Presence of endothelial progenitor cells, distinct from mature endothelial cells, within human CD146+ blood cells

Bruno Delorme; Agnès Basire; Carla Gentile; Florence Sabatier; Frédéric Monsonis; Christophe Desouches; Marcel Blot-Chabaud; Georges Uzan; José Sampol; Françoise Dignat-George

doi:10.1160/TH05-07-0499

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 2005; 94(06): 1270-1279
DOI: 10.1160/TH05-07-0499

Endothelium and Vascular Development

Schattauer GmbH

Presence of endothelial progenitor cells, distinct from mature endothelial cells, within human CD146⁺ blood cells

Authors

Bruno Delorme

¹INSERM Unité 608, Laboratoire d’Hématologie et d’Immunologie, UFR de Pharmacie, Université de la Méditerranée, Marseille, France
Agnès Basire

¹INSERM Unité 608, Laboratoire d’Hématologie et d’Immunologie, UFR de Pharmacie, Université de la Méditerranée, Marseille, France
Carla Gentile

³INSERM Unité 602, Groupe Hospitalier Paul Brousse, Villejuif, France
Florence Sabatier

¹INSERM Unité 608, Laboratoire d’Hématologie et d’Immunologie, UFR de Pharmacie, Université de la Méditerranée, Marseille, France

²Laboratoire d’Hématologie, Centre Hospitalier Universitaire La Conception, Marseille, France
Frédéric Monsonis

⁴Beckman Coulter Immunotech Company, Marseille, France
Christophe Desouches

⁵Service de Chirurgie Plastique, Réparatrice et Esthétique, Centre Hospitalier Universitaire La Conception, Marseille, France
Marcel Blot-Chabaud

¹INSERM Unité 608, Laboratoire d’Hématologie et d’Immunologie, UFR de Pharmacie, Université de la Méditerranée, Marseille, France
Georges Uzan

³INSERM Unité 602, Groupe Hospitalier Paul Brousse, Villejuif, France
José Sampol

¹INSERM Unité 608, Laboratoire d’Hématologie et d’Immunologie, UFR de Pharmacie, Université de la Méditerranée, Marseille, France

²Laboratoire d’Hématologie, Centre Hospitalier Universitaire La Conception, Marseille, France
Françoise Dignat-George

¹INSERM Unité 608, Laboratoire d’Hématologie et d’Immunologie, UFR de Pharmacie, Université de la Méditerranée, Marseille, France

²Laboratoire d’Hématologie, Centre Hospitalier Universitaire La Conception, Marseille, France

Abstract

Summary

CD146 is an adhesion molecule present on endothelial cells throughout the vascular tree. CD146 is also expressed by circulating endothelial cells (CECs) widely considered to be mature endothelial cells detached from injured vessels. The discovery of circulating endothelial progenitor cells (EPCs) originating from bone marrow prompted us to investigate whether CD146 circulating cells could also contains EPCs. We tested this hypothesis using an approach combining elimination of CECs by an adhesion step, followed by immunomagnetic sorting of remaining CD146⁺ cells from the non adherent fraction of cord blood mononuclear cells. When cultured under endothelial-promoting conditions, these cells differentiated as late outgrowth endothelial colonies: they grew as a cobblestone monolayer, were uniformly positive for endothelial markers and did not express leukocyte antigens. They highly proliferated and were expanded in long-term culture without alterations of their phenotypic and functional properties (DiI-ac-LDL uptake, wound repair, capillary-like network formation, and TNFα response). Moreover, these cells colonized a Matrigel plug in immunodeficient mice (NOD/SCID). Finally, using 4-color flow cytometry analysis of purified CD34+ cells, we clearly discriminated, CD146⁺ EPCs (CD146⁺ CD34⁺ CD45⁺ CD133⁺ or CD117⁺), and CD146⁺ CECs (CD146⁺ CD34⁺, CD45⁻ CD133⁻ or CD117⁻), both in cord and adult peripheral blood. The relative proportions of the two CD146⁺ subsets varied in patients with myocardial infarction as compared to healthy subjects. Our study establishes that, beside CECs, CD146⁺ circulating cells contain a subpopulation of EPCs with potential use in proangiogenic therapy. In addition, the dual measurement of CD146⁺ CECs and CD146⁺ EPCs offers a promising tool for monitoring vascular injury/regeneration processes in clinical situations.

