Circulating Endothelial Progenitor Cells
http://www.100md.com
《新英格兰医药杂志》
To the Editor: Recent studies have shown that endothelial progenitor cells are mobilized within a few hours after acute myocardial infarction, and the level of these cells in peripheral blood remains elevated for up to several days or weeks in patients with acute myocardial infarction, as compared with patients with stable angina and healthy control subjects.1,2 Among 507 participants in the study by Werner et al. (Sept. 8 issue)3 who had coronary artery disease on angiography, a history of acute, subacute, or myocardial infarction was present in 3.4 percent, 8.5 percent, and 32.1 percent, respectively. The level of endothelial progenitor cells in these patients was elevated to various degrees as a result of previous myocardial infarction. Therefore, categorizing patients according to the level of endothelial progenitor cells without considering the history of myocardial infarction may have caused a misclassification in this study. It is possible that patients who had recently had a myocardial infarction (and therefore were at higher risk for future infarction and death) were wrongly assigned to the group of patients with a medium level or high level of endothelial progenitor cells among those who had never had a myocardial infarction. This potential misclassification may explain the lack of the association between the level of endothelial progenitor cells and the incidence of myocardial infarction and death from all causes or acute myocardial infarction.
Dae Hyun Kim, M.D., M.P.H.
Thomas Jefferson University
Philadelphia, PA 19107
dae-hyun.kim@mail.tju.edu
References
Wojakowski W, Tendera M, Michalowska A, et al. Mobilization of CD34/CXCR4+, CD34/CD117+, c-met+ stem cells, and mononuclear cells expressing early cardiac, muscle, and endothelial markers into peripheral blood in patients with acute myocardial myocardial infarction. Circulation 2004;110:3213-3220.
Massa M, Rosti V, Ferrario M, et al. Increased circulating hematopoietic and endothelial progenitor cells in the early phase of acute myocardial infarction. Blood 2005;105:199-206.
Werner N, Kosiol S, Schiegl T, et al. Circulating endothelial progenitor cells and cardiovascular outcomes. N Engl J Med 2005;353:999-1007.
To the Editor: Werner et al. reported that increased levels of circulating endothelial progenitor cells were associated with a reduced risk of future cardiovascular events among patients with coronary artery disease, and the significant difference in the cumulative event rate according to the levels of endothelial progenitor cells could be shown early in the first four months of follow-up. However, several clinical observational studies, such as that by George et al.,1 demonstrated that patients with acute coronary syndromes had elevated levels of circulating endothelial progenitor cells. Furthermore, George et al. showed that the level of circulating endothelial progenitor cells decreased by nearly 50 percent, as did the level of C-reactive protein (CRP), in patients with unstable angina after their clinical condition had been stabilized for three months,1 suggesting that tissue ischemia has a role in triggering the mobilization of endothelial progenitor cells and may be correlated with systemic inflammation. In the article by Werner et al., half the patients had acute coronary syndromes or had undergone coronary angioplasty, and the levels of circulating endothelial progenitor cells may have been altered by these events. The CRP level is an independent predictor of adverse cardiovascular events.2 Additional information, such as serum level of CRP, might further elucidate the important role of endothelial progenitor cells in determining future events in patients with cardiovascular disease.
Hsin-Bang Leu, M.D.
Jaw-Wen Chen, M.D.
Shing-Jong Lin, M.D., Ph.D.
National Yang-Ming University
Taipei 112, Taiwan
doc1958h@yahoo.com.tw
References
George J, Goldstein E, Abashidze S, et al. Circulating endothelial progenitor cells in patients with unstable angina: association with systemic inflammation. Eur Heart J 2004;25:1003-1008.
Ridker PM, Hennekens CH, Buring JE, et al. C-reactive protein and other markers of inflammation in the prediction of cardiovascular disease in women. N Engl J Med 2000;342:836-843.
To the Editor: Werner and coauthors stated that "the level of circulating CD34+KDR+ endothelial progenitor cells predicts the occurrence of cardiovascular events and death from cardiovascular causes." They calculated the percentage of endothelial progenitor cells in the lymphocytic fraction of peripheral-blood mononuclear cells with fluorescence-activated cell-sorting analysis and as colony-forming units in 1x107 peripheral-blood mononuclear cells.
