Laboratory of Translational Tumorimmunolgy, Department of Medical Oncology, Erasmus University Medical Center, Daniel den Hoed Cancer Center, Rotterdam, The Netherlands.
Thromb Haemost. 2010 Aug;104(2):318-26. doi: 10.1160/TH09-08-0594. Epub 2010 Jun 29.
Circulating endothelial cells (CEC) are considered a promising marker to determine the extent of vascular damage. However, currently available and validated CEC enumeration assays are laborious, time consuming and costly, which limits their clinical utility. Here, we evaluated the feasibility of quantifying mRNA levels of the endothelium-associated markers CD31, CD144, CD146 and von Willebrand factor (vWf) in peripheral blood (PB) of healthy donors, patients, and human umbilical veins by real-time reverse transcriptase polymerase chain reaction (RT-PCR) and their use as surrogate markers for CEC. Whole blood samples and CD146+ cell-enriched fractions were assessed for mRNA and protein expression of CD31, CD144, CD146 and vWf by RT-PCR and flow cytometry, respectively. We showed the feasibility to detect endothelial mRNA isolated from HUVEC numbers as low as 10. However, no endothelial mRNA could be measure in whole blood samples, and only low levels of CD31 and CD146 mRNA were detected in suspensions of isolated CEC with numbers up to 4,450 CEC per sample. We conclude that mRNA levels of CD31, CD144, CD146 and vWf in whole blood as detected by real time RT-PCR cannot be used as biomarkers for end-stage endothelial cells such as CEC.
循环内皮细胞（CEC）被认为是确定血管损伤程度的有前途的标志物。然而，目前可用的和经过验证的 CEC 计数检测方法既繁琐又耗时耗力，成本高昂，限制了它们的临床应用。在这里，我们通过实时逆转录聚合酶链反应（RT-PCR）评估了定量健康供体、患者和人脐静脉外周血（PB）中与内皮相关的标记物 CD31、CD144、CD146 和血管性血友病因子（vWf）的 mRNA 水平的可行性，并将其作为 CEC 的替代标志物。通过 RT-PCR 和流式细胞术分别评估全血样本和 CD146+细胞富集部分中 CD31、CD144、CD146 和 vWf 的 mRNA 和蛋白表达。我们表明，从 HUVEC 数量低至 10 即可检测到内皮 mRNA 的可行性。然而，在全血样本中无法测量到内皮 mRNA，并且在分离的 CEC 悬浮液中仅检测到低水平的 CD31 和 CD146 mRNA，每个样本的 CEC 数量高达 4,450 个。我们得出结论，实时 RT-PCR 检测全血中 CD31、CD144、CD146 和 vWf 的 mRNA 水平不能用作终末期内皮细胞（如 CEC）的生物标志物。
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