Keywords

CD146 - angiogenesis - myocardial infarction - circulating endothelial cells - progenitor endothelial cells

Full Text

References

References
1 Bardin N. et al S-Endo 1, a pan-endothelial monoclonal antibody recognizing a novel human endothelial antigen. Tissue Antigens 1996; 48: 531-9.
2 Bardin N. et al Identification of the S-Endo 1 endothelial-associated antigen. Biochem Biophys Res Commun 1996; 218: 210-6.
3 Bardin N. et al Identification of CD146 as a component of the endothelial junction involved in the control of cell-cell cohesion. Blood 2001; 98: 3677-84.
4 Anfosso F. et al Activation of human endothelial cells via S-endo-1 antigen (CD146) stimulates the tyrosine phosphorylation of focal adhesion kinase p125(FAK). J Biol Chem 1998; 273: 26852-6.
5 Anfosso F. et al Outside-in signaling pathway linked to CD146 engagement in human endothelial cells. J Biol Chem 2001; 276: 1564-9.
6 Blann AD. et al Circulating endothelial cells : biomarker of vascular disease. Thromb Haemost 2005; 93: 228-35.
7 George F. et al Demonstration of Rickettsia conorii-induced endothelial injury in vivo by measuring circulating endothelial cells, thrombomodulin, and von Willebrand factor in patients with Mediterranean spotted fever. Blood 1993; 82: 2109-16.
8 Quilici J. et al Circulating endothelial cell count as a diagnostic marker for non-ST-elevation acute coronary syndromes. Circulation 2004; 110: 1586-91.
9 Woywodt A. et al Circulating endothelial cells as markers for ANCA-associated small-vessel vasculitis. Lancet 2003; 361: 206-10.
10 Solovey A. et al Circulating activated endothelial cells in sickle cell anemia. N Engl J Med 1997; 337: 1584-90.
11 Popa ER. et al Donor-derived circulating endothelial cells after kidney transplantation. Transplantation 2002; 74: 1320-7.
12 Dignat-George F, Sampol J. Circulating endothelial cells in vascular disorders: new insights into an old concept. Eur J Haematol 2000; 65: 215-20.
13 Rajagopalan S. et al Endothelial cell apoptosis in systemic lupus erythematosus: a common pathway for abnormal vascular function and thrombosis propensity. Blood 2004; 103: 3677-83.
14 Clancy R. et al Circulating activated endothelial cells in systemic lupus erythematosus: further evidence for diffuse vasculopathy. Arthritis Rheum 2001; 44: 1203-8.
15 Beerepoot LV. et al Increased levels of viable circulating endothelial cells are an indicator of progressive disease in cancer patients. Ann Oncol 2004; 15: 139-45.
16 Rafii S, Lyden D. Therapeutic stem and progenitor cell transplantation for organ vascularization and regeneration. Nat Med 2003; 9: 702-12.
17 Asahara T. et al Isolation of putative progenitor endothelial cells for angiogenesis. Science 1997; 275: 964-7.
18 Lin Y. et al Origins of circulating endothelial cells and endothelial outgrowth from blood. J Clin Invest 2000; 105: 71-7.
19 Gulati R. et al Diverse origin and function of cells with endothelial phenotype obtained from adult human blood. Circ Res 2003; 93: 1023-5.
20 Hur J, Yoon CH, Kim HS. et al Characterization of two types of endothelial progenitor cells and their different contributions to neovasculogenesis. Arterioscler Thromb Vasc Biol 2004; 24: 288-93.
21 Hill JM, Zalos G, Halcox JP. et al Circulating endothelial progenitor cells, vascular function, and cardiovascular risk. N Engl J Med 2003; 348: 593-600.
22 Choi JH. et al Decreased number and impaired angiogenic function of endothelial progenitor cells in patients with chronic renal failure. Arterioscler Thromb Vasc Biol 2004; 24: 1246-52.
23 Vasa M. et al Number and migratory activity of circulating endothelial progenitor cells inversely correlate with risk factors for coronary artery disease. Circ Res 2001; 89: E1-7.