The fraction of mononuclear cells, as isolated by density gradient, is heterogeneous and contains cell populations that can vary considerably with disease or over time.1 Our concern is that measuring the percentage of endothelial progenitor cells in the lymphocytic fraction could underestimate or overestimate the concentration of endothelial progenitor cells if another cell population within that fraction is altered. To obtain an accurate concentration of blood endothelial progenitor cells, the percentage of such cells measured in the lymphocytic fraction by fluorescence-activated cell sorting or the number of colony-forming units in mononuclear cells should be multiplied by the total lymphocyte or mononuclear-cell count.2,3 To gain more insight into the mechanism and to detect a possible causative relation, it would be important to determine the prognostic value of absolute counts of endothelial progenitor cells.
Harald C. Ott, M.D.
Doris A. Taylor, Ph.D.
University of Minnesota
Minneapolis, MN 55455
ottxx057@umn.edu
References
B?yum A. Separation of white blood cells. Nature 1964;204:793-794.
Elliott C, Samson DM, Armitage S, et al. When to harvest peripheral-blood stem cells after mobilization therapy: prediction of CD34-positive cell yield by preceding day CD34-positive concentration in peripheral blood. J Clin Oncol 1996;14:970-973.
Fukuda J, Kaneko T, Egashira M, Oshimi K. Direct measurement of CD34+ blood stem cell absolute counts by flow cytometry. Stem Cells 1998;16:294-300.
To the Editor: Werner et al. show that the number of blood cells with a CD34+KDR+ phenotype predicts the occurrence of cardiovascular events and death from cardiovascular causes. They indicate that these cells are circulating endothelial progenitors. However, CD34 and KDR are antigens expressed not only by endothelial progenitors but also by endothelial cells with a mature phenotype, which make up the large majority of all circulating endothelial cells.1,2 This is a subtle but relevant issue, because it is still unclear whether in adult life most of vascular remodeling is due to angiogenesis (mature cell–derived generation of new vessels) or vasculogenesis (endothelial progenitor cell–dependent generation of new vessels).3,4 So far, only the CD133 antigen is known to be expressed by endothelial progenitors and not by mature endothelial cells.4 Thus, in the population of patients in the study by Werner et al., it would be useful to dissect clinical outcomes associated with the number of CD133+CD34+KDR+ endothelial progenitors from those associated with the number of CD34+KDR+ cells, a majority of which are mature circulating endothelial cells.
Francesco Bertolini, M.D., Ph.D.
Patrizia Mancuso, Ph.D.
European Institute of Oncology
20141 Milan, Italy
francesco.bertolini@ieo.it
Robert S. Kerbel, Ph.D.
Sunnybrook and Women's College Health Sciences Centre
Toronto, ON M4N 3M5, Canada
References
Mancuso P. Resting and activated endothelial cells are increased in the peripheral blood of cancer patients. Blood 2001;97:3658-3661.
Blann AD, Woywodt A, Bertolini F, et al. Circulating endothelial cells: biomarker of vascular disease. Thromb Haemost 2005;93:228-235.
Jain RK. Molecular regulation of vessel maturation. Nat Med 2003;9:685-693.
Rafii S, Lyden D, Benezra R, Hattori K, Heissig B. Vascular and haematopoietic stem cells: novel targets for anti-angiogenesis therapy? Nat Rev Cancer 2002;2:826-835.
To the Editor: With regard to the study by Werner et al., we would like to highlight a distinctly different endothelial cell type, the CD146+ circulating endothelial cell. These are mature cells that have detached from the endothelium as a result of endothelial injury or damage. The number of circulating endothelial cells has been shown to correlate closely with several well-recognized indexes of endothelial damage or dysfunction.1,2 We have previously reported an increased number of circulating endothelial cells in a cohort of 156 patients presenting with acute coronary syndromes.3 The number of circulating endothelial cells at 48 hours after admission was a strong and independent predictor of the risk of death and major adverse cardiac events at 30 days and at 1 year.3 There is also a clear relationship between an increased number of circulating endothelial cells and high Thrombolysis in Myocardial Infarction (TIMI) risk scores in acute coronary syndromes.4
Current data suggest that circulating endothelial cells and endothelial progenitor cells are separate cell populations. The former are mature endothelial cells that are shed from the vessel wall in response to damage, whereas the latter are involved in vascular regeneration or repair. Clearly, more information is needed for an adequate definition of the interrelationships between these two types of cells.
