机械牵拉诱导人RPE细胞IL-8分泌及其对PI3K/AKT通路的影响

IL-8 Secretion Induced by Mechanical Stretch in Human RPE Cells and Its Effect on PI3K/AKT Pathway

目的:视网膜脱离发生机制尚不明确,本文旨在研究机械应力牵拉后视网膜色素上皮细胞上清中IL-8分泌量的变化以及人其对RPE细胞PI3K/AKT通路变化影响。方法:培养人RPE细胞,应用Flex-cell细胞应力加载系统牵拉人RPE细胞不同时间(0h、1 h、3 h、6 h、9 h)建立不同牵拉时间的模型。分别标记为对照组、牵拉1 h组、牵拉3 h组、牵拉6 h组、牵拉9 h组。随后用ELISA方法检测人RPE细胞不同实验组上清中IL-8分泌量的变化。细胞免疫荧光和Western blot方法观察PI3K、PPI3K、T-AKT、P-AKT的表达。结果:随着牵拉时间的延长,ELISA方法检测人RPE细胞不同实验组上清中IL-8分泌量逐渐增加[924.79±5.92 pg/m L、947.73±5.34 pg/m L、974.53±5.74 pg/m L、979.57±1.12 pg/m L、1019.22±4.25 pg/m L],差异有统计学意义(F=166,p<0.01),各牵拉组与对照组分别对比,差异有统计学意义(P<0.05)。细胞中P-AKT蛋白表达增加[0.61±0.02、0.97±0.05、0.99±0.04、1.21±0.11、1.20±0.07],差异有统计学意义(F=41.24,p<0.01),各牵拉组与对照组分别对比,差异有统计学意义(P<0.05),AKT表达未见明显变化。结论:机械应力牵拉人RPE细胞时,细胞上清中IL-8分泌量逐渐增加同时RPE细胞内PI3K/AKT通路明显激活,并与牵拉时间有关,为探究EMT原因和对防治PVR提供了理论基础。

Objective： The mechanism of retinal detachment is not clear, this paper aims to study volume changes of supematant IL-8 and the changes of of PI3K / Akt pathway in human RPE cells which were stretched by the Device of Mechanical Stress. Methods： Human RPE cells were stretched by the Device of Mechanical Stress for 0 h,1 h,3 h,6 h,9 h. Labeled as the control group, 1 h group, 3 h group, 6 h group, 9 h group. The changes of 1L-8 secreted by human RPE cells in different experimental groups were detected by ELISA. Expression levels of PPI3K, T-AKT, PI3K P-AKT were observed by the immunofluorescence and Western blot method. Results： With the extension of stretch time, the volume of supematant IL-8 in different groups increased gradually [924.79± 5.92 pg/mL, 947.73± 5.34 pg/mL, 974.53± 5.74 pg/mL, 979.57± 1.12 pg/mL, 1019.22± 4.25pg/mL]. The results reviewed that the differences between different groups were significant （F=166, P〈0.01）. The differences between stretch groups and control group were significant （P 〈 0.05）. The protein expression of PAKT increased [0.61＋ 0.02, 0.97± 0.05, 0.99± 0.04, 1.21± 0.11, 1.20± 0.07]. The differences between different groups were significant （F=41.24, P 〈 0.01）. The differences between stretch groups and control group were significant （P 〈 0.05）. Conclusion： With the extension of stretch time, the volumes of supematant IL-8 in different groups increased gradually and the PI3K / Akt pathway was activated in human RPE cells which was stretched by the Device of Mechanical Stress. The results offer a method to explore the reasons of EMT and provide a basis for the prevention and treatment of PVR.