24 Woywodt A. et al Circulating endothelial cells as a marker of endothelial damage in allogeneic hematopoietic stem cell transplantation. Blood 2004; 103: 3603-5.
25 Burger PE. et al Fibroblast growth factor receptor-1 is expressed by endothelial progenitor cells. Blood 2002; 100: 3527-35.
26 Rehman J. et al Peripheral blood “endothelial progenitor cells” are derived from monocyte/macrophages and secrete angiogenic growth factors. Circulation 2003; 107: 1164-9.
27 Schmeisser A. et al Monocytes coexpress endothelial and macrophagocytic lineage markers and form cord-like structures in Matrigel under angiogenic conditions. Cardiovasc Res 2001; 49: 671-80.
28 Miraglia S. et al A novel five-transmembrane hematopoietic stem cell antigen: isolation, characterization, and molecular cloning. Blood 1997; 90: 5013-21.
29 Kalka C. et al Transplantation of ex vivo expanded endothelial progenitor cells for therapeutic neovascularization. Proc Natl Acad Sci U S A 2000; 97: 3422-7.
30 Kocher AA. et al Neovascularization of ischemic myocardium by human bone-marrow-derived angioblasts prevents cardiomyocyte apoptosis, reduces remodeling and improves cardiac function. Nat Med 2001; 7: 430-6.
31 Tateishi-Yuyama E. et al Therapeutic angiogenesis for patients with limb ischaemia by autologous transplantation of bone-marrow cells: a pilot study and a randomised controlled trial. Lancet 2002; 360: 427-35.
32 Assmus B. et al Transplantation of progenitor cells and regeneration enhancement in acute myocardial infarction. Circulation 2002; 106: 3009-17.
33 Kudo FA. et al Autologous transplantation of peripheral blood endothelial progenitor cells (CD34+) for therapeutic angiogenesis in patients with critical limb ischemia. Int Angiol 2003; 22: 344-8.
34 Peichev M, Naiyer AJ, Pereira D. et al Expression of VEGFR-2 and AC133 by circulating human CD34(+) cells identifies a population of functional endothelial precursors. Blood 2000; 95: 952-8.
35 Gehling UM. et al In vitro differentiation of endothelial cells from AC133-positive progenitor cells. Blood 2000; 95: 3106-12.
36 Bompais H. et al Human endothelial cells derived from circulating progenitors display specific functional properties compared with mature vessel wall endothelial cells. Blood 2004; 103: 2577-84.
37 Pelosi E. et al Identification of the hemangioblast in postnatal life. Blood 2002; 100: 3203-8.
38 Schatteman GC, Awad O. Hemangioblasts, angioblasts, and adult endothelial cell progenitors. Anat Rec 2004; 276A: 13-21.
39 Grant MB. et al Adult hematopoietic stem cells provide functional hemangioblast activity during retinal neovascularization. Nat Med 2002; 8: 607-12.
40 Filshie RJ. et al MUC18, a member of the immunoglobulin superfamily, is expressed on bone marrow fibroblasts and a subset of hematological malignancies. Leukemia 1998; 12: 414-21.
41 Ingram DA. et al Identification of a novel hierarchy of endothelial progenitor cells using peripheral and umbilical cord blood. Blood 2004; 104: 2752-60.
42 Alais S. et al HEMCAM/CD146 downregulates cell surface expression of β1 integrins. J Cell Sci 2001; 114: 1847-59.
43 Solovey AN. et al Identification and functional assessment of endothelial P1H12. J Lab Clin Med 2001; 138: 322-31.
44 Yan X. et al A novel anti-CD146 monoclonal antibody, AA98, inhibits angiogenesis and tumor growth. Blood 2003; 102: 184-91.
45 Shintani S. et al Mobilization of endothelial progenitor cells in patients with acute myocardial infarction. Circulation 2001; 103: 2776-9.
46 Tepper OM. et al Adult vasculogenesis occurs through the in situ recruitment, proliferation and tubulization of circulating bone marrow-derived cells. Blood 2004; 105: 1068-77.