Thill Michelle, Strunnikova Natalya V, Berna Marc J, Gordiyenko Nataliya, Schmid Kristin, Cousins Scott W, Thompson Darby J S, Csaky Karl G
National Eye Institute, National Institutes of Health, Bethesda, Maryland, USA.
Invest Ophthalmol Vis Sci. 2008 Jun;49(6):2696-708. doi: 10.1167/iovs.07-0955.
To evaluate the feasibility of isolating and expanding endothelial progenitor cells (EPCs), in the form of late outgrowth endothelial progenitor cells (OECs), from the peripheral blood of an aged population, particularly patients affected by different forms of AMD.
Peripheral blood mononuclear cells were collected from young control subjects (n = 18) and from elderly subjects with non-AMD/low-risk dry AMD (n = 15), high-risk dry AMD (n = 6), or neovascular AMD (nvAMD; n = 32); cultured in established conditions; and observed for appearance of OEC clusters and growth characteristics on expansion. Expression of VEGF receptor-2 (KDR) in OECs after expansion was determined by Western blot. Plasma samples of study subjects were analyzed for CRP and VEGF levels.
OEC cultures were successfully generated from a similar number of subjects in each group. After adjustment for all other variables, subjects with high-risk dry AMD had a 5.6-fold higher number of OEC clusters per 20 mL blood, and subjects with nvAMD had a 5.1-fold high number than did subjects with non-AMD/low-risk dry AMD (P < 0.05). High-risk dry AMD generated 63 times more (NS) and nvAMD 32-times more (P < 0.05) OECs on expansion of clusters than did non-AMD/low-risk dry AMD. Population doubling occurred significantly faster in cultures from nvAMD eyes compared to non-AMD/low-risk dry AMD eyes. In addition, a significant correlation between the number of OEC clusters, expanded OECs and levels of KDR was demonstrated.
An OEC population was isolated and expanded from the blood of elderly control and AMD-affected patients and demonstrated significantly higher number of initial OEC clusters and expansion potential of OECs in patients at risk for or already affected by nvAMD. OECs may be used for further phenotypic, genetic, and functional analyses in patients with nvAMD.
评估从老年人群，尤其是受不同形式年龄相关性黄斑变性（AMD）影响的患者外周血中分离并扩增晚期内皮祖细胞（OECs）形式的内皮祖细胞（EPCs）的可行性。
从年轻对照受试者（n = 18）以及患有非AMD/低风险干性AMD（n = 15）、高风险干性AMD（n = 6）或新生血管性AMD（nvAMD；n = 32）的老年受试者中采集外周血单个核细胞；在既定条件下培养；观察OEC簇的出现情况以及扩增时的生长特性。通过蛋白质印迹法测定扩增后OEC中血管内皮生长因子受体-2（KDR）的表达。对研究受试者的血浆样本进行CRP和VEGF水平分析。
每组中相似数量的受试者均成功培养出OEC。在对所有其他变量进行校正后，高风险干性AMD受试者每20 mL血液中的OEC簇数量比非AMD/低风险干性AMD受试者高5.6倍，nvAMD受试者的该数量比非AMD/低风险干性AMD受试者高5.1倍（P < 0.05）。与非AMD/低风险干性AMD相比，高风险干性AMD在簇扩增时产生的OEC数量多63倍（无统计学意义），nvAMD产生的OEC数量多32倍（P < 0.05）。与非AMD/低风险干性AMD眼相比，nvAMD眼培养物中的群体倍增明显更快。此外，OEC簇数量、扩增后的OEC数量与KDR水平之间存在显著相关性。
从老年对照和AMD患者血液中分离并扩增出了OEC群体，且在有nvAMD风险或已受其影响的患者中，初始OEC簇数量和OEC扩增潜力显著更高。OEC可用于对nvAMD患者进行进一步的表型、基因和功能分析。