Christopher J. Boos, M.B., B.S., M.R.C.P.
Patrick K.Y. Goon, M.R.C.S.
Gregory Y.H. Lip, M.D.
City Hospital
Birmingham B18 7QH, United Kingdom
g.y.h.lip@bham.ac.uk
References
Schmidt-Lucke C, Rossig L, Fichtlscherer S, et al. Reduced number of circulating endothelial progenitor cells predicts future cardiovascular events: proof of concept for the clinical importance of endogenous vascular repair. Circulation 2005;111:2981-2987.
Chong AY, Blann AD, Patel J, Freestone B, Hughes E, Lip GY. Endothelial dysfunction and damage in congestive heart failure: relation of flow-mediated dilation to circulating endothelial cells, plasma indexes of endothelial damage, and brain natriuretic peptide. Circulation 2004;110:1794-1798.
Lee KW, Lip GY, Tayebjee M, Foster W, Blann AD. Circulating endothelial cells, von Willebrand factor, interleukin-6, and prognosis in patients with acute coronary syndromes. Blood 2005;105:526-532.
Lee KW, Blann AD, Lip GYH. Plasma markers of endothelial damage/dysfunction, inflammation and thrombogenesis in relation to TIMI risk stratification in acute coronary syndromes. Thromb Haemost (in press).
The authors reply: We thank the letter writers for their interesting comments. We disagree with Bertolini et al. that CD34+KDR+ cells are mature circulating endothelial cells. Endothelial cells are predominantly identified by the presence of CD1461 (and results of Boos et al., as described in their letter). In our study, CD146+ circulating endothelial cells were not predictive of cardiovascular outcomes, which indicates that CD34+KDR+ cells differ substantially from circulating endothelial cells (unpublished data).
It is an accepted standard to identify circulating endothelial progenitor cells by the presence of CD34 and KDR. To confirm the results, we measured CD133+ endothelial progenitor cells and obtained similar results, which appear in the online Supplementary Appendix to our article.
Drs. Ott and Taylor suggest calculating the absolute number of endothelial progenitor cells with the use of peripheral-blood mononuclear cells or lymphocytes. However, absolute cell counts measured by flow cytometry can be determined only with the use of enumeration systems (e.g., flow count beads).2 At present, we cannot think of a major advantage to measuring the absolute number of endothelial progenitor cells. The method provided allows a single measurement that is easy to perform, highly predictive, and transferable to other laboratories.
Dr. Kim and Dr. Leu and colleagues address the role of endothelial progenitor cells in acute coronary syndromes and acute myocardial infarction. Only one study has investigated the mobilization of CD34+KDR+ endothelial progenitor cells in myocardial infarction, whereas other studies have measured CD34+ cells or non–endothelial progenitor cell subfractions.3 None of the studies have systematically looked at the time course directly after acute myocardial infarction, owing to the fact that the exact onset of myocardial infarction is difficult to determine. Treatment of myocardial infarction requires the administration of multiple drugs that may influence the number of endothelial progenitor cells. Therefore, current data on progenitor cells in myocardial infarction are questionable. In order to elucidate the mobilization of endothelial progenitor cells after myocardial infarction, we measured the number of CD34+KDR+ cells in patients undergoing transcoronary ablation of septal hypertrophy (unpublished data). Preliminary results indicate that directly after myocardial infarction, the number of endothelial progenitor cells decreases as a result of consumption of cells within the ischemic region. The increase in cells described previously may be due to medical treatment. No patient who was included in the study had had a recent ischemic event, so misclassification of patients was not an issue.
We did not find an association between high-sensitivity CRP measures and the number of endothelial progenitor cells. To our knowledge, there are no data available on the association between endothelial progenitor cells and inflammatory markers in a similar population of patients. Data that are available come from in vitro, animal, and small-scale studies investigating the role of endothelial progenitor cells in acute coronary syndromes. Since our study population consisted mainly of patients with stable coronary artery disease, this may explain the lack of an association.
Nikos Werner, M.D.
Georg Nickenig, M.D.
