一种快速、可靠地计数患者循环内皮细胞的新方法。

A new approach for rapid and reliable enumeration of circulating endothelial cells in patients.

机构信息

Department of Medical Oncology, Erasmus Medical Center, Daniel den Hoed Cancer Center, Rotterdam, Netherlands.

出版信息

J Thromb Haemost. 2012 May;10(5):931-9. doi: 10.1111/j.1538-7836.2012.04681.x.

DOI:10.1111/j.1538-7836.2012.04681.x

PMID:22385979

Abstract

BACKGROUND

Mature circulating endothelial cells (CECs) are surrogate markers of endothelial damage/dysfunction. A lack of standardized assays and consensus on CEC phenotype has resulted in a wide variation of reported CEC numbers (4-1300 per mL).

OBJECTIVES

Given the need for a quick, reliable, robust and validated CEC assay at an affordable price, we present a novel approach to enumerate CECs using a multi-parameter flow cytometric (FCM) method without immunological pre-enrichment.

METHODS

CECs were defined as CD34+, CD45neg, CD146+ and DNA+ events based on the immunophenotype of endothelial cells from vein-wall dissections. As CECs express high levels of CD34, we based our assay on absolute CD34 counts after analyzing all CD34 positive events in a total blood volume of 4 mL needed for a precise enumeration of CECs at a frequency of < 1 cell μL(-1).

RESULTS

The endothelial origin of CECs was confirmed by morphology, immunohistochemistry and gene expression. The new FCM assay was tested in parallel with a validated assay (i.e. CellSearch). CEC levels ranged from 4 to 79 CEC mL(-1) in healthy individuals and were significantly higher in patients with advanced solid malignancies (P = 0.0008) and in patients with hematological malignancies (P < 0.0001).

CONCLUSIONS

This flow cytometric method should be useful as a fast and economical assay to enumerate and characterize CECs.

摘要

背景

成熟的循环内皮细胞（CEC）是内皮损伤/功能障碍的替代标志物。由于缺乏标准化的检测方法和对 CEC 表型的共识，导致报告的 CEC 数量差异很大（4-1300 个/ml）。

目的

鉴于需要一种快速、可靠、稳健且经过验证的、价格合理的 CEC 检测方法，我们提出了一种使用多参数流式细胞术（FCM）方法来计数 CEC 的新方法，无需免疫预富集。

方法

CEC 被定义为 CD34+、CD45neg、CD146+和 DNA+事件，基于静脉壁切片内皮细胞的免疫表型。由于 CEC 表达高水平的 CD34，我们的检测方法基于在 4ml 全血体积中分析所有 CD34 阳性事件后的绝对 CD34 计数，以在 < 1 个细胞/μl(-1)的频率下精确计数 CEC。

结果

CEC 的内皮起源通过形态学、免疫组织化学和基因表达得到证实。新的 FCM 检测方法与经过验证的检测方法（即 CellSearch）并行测试。健康个体的 CEC 水平范围为 4-79 CEC ml(-1)，在晚期实体恶性肿瘤患者（P = 0.0008）和血液恶性肿瘤患者中显著更高（P < 0.0001）。