.55±1.48KPa,P <0.01)。另外,Y-27632抑制ROCK活性导致细胞伸长和细胞骨架微丝和微管的重组。 结论:综合起来,我们的数据表明Y-27632可以改变生物力学特性并重组细胞骨架以促进RPE细胞存活。这些结果是Y-27632在未来视网膜退行性疾病中应用的重要一步。
关键词: 视网膜色素上皮细胞,ROCK抑制剂,Y-27632,增殖,细胞骨架,机械性能。
Current Molecular Medicine
Title:ROCK Inhibitor Y-27632 Promotes Human Retinal Pigment Epithelium Survival by Altering Cellular Biomechanical Properties
Volume: 17 Issue: 9
关键词: 视网膜色素上皮细胞,ROCK抑制剂,Y-27632,增殖,细胞骨架,机械性能。
摘要: Purpose: Dysfunction or death of retinal pigment epithelial (RPE) cells is a common pathogenesis of various types of retinal degenerative diseases. Recent reports indicated that ROCK pathway inhibitors regulate cell proliferation or apoptosis in a celltype- dependent manner. Here, we aim to investigate the effect of ROCK inhibitor Y- 27632 on the human retinal pigment epithelium (RPE) in vitro.
Methods: Cell proliferation and apoptosis were analyzed by CCK-8 and flow cytometry respectively. Cell proliferation markers were detected by immunofluorescence and western blot. Cell morphology was evaluated using scanning electron microscopy. The topography and biomechanical properties of living cells were assessed using atomic force microscope (AFM). In addition, cytoskeleton and epithelial-mesenchymal transition (EMT) markers were detected by western blot and immunofluorescence.
Results: 30µM Y-27632 significantly promoted cell proliferation and decreased apoptosis. Compared with control group, human retinal pigment epithelial cell line ARPE-19 cells treated with 30µM Y-27632 exhibited significantly decreased cytomembrane roughness (Ra: 41.04±1.63nm vs. 24.41±0.75nm, P<0.01; Rq: 51.56±2.03nm vs. 30.81±0.95nm, P<0.01) and increased elasticity modulus (16.66±0.83KPa vs. 32.55±1.48KPa, P<0.01). In addition, the inhibition of ROCK activity by Y-27632 caused cell elongation and reorganization of microfilaments and microtubules of cytoskeletons.
Conclusion: Taken together, our data demonstrated that Y-27632 could alter biomechanical properties and reorganized cytoskeletons to promote RPE cell survival. These results are an important step toward the future application of Y-27632 in retinal degenerative diseases.
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Cite this article as:
ROCK Inhibitor Y-27632 Promotes Human Retinal Pigment Epithelium Survival by Altering Cellular Biomechanical Properties, Current Molecular Medicine 2017; 17 (9) . https://dx.doi.org/10.2174/1566524018666180316150936
DOI https://dx.doi.org/10.2174/1566524018666180316150936 |
Print ISSN 1566-5240 |
Publisher Name Bentham Science Publisher |
Online ISSN 1875-5666 |
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