University of Bonn
D-53105 Bonn, Germany
georg.nickenig@ukb.uni-bonn.de
References
Makin AJ, Blann AD, Chung NA, Silverman SH, Lip GY. Assessment of endothelial damage in atherosclerotic vascular disease by quantification of circulating endothelial cells: relationship with von Willebrand factor and tissue factor. Eur Heart J 2004;25:371-376.
Sims LC, Brecher ME, Gertis K, et al. Enumeration of CD34-positive stem cells: evaluation and comparison of three methods. J Hematother 1997;6:213-226.
Shintani S, Murohara T, Ikeda H, et al. Mobilization of endothelial progenitor cells in patients with acute myocardial infarction. Circulation 2001;103:2776-2779.
Dae Hyun Kim, M.D., M.P.H.
Thomas Jefferson University
Philadelphia, PA 19107
dae-hyun.kim@mail.tju.edu
References
Wojakowski W, Tendera M, Michalowska A, et al. Mobilization of CD34/CXCR4+, CD34/CD117+, c-met+ stem cells, and mononuclear cells expressing early cardiac, muscle, and endothelial markers into peripheral blood in patients with acute myocardial myocardial infarction. Circulation 2004;110:3213-3220.
Massa M, Rosti V, Ferrario M, et al. Increased circulating hematopoietic and endothelial progenitor cells in the early phase of acute myocardial infarction. Blood 2005;105:199-206.
Werner N, Kosiol S, Schiegl T, et al. Circulating endothelial progenitor cells and cardiovascular outcomes. N Engl J Med 2005;353:999-1007.
To the Editor: Werner et al. reported that increased levels of circulating endothelial progenitor cells were associated with a reduced risk of future cardiovascular events among patients with coronary artery disease, and the significant difference in the cumulative event rate according to the levels of endothelial progenitor cells could be shown early in the first four months of follow-up. However, several clinical observational studies, such as that by George et al.,1 demonstrated that patients with acute coronary syndromes had elevated levels of circulating endothelial progenitor cells. Furthermore, George et al. showed that the level of circulating endothelial progenitor cells decreased by nearly 50 percent, as did the level of C-reactive protein (CRP), in patients with unstable angina after their clinical condition had been stabilized for three months,1 suggesting that tissue ischemia has a role in triggering the mobilization of endothelial progenitor cells and may be correlated with systemic inflammation. In the article by Werner et al., half the patients had acute coronary syndromes or had undergone coronary angioplasty, and the levels of circulating endothelial progenitor cells may have been altered by these events. The CRP level is an independent predictor of adverse cardiovascular events.2 Additional information, such as serum level of CRP, might further elucidate the important role of endothelial progenitor cells in determining future events in patients with cardiovascular disease.
Hsin-Bang Leu, M.D.
Jaw-Wen Chen, M.D.
Shing-Jong Lin, M.D., Ph.D.
National Yang-Ming University
Taipei 112, Taiwan
doc1958h@yahoo.com.tw
References
George J, Goldstein E, Abashidze S, et al. Circulating endothelial progenitor cells in patients with unstable angina: association with systemic inflammation. Eur Heart J 2004;25:1003-1008.
Ridker PM, Hennekens CH, Buring JE, et al. C-reactive protein and other markers of inflammation in the prediction of cardiovascular disease in women. N Engl J Med 2000;342:836-843.
To the Editor: Werner and coauthors stated that "the level of circulating CD34+KDR+ endothelial progenitor cells predicts the occurrence of cardiovascular events and death from cardiovascular causes." They calculated the percentage of endothelial progenitor cells in the lymphocytic fraction of peripheral-blood mononuclear cells with fluorescence-activated cell-sorting analysis and as colony-forming units in 1x107 peripheral-blood mononuclear cells.
The fraction of mononuclear cells, as isolated by density gradient, is heterogeneous and contains cell populations that can vary considerably with disease or over time.1 Our concern is that measuring the percentage of endothelial progenitor cells in the lymphocytic fraction could underestimate or overestimate the concentration of endothelial progenitor cells if another cell population within that fraction is altered. To obtain an accurate concentration of blood endothelial progenitor cells, the percentage of such cells measured in the lymphocytic fraction by fluorescence-activated cell sorting or the number of colony-forming units in mononuclear cells should be multiplied by the total lymphocyte or mononuclear-cell count.2,3 To gain more insight into the mechanism and to detect a possible causative relation, it would be important to determine the prognostic value of absolute counts of endothelial progenitor cells.
