Abstract
We thank Dr Hubel for his comments. The methodology to identify circulating endothelial progenitor cells (EPCs) used in our study has been described in detail.1Savvidou M.D. Xiao Q. Kaihura C. Anderson J.M. Nicolaides K.H. Maternal circulating endothelial progenitor cells in normal singleton and twin pregnancy.Am J Obstet Gynecol. 2008; 198 (414.e1-5)Abstract Full Text Full Text PDF PubMed Scopus (19) Google Scholar In many previous studies, EPCs are described as a peripheral blood mononuclear cell (PBMC) population expressing CD34, kinase insert domain receptor (KDR)/vascular endothelial growth factor receptor-2, and CD133/AC133 with adherent growth characteristics. Whereas the function and the clonogenic capacity of EPC should be evaluated using colony-forming unit assays, their number is usually assessed by flow cytometry using either antibodies against CD34 and KDR or CD133. However, very strict and rigorous technology in flow cytometry analysis is needed, and a completed negative/positive control should be included in this analysis. This is because it is very difficult to reproduce the data of EPC numbers because of the limitation of nonspecific background of this measurement and because of the fact that just a very small fraction (0.001% to 0.03% of PBMCs or less than 5 per mL of blood) of CD34-AC133-KDR-triple positive EPCs exist in the circulation. Alternatively, phenotypes of human EPCs can be confirmed by the uptake of 1, 1'-dioctadecyl-3, 3' 3'-tetramethylindocarbocyanine–labeled acetyl low-density lipoprotein (DiI-Ac-LDL) and binding of Ulex europeus agglutinin (lectin). Our previous study2Xiao Q. Kiechl S. Patel S. et al.Endothelial progenitor cells, cardiovascular risk factors, cytokine levels and atherosclerosis—results from a large population-based study.PLoS ONE. 2007; 2: e975Crossref PubMed Scopus (134) Google Scholar demonstrated that a large proportion (from 25% to 75%) of DiI/lectin double-positive cells were endothelial cell–specific markers positive, under our culture conditions, such as CD31, CD144, von Willebrand factor, KDR, and endothelial nitric oxide synthase, but conducting all the experiments to check the endothelial-specific marker expression in DiI/lectin double-positive cells was not feasible. Therefore, double-positive cells for DiI-Ac-LDL and lectin were counted as circulating EPCs and used in the present study. We acknowledge that all termed EPCs have recently been reclassified as 3 kinds of cells: colony-forming unit–endothelial cells, circulating angiogenic cells, and endothelial colony-forming cells.3Prater D.N. Case J. Ingram D.A. Yoder M.C. Working hypothesis to redefine endothelial progenitor cells.Leukemia. 2007; 21: 1141-1149Crossref PubMed Scopus (257) Google Scholar Although we recognize the controversy in the EPC definition, based on the previously cited information, we can conclude that our measurements reflect the changes of termed EPCs during the different stages of pregnancy. Undoubtedly, a consensus among scientists would be desirable to help further the research in this field. Endothelial progenitor cells and pregnancyAmerican Journal of Obstetrics & GynecologyVol. 200Issue 1PreviewSavvidou et al1 reported a decline in endothelial progenitor cells (EPCs) in the maternal circulation with advancing gestation. The available literature on EPCs during pregnancy is conflicting, however, as appropriately acknowledged by the authors. Full-Text PDF
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