Endothelial Progenitor Cells
Angiogenesis, the process of the formation and sprouting of new blood vessels, occurs during embryonic development and adult life. Traditionally, it was believed that the formation of the fetal capillary network from migrating endothelial precursor cells was restricted to early embryonic development, while blood vessels in an adult organism are derived from endothelial cells located in situ. However, recent evidence suggests that endothelial precursor cells residing remotely from the site of neoangiogenesis may play an important physiological role in postnatal endothelialization.
Understanding of Endothelial Cell (EC) biology is essential for the development of new methodologies to treat cancer, promote vascular healing, provide suitable coating for vascular grafts and for the delivery of toxins to tumor vascular beds. The therapeutic potential of endothelial progenitor cells was recently demonstrated by the improved blood flow rates and capillary density following the transplantation of endothelial progenitor cells in hindlimb ischemic mice (Kalka et al., 2000, PNAS).
Kits for Differentiation Analysis
The ability of CD34+ cells to differentiate into ECs can be determined in vitro by plating endothelial cells onto an extracellular matrix, such as collagen, fibronectin and/or gelatin. Adherent colonies of ECs can subsequently be detected by their morphology and immunoreactivity to EC markers (Rafii et al., 1995, Ann. Thorac. Surg.; Rafii, 2000, J. Clin. Invest.; Shi et al.,1998, Blood).
Further analysis of endothelial progenitor cell homing and incorporation into sites of neovascularization (e.g. following bone marrow transplant into recipient mice) can be performed using antibodies specific for human endothelial cell markers (Kalka et al., 2000, PNAS).
Millipore supplies a number of growth factors for the in vitro expansion of endothelial precursor cells. One such combination, comprising VEGF, bFGF, IGF and EGF, resulted in an 80 to 90 fold expansion of cells expressing EC antigens (Kalka et al., 2000, PNAS). Other growth factor combinations described in the literature include VEGF, bFGF and IGF (Suhonen et al., 1996, Nature), and bFGF together with VEGF (Peichev et al., 2000, Blood).
Endothelial Progenitor Cell Markers
The identification and isolation of endothelial progenitor cells is difficult due to the absence of specific endothelial markers and functional assays to distinguish migrating endothelial progenitor cells from sloughed mature ECs (Rafii et al., 1995, Ann. Thorac. Surg.). ECs and mature vessel wall-derived ECs express common endothelial-specific markers including; VEGFR-2 (KDR, Flk-1) (Eichmann et al., 1997, PNAS), Tie-2 and Tie-1 (Sato et al., 1993, PNAS; Suri et al., 1996, Cell), VE-Cadherin (Johnstone et al., 1998, Exp. Cell. Res.), CD34, PECAM (CD31) and E-Selectin (Peichev et al., 2000, Blood). Other endothelial markers include von Willebrand factor (vWF), P1H12 (CD146), thrombomodulin, CD36, Endoglin and Integrin aVß3. Identification of ECs is further complicated by the fact that hematopoietic stem cells and their progeny (particularly monocytes) express markers similar to those expressed by endothelial cells, such as VEGFR-1 (Flt-1), CD34, PECAM-1, Tie-1, Tie-2 and von Willebrand’s factor (Peichev et al., 2000, Blood). The antibody P1H12 (CD146) has been used successfully for the enrichment of circulating ECs from peripheral blood prior to analysis (Lin et al., 2000, J. Clin. Invest.).
Pure populations of CD34+/VEGFR-2+ circulating endothelial progenitor cells have been isolated from fetal liver, mobilized peripheral blood and cord blood (Peichev et al., 2000, Blood). These cells express EC specific markers including VE-Cadherin and E-selectin. Additionally, the majority of these cells express the novel hematopoietic stem cell marker AC133 (CD133) and the chemokine receptor CXCR4.
When isolating endothelial progenitor cells from whole blood by flow cytometry, contaminating monocytes and other blood cells can be removed using the markers CD14, CD15 and CD45.
The application of Millipore’s antibodies to EC markers will facilitate the study of EC biology and the development of therapeutic applications.
Endothelial Progenitor Cell Markers
| CD14 [LPS RECEPTOR] | C-X-X-X-C CHEMOKINE RECEPTOR 1 |
| CD15 [LEWIS X; 3-FAL] | ENDOGLIN [CD105] |
| CD31 [PECAM-1] | E-SELECTIN [CD62E] |
| CD34 CLASS I | INTEGRIN aV [CD51] |
| CD34 CLASS II | INTEGRIN aVß3 [CD51/CD61] |
| CD34 CLASS III | PECAM-1 [CD31] |
| CD36 [PLATELET GLYCOPROTEIN IV] | TIE-1 |
| CD45 | TIE-2 |
| CD133 [PROMININ-1] | VE-CADHERIN [CD144] |
| CD141 [THROMBOMODULIN; FETOMODULIN] | VEGF RECEPTOR-2 |
| CD146 [MUC18; MCAM; P1H12] | VON WILLEBR AND FACTOR |
| C-X-C CHEMOKINE RECEPTOR 4 |  |
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