Harald C. Ott, M.D.
Doris A. Taylor, Ph.D.
University of Minnesota
Minneapolis, MN 55455
ottxx057@umn.edu
References
B?yum A. Separation of white blood cells. Nature 1964;204:793-794.
Elliott C, Samson DM, Armitage S, et al. When to harvest peripheral-blood stem cells after mobilization therapy: prediction of CD34-positive cell yield by preceding day CD34-positive concentration in peripheral blood. J Clin Oncol 1996;14:970-973.
Fukuda J, Kaneko T, Egashira M, Oshimi K. Direct measurement of CD34+ blood stem cell absolute counts by flow cytometry. Stem Cells 1998;16:294-300.
To the Editor: Werner et al. show that the number of blood cells with a CD34+KDR+ phenotype predicts the occurrence of cardiovascular events and death from cardiovascular causes. They indicate that these cells are circulating endothelial progenitors. However, CD34 and KDR are antigens expressed not only by endothelial progenitors but also by endothelial cells with a mature phenotype, which make up the large majority of all circulating endothelial cells.1,2 This is a subtle but relevant issue, because it is still unclear whether in adult life most of vascular remodeling is due to angiogenesis (mature cell–derived generation of new vessels) or vasculogenesis (endothelial progenitor cell–dependent generation of new vessels).3,4 So far, only the CD133 antigen is known to be expressed by endothelial progenitors and not by mature endothelial cells.4 Thus, in the population of patients in the study by Werner et al., it would be useful to dissect clinical outcomes associated with the number of CD133+CD34+KDR+ endothelial progenitors from those associated with the number of CD34+KDR+ cells, a majority of which are mature circulating endothelial cells.
Francesco Bertolini, M.D., Ph.D.
Patrizia Mancuso, Ph.D.
European Institute of Oncology
20141 Milan, Italy
francesco.bertolini@ieo.it
Robert S. Kerbel, Ph.D.
Sunnybrook and Women's College Health Sciences Centre
Toronto, ON M4N 3M5, Canada
References
Mancuso P. Resting and activated endothelial cells are increased in the peripheral blood of cancer patients. Blood 2001;97:3658-3661.
Blann AD, Woywodt A, Bertolini F, et al. Circulating endothelial cells: biomarker of vascular disease. Thromb Haemost 2005;93:228-235.
Jain RK. Molecular regulation of vessel maturation. Nat Med 2003;9:685-693.
Rafii S, Lyden D, Benezra R, Hattori K, Heissig B. Vascular and haematopoietic stem cells: novel targets for anti-angiogenesis therapy? Nat Rev Cancer 2002;2:826-835.
To the Editor: With regard to the study by Werner et al., we would like to highlight a distinctly different endothelial cell type, the CD146+ circulating endothelial cell. These are mature cells that have detached from the endothelium as a result of endothelial injury or damage. The number of circulating endothelial cells has been shown to correlate closely with several well-recognized indexes of endothelial damage or dysfunction.1,2 We have previously reported an increased number of circulating endothelial cells in a cohort of 156 patients presenting with acute coronary syndromes.3 The number of circulating endothelial cells at 48 hours after admission was a strong and independent predictor of the risk of death and major adverse cardiac events at 30 days and at 1 year.3 There is also a clear relationship between an increased number of circulating endothelial cells and high Thrombolysis in Myocardial Infarction (TIMI) risk scores in acute coronary syndromes.4
Current data suggest that circulating endothelial cells and endothelial progenitor cells are separate cell populations. The former are mature endothelial cells that are shed from the vessel wall in response to damage, whereas the latter are involved in vascular regeneration or repair. Clearly, more information is needed for an adequate definition of the interrelationships between these two types of cells.
Christopher J. Boos, M.B., B.S., M.R.C.P.
Patrick K.Y. Goon, M.R.C.S.
Gregory Y.H. Lip, M.D.
City Hospital
Birmingham B18 7QH, United Kingdom
g.y.h.lip@bham.ac.uk
References
Schmidt-Lucke C, Rossig L, Fichtlscherer S, et al. Reduced number of circulating endothelial progenitor cells predicts future cardiovascular events: proof of concept for the clinical importance of endogenous vascular repair. Circulation 2005;111:2981-2987.
