Department of Medical Sciences, Section of Hematology, Azienda Ospedaliero-Universitaria, Arcispedale Sant'Anna, University of Ferrara, Ferrara, Italy.
PLoS One. 2013;8(2):e56377. doi: 10.1371/journal.pone.0056377. Epub 2013 Feb 11.
The current understanding of the functional characteristics of circulating endothelial progenitor cells (EPC) is limited, especially in patients affected by cardiovascular diseases. In this study, we have analyzed the in vitro clonogenic capacity of circulating EPC, also known as endothelial colony-forming cells (ECFC), in patients with acute coronary syndrome (ACS), in comparison to the colony forming unit-endothelial-like cells (CFU-EC) of hematopoietic/monocytic origin.
METHODOLOGY/PRINCIPAL FINDINGS: By culturing peripheral blood mononuclear cells (PBMC) of patients with ACS (n = 70), CFU-EC were frequently isolated (from 77% of ACS patients), while EPC/ECFC were obtained only in a small subset (13%) of PBMC samples, all harvested between 7-14 days after the acute cardiovascular event. Notably, ex-vivo generation of EPC/ECFC was correlated to a higher in vitro release of PDGF-AA by the corresponding ACS patient PBMC. By using specific endothelial culture media, EPC/ECFC displayed in vitro expansion capacity, allowing the phenotypic and functional characterization of the cells. Indeed, after expansion, EPC/ECFC exhibited a normal diploid chromosomal setting by FISH analysis and an immunophenotype characterized by: i) uniform positivity for the expression of CD105, CD31, CD146 and Factor VIII, i) variable expression of the CD34, CD106 and CD184 markers, and iii) negativity for CD45, CD90, CD117 and CD133. Of interest, in single-cell replanting assays EPC/ECFC exhibited clonogenic expansion capacity, forming secondary colonies characterized by variable proliferation capacities.
CONCLUSION/SIGNIFICANCE: Our data indicate that a careful characterization of true EPC is needed in order to design future studies in the clinical autologous setting of patients with ACS.
目前对循环内皮祖细胞（EPC）功能特征的了解有限，尤其是在患有心血管疾病的患者中。在这项研究中，我们分析了急性冠状动脉综合征（ACS）患者循环 EPC（也称为内皮集落形成细胞 [ECFC]）的体外克隆形成能力，并与造血/单核来源的集落形成单位-内皮样细胞（CFU-EC）进行了比较。
方法/主要发现：通过培养 ACS 患者的外周血单核细胞（PBMC）（n = 70），我们经常分离 CFU-EC（来自 77%的 ACS 患者），而 EPC/ECFC 仅在一小部分（13%）的 PBMC 样本中获得，所有样本均在急性心血管事件发生后 7-14 天采集。值得注意的是，EPC/ECFC 的体外生成与相应 ACS 患者 PBMC 中 PDGF-AA 的体外释放量呈正相关。通过使用特定的内皮细胞培养基，EPC/ECFC 显示出体外扩增能力，允许对细胞进行表型和功能特征分析。事实上，经过扩增，EPC/ECFC 通过 FISH 分析显示出正常的二倍体染色体设置，并具有以下免疫表型特征：i）CD105、CD31、CD146 和因子 VIII 的表达均匀阳性，ii）CD34、CD106 和 CD184 标志物的表达可变，以及 iii）CD45、CD90、CD117 和 CD133 阴性。有趣的是，在单细胞再种植实验中，EPC/ECFC 表现出克隆扩增能力，形成具有可变增殖能力的次级集落。
结论/意义：我们的数据表明，需要对真正的 EPC 进行仔细的特征描述，以便在 ACS 患者的临床自体环境中设计未来的研究。
