Flow cytometric quantitation of red blood cell vesicles in thalassemia
Flow cytometric quantitation of red blood cell vesicles in thalassemia
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DOI：
10.1002/cyto.b.10064
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发表时间：
2004-01-01
影响因子：
3.4
通讯作者：
Webster, HK
中科院分区：
文献类型：
--
作者：
Pattanapanyasat, K;Noulsri, E;Webster, HK
背景：地中海贫血是一种由珠蛋白基因复合体突变引起的遗传性溶血性贫血。在这些患者中也发现了循环障碍，包括动脉和静脉血栓形成。异常红细胞的聚集性和来自激活的血小板和其他血细胞的高水平膜衍生微粒被认为是相关的血栓风险的原因。红细胞的破坏也被认为是一个重要的病理生理后果，特别是通过形成循环小泡。据我们所知，还没有尝试对地中海贫血患者的红细胞囊泡数量进行定量评估。这促使我们使用定量流式细胞术来研究地中海贫血患者外周血中RBC囊泡的水平。方法：对每个受试者的全血进行RBC和血小板或Annexin V标记的双重染色，以及已知密度的TruCount(TM)珠子。根据RBC的前、侧向散射和RBC标志物对RBC囊泡进行门控。用流式细胞术分析红细胞囊泡的百分率及其绝对数。结果：我们的数据显示，红细胞囊泡呈膜联蛋白V阳性。膜联蛋白V阳性事件的数量高于其完整的红细胞。在正常和地中海贫血的血液样本中都存在红细胞囊泡，但地中海贫血患者的红细胞囊泡数量显著增加。结论：流式细胞术定量检测红细胞囊泡方法简便、可靠，为探讨地中海贫血患者血红蛋白合成缺陷、红细胞紊乱与病理生理并发症之间的关系提供了新的思路。(C)2003年Wiley-Liss，Inc.
Background: Thalassemia is a hereditary hemolytic anemia caused by mutations in the globin gene complex. Circulatory disturbances including arterial and venous thrombosis have also been noted in these patients. Aggregability of abnormal RBC and the high level of membrane-derived microparticles stemming from activated platelets and other blood cells are thought to be responsible for the associated thrombotic risk. Destruction of RBC is also thought to be an important pathophysiological consequence, particularly through the formation of circulating vesicles. To our knowledge, there has been no attempt to quantitatively evaluate the number of RBC vesicles in thalassemia. This prompted us to study the level of RBC vesicles in the peripheral blood of thalassemia patients using quantitative flow cytometry.Methods: Whole blood from each subject was doubly stained for RBC and platelet or annexin V markers, together with the known density TruCount(TM) beads. RBC vesicles were gated according to their forward/side scatter and RBC marker. Percentage of RBC vesicles and their absolute number were analyzed by flow cytometry.Results: Our data indicated that RBC vesicles were annexin V-positive. The number of annexin V-positive events was higher than their intact RBCs. RBC vesicles were present in both normal and thalassemic blood samples, but the numbers of RBC vesicles were significantly higher in thalassemia. Both the percentage and the absolute number of RBC vesicles were especially marked in splenectomized subjects with beta-thalassemia/Hemoglobin E. When clinical and hematological indices were compared with RBC vesicles, there was an inverse relationship between the degree of severity in thalassemia patients and the number of RBC vesicles.Conclusion: Flow cytometric quantitation of RBC vesicles is simple, reliable and may offer new insights in to study of the relationship between defective hemoglobin synthesis, RBC perturbation and pathophysiological complications in thalassemia. (C) 2003 Wiley-Liss, Inc.