Chong AY, Blann AD, Patel J, Freestone B, Hughes E, Lip GY. Endothelial dysfunction and damage in congestive heart failure: relation of flow-mediated dilation to circulating endothelial cells, plasma indexes of endothelial damage, and brain natriuretic peptide. Circulation 2004;110:1794-1798.
Lee KW, Lip GY, Tayebjee M, Foster W, Blann AD. Circulating endothelial cells, von Willebrand factor, interleukin-6, and prognosis in patients with acute coronary syndromes. Blood 2005;105:526-532.
Lee KW, Blann AD, Lip GYH. Plasma markers of endothelial damage/dysfunction, inflammation and thrombogenesis in relation to TIMI risk stratification in acute coronary syndromes. Thromb Haemost (in press).
The authors reply: We thank the letter writers for their interesting comments. We disagree with Bertolini et al. that CD34+KDR+ cells are mature circulating endothelial cells. Endothelial cells are predominantly identified by the presence of CD1461 (and results of Boos et al., as described in their letter). In our study, CD146+ circulating endothelial cells were not predictive of cardiovascular outcomes, which indicates that CD34+KDR+ cells differ substantially from circulating endothelial cells (unpublished data).
It is an accepted standard to identify circulating endothelial progenitor cells by the presence of CD34 and KDR. To confirm the results, we measured CD133+ endothelial progenitor cells and obtained similar results, which appear in the online Supplementary Appendix to our article.
Drs. Ott and Taylor suggest calculating the absolute number of endothelial progenitor cells with the use of peripheral-blood mononuclear cells or lymphocytes. However, absolute cell counts measured by flow cytometry can be determined only with the use of enumeration systems (e.g., flow count beads).2 At present, we cannot think of a major advantage to measuring the absolute number of endothelial progenitor cells. The method provided allows a single measurement that is easy to perform, highly predictive, and transferable to other laboratories.
Dr. Kim and Dr. Leu and colleagues address the role of endothelial progenitor cells in acute coronary syndromes and acute myocardial infarction. Only one study has investigated the mobilization of CD34+KDR+ endothelial progenitor cells in myocardial infarction, whereas other studies have measured CD34+ cells or non–endothelial progenitor cell subfractions.3 None of the studies have systematically looked at the time course directly after acute myocardial infarction, owing to the fact that the exact onset of myocardial infarction is difficult to determine. Treatment of myocardial infarction requires the administration of multiple drugs that may influence the number of endothelial progenitor cells. Therefore, current data on progenitor cells in myocardial infarction are questionable. In order to elucidate the mobilization of endothelial progenitor cells after myocardial infarction, we measured the number of CD34+KDR+ cells in patients undergoing transcoronary ablation of septal hypertrophy (unpublished data). Preliminary results indicate that directly after myocardial infarction, the number of endothelial progenitor cells decreases as a result of consumption of cells within the ischemic region. The increase in cells described previously may be due to medical treatment. No patient who was included in the study had had a recent ischemic event, so misclassification of patients was not an issue.
We did not find an association between high-sensitivity CRP measures and the number of endothelial progenitor cells. To our knowledge, there are no data available on the association between endothelial progenitor cells and inflammatory markers in a similar population of patients. Data that are available come from in vitro, animal, and small-scale studies investigating the role of endothelial progenitor cells in acute coronary syndromes. Since our study population consisted mainly of patients with stable coronary artery disease, this may explain the lack of an association.
Nikos Werner, M.D.
Georg Nickenig, M.D.
University of Bonn
D-53105 Bonn, Germany
georg.nickenig@ukb.uni-bonn.de
References
Makin AJ, Blann AD, Chung NA, Silverman SH, Lip GY. Assessment of endothelial damage in atherosclerotic vascular disease by quantification of circulating endothelial cells: relationship with von Willebrand factor and tissue factor. Eur Heart J 2004;25:371-376.
Sims LC, Brecher ME, Gertis K, et al. Enumeration of CD34-positive stem cells: evaluation and comparison of three methods. J Hematother 1997;6:213-226.
Shintani S, Murohara T, Ikeda H, et al. Mobilization of endothelial progenitor cells in patients with acute myocardial infarction. Circulation 2001;103:2776-2779.