Transwell小室共培养条件下缺氧时视网膜色素上皮细胞对内皮细胞增殖的影响

Effects of retinal pigment epithelium cells on proliferation of endothelial cells under hypoxia in a co-culture system

目的研究在Transwell小室共培养系统中,缺氧时视网膜色素上皮细胞(retinal pigment epithelium,RPE)对血管内皮细胞增殖的影响。方法培养并鉴定RPE细胞和人脐静脉血管内皮细胞,利用200μmol/LCoCl2造成细胞缺氧、Transwell小室建立细胞共培养模型。实验分为4组:对照组:内皮细胞;共培养组:内皮细胞+RPE细胞;缺氧对照组:内皮细胞+CoCl2;缺氧共培养组:内皮细胞+RPE细胞+CoCl2,各组分别培养0 h2、4 h4、8 h后,应用MTT法检测内皮细胞增殖情况。结果常氧时,对照组内皮细胞光吸收值(A值)在24 h达最高;对照组和共培养组内皮细胞光吸收值在各时间点差别均无统计学意义(P>0.05)。缺氧条件下,缺氧对照组A值在24 h4、8 h均低于对照组(P<0.05);缺氧对照组A值在24 h、48 h低于缺氧共培养组(P<0.05)。结论缺氧早期内皮细胞增殖受到抑制;缺氧条件下,RPE促进内皮细胞的增殖。

Objective Using Transwell system to investigate the role of retinal pigment epithelium （RPE） cells in growth of endothelial cells in hypoxia. Methods Endothelial cells and RPE cells were cultured in a contact co-culture system by Transwell chamber, hypoxic conditions were achieved by adding 200 μmoL/L CoCl2 to medium to mimic hypoxia. Endothelial cells proliferation was evaluated by MTT. Experimental endothelial cells were divided into four groups, including Endothelial cells cultured under normoxia, endothelial cells and RPE cells co-cultured under normoxia, endothelial cells cultured under hypoxia, and endothelial cells and RPE cells co-cultured uniter hypoxia. M3T assay was carried out to determine the number of cells at the 0, 24th and 48th hour in each well. Results Under normoxia, the A value of endothelial cells was the highest at the 24th hour. There was no significant difference between the group of the endothelial and the group of the endothelial and the RPE co-culture （ P 〉 0. 05 ） ; Under hypoxia, the A value of the group of the en-dothelial cell with CoCl2 was lower than that in the normal group at the time points of 24 and 48 hours （P 〈 0.05 ）. The A value of the endothelial cells cultured under hy.poxia was lower than the endothelial ceils and RPE ceils co-cultured under hypoxia at the time points of 24 and 48 hours （P 〈 0. 05 ）. Conclusion The proliferation of endothelial cells could be inhibited at the early stage of hypoxia. Under hypoxia, the RPE cells could protect the change of endothelial＇cells.