Application Of Endothelial Progenitor Cells Research Articles

Letter Regarding Article by Rotmans et al, “In Vivo Cell Seeding With Anti-CD34 Antibodies Successfully Accelerates Endothelialization but Stimulates Intimal Hyperplasia in Porcine Arteriovenous Expanded Polytetrafluoroethylene Grafts”

To the Editor: Rapid (re)endothelialization of autologous and synthetic vascular grafts is regarded as a key element in reducing the risk of acute thrombosis and, by limiting adverse vessel wall remodeling, which includes intimal hyperplasia, preserving long-term patency. Rotmans et al1 (and the accompanying editorial2) challenge this tenet of vascular biology in their report of human anti-CD34 antibody–coated polytetrafluoroethylene (PTFE) grafts in a porcine AV fistula model. Rapid “endothelialization” of grafts is described, which, rather than improving functional outcome, was associated with significantly increased intimal hyperplasia at the venous anastomosis. The authors conclude that future studies will need to address a putative deficiency in the functional capability of captured endothelial progenitor cells (EPCs), highlighting approaches based on the use of regulatory proteins or alternative cell combinations. In our opinion, the data do not support these conclusions. The graft endoluminal cells were inadequately characterized with lectin (a surface antigen shared by several blood-borne …

Therapeutic Potential of Endothelial Progenitor Cells in Cardiovascular Diseases

Endothelial dysfunction and cell loss are prominent features in cardiovascular disease. Endothelial progenitor cells (EPCs) originating from the bone marrow play a significant role in neovascularization of ischemic tissues and in re-endothelialization of injured blood vessels. Several studies have shown the therapeutic potential of EPC transplantation in rescue of tissue ischemia and in repair of blood vessels and bioengineering of prosthetic grafts. Recent small-scale trials have provided preliminary evidence of feasibility, safety, and efficacy in patients with myocardial and critical limb ischemia. However, several studies have shown that age and cardiovascular disease risk factors reduce the availability of circulating EPCs (CEPCs) and impair their function to varying degrees. In addition, the relative scarcity of CEPCs limits the ability to expand these cells in sufficient numbers for some therapeutic applications. Priority must be given to the development of strategies to enhance the number and improve the function of CEPCs. Furthermore, alternative sources of EPC such as chord blood need to be explored. Strategies for improvement of cell adhesion, survival, and prevention of cell senescence are also essential to ensure therapeutic viability. Genetic engineering of EPCs may be a useful approach to developing these cells into efficient therapeutic tools. In the clinical arena there is pressing need to standardize the protocols for isolation, culture, and therapeutic application of EPC. Large-scale multi-center randomized trials are required to evaluate the long-term safety and efficacy of EPC therapy. Despite these hurdles, the outlook for EPC-based therapy for cardiovascular disease is promising.

Endothelial Progenitor Cells (EPCs) Isolated from the Rhesus Macaque but Not the Mouse Are Phenotypically and Functionally Similar to Human EPCs.

We recently identified a novel hierarchy of human endothelial progenitor cells (EPCs), which are functionally defined by their proliferative and clonogenic potential (Blood, 2004). Emerging evidence suggests that EPCs may be used as angiogenic therapies, or as biomarkers to assess cardiovascular disease risk. Thus, identification of animal models, which phenocopy the human EPC hierarchy, is an important priority for preclinical testing of experimental therapeutics. Given the importance of the Rhesus Macaque as a preclinical model, we tested whether EPCs could be isolated from the peripheral blood of the Rhesus Macaque and compared to EPCs isolated from human adult peripheral blood. Mononuclear cells were isolated from 20 ml of Rhesus peripheral blood and cultured in EGM-2 medium, which promotes the formation of EPC colonies. After 7 days in culture, we identified approximately 20 endothelial cell colonies (n=9), which appeared identical to human EPC colonies. We subcultured the endothelial cell colonies into monolayers for immunophenotyping and functional analysis. Endothelial cells (ECs) derived from the Rhesus EPC colonies formed vessels in matrigel, and demonstrated uptake of acetylated LDL, which are characteristics of ECs. Similar to ECs derived from human EPCs, Rhesus ECs expressed the endothelial cell antigens, CD31, CD144, CD105, CD146, and Flk1. Importantly, Rhesus ECs did not express the hematopoietic cell specific antigens, CD45 and CD14. Similar to ECs derived from human peripheral blood EPC colonies, Rhesus ECs could be serially passaged for at least 40 population doublings without signs of cellular senescence. A hallmark of stem and progenitor cells is their ability to proliferate and give rise to functional progeny. Analogous to a paradigm established in the hematopoietic cell system, we recently developed a single cell deposition assay to reproducibly identify the following human EPCs: (1) high proliferative potential - endothelial colony forming cells (HPP-ECFC), which form macroscopic colonies that form secondary and tertiary colonies upon replating, (2) low proliferative potential - endothelial colony forming cells (LPP-ECFC), which form colonies greater than 50 cells, but do not form secondary colonies upon replating, (3) endothelial cell clusters (