压力仿生培养对兔角膜内皮细胞调控的研究

发布时间：2018-01-22 11:36

本文关键词： 压力微环境仿生培养角膜内皮细胞　出处：《眼科》2017年01期 　论文类型：期刊论文

【摘要】：目的探讨体外压力仿生培养系统下不同梯度压力对角膜内皮细胞形态和功能的调控作用。设计实验性研究。研究对象兔角膜内皮细胞。方法将体外培养的第一代兔角膜内皮细胞分为五组:A组为无压力常规培养(空白对照),B组为正常压力仿生培养(15 mmHg),C组为压力波动组,将压力设为15mmHg、25 mmHg、20 mmHg、10 mmHg,D组30 mmHg压力培养,E组50 mmHg压力培养。细胞均培养24h,免疫法鉴定原代角膜内皮细胞,HE染色和电镜观察细胞形态的改变,流式细胞术检测细胞活性。主要指标角膜内皮细胞的形态、存活率。结果获取的所有细胞证实为角膜内皮细胞表型,无角膜上皮细胞及基质细胞污染。五组细胞分别培养24h后,经HE染色和电镜检测发现正常压力微环境培养的角膜内皮细胞排列紧密,六边形细胞居多,细胞表面微绒毛丰富,细胞核染色质丰富,而高压力培养的角膜内皮细胞活性差,细胞间隙加大。经流式细胞术分析显示,正常压力组、30 mmHg组、压力波动组、50 mmHg组的角膜内皮细胞培养24 h后细胞存活率分别为(98.16±0.45)%、(78.83±1.65)%、(70.2±3.54)%、(41.33±0.25)%(P=0.016)。高压力培养组中随着压力的升高和持续时间延长,细胞活性显著下降。结论正常压力微环境培养对角膜内皮细胞形态和功能具有正向调节作用,而高压力对角膜内皮细胞具有损伤性,并随时间延长而加重。
[Abstract]:Objective to investigate the effects of different gradient pressure on the morphology and function of corneal endothelial cells in vitro under the pressure bionic culture system. To design an experimental study on rabbit corneal endothelial cells. Methods the first generation of cultured rabbit corneal endothelial cells in vitro was used to investigate the effects of different gradient pressures on the morphology and function of corneal endothelial cells. Rabbit corneal endothelial cells were divided into five groups:. Group A was cultured without stress (. Blank control). Group B was a normal pressure bionic culture group with 15 mm HgG and C group was a pressure fluctuation group. The pressure was set at 15mm HgG 25 mmHg + 20 mmHg for 10 mmHg. Group D was cultured under pressure for 30 mmHg and group E was cultured under pressure for 50 mmHg. All the cells were cultured for 24 hours. The primary corneal endothelial cells were identified by HE staining and the morphological changes were observed by electron microscope. Main outcome measures: morphology and survival rate of corneal endothelial cells. Results all the obtained cells were confirmed as corneal endothelial cell phenotypes. No corneal epithelial cells and stromal cells were contaminated. After cultured for 24 hours in the five groups, HE staining and electron microscopy showed that the corneal endothelial cells cultured under normal pressure microenvironment were closely arranged, and hexagonal cells were the majority. The microvilli on the surface of the cells were abundant, the nuclear chromatin was abundant, but the activity of corneal endothelial cells in high pressure culture was poor, and the intercellular gap was enlarged. Flow cytometry analysis showed that the normal pressure group was 30 mmHg group. The survival rate of corneal endothelial cells cultured for 24 hours in the pressure fluctuating group was 98.16 卤0.45 and 78.83 卤1.65% respectively. 70.2 卤3.54 and 41.33 卤0.25 respectively. In the high pressure culture group, the pressure increased and the duration prolonged with the increase of pressure. Conclusion normal pressure microenvironment culture can positively regulate the morphology and function of corneal endothelial cells, while high pressure can damage corneal endothelial cells and increase with time.
【作者单位】：南昌大学;江西新视界眼科医院;中山眼科中心;南昌爱尔眼科;南昌大学附属眼科医院;
【基金】：国家自然科学基金(30801263)
【分类号】：R772.2
【正文快照】： 近年干细胞领域的研究证实,组织的微环境不同,导致组织干细胞的发育方向也不同,这提示细胞的微环境对其生理功能具有重要的调控作用[1]。Koizumi和Yuriko等[2-3]运用气-液相技术,模拟活体角膜上皮细胞处于空气和泪液交互作用形成的气-液面环境,培养的角膜上皮细胞连接更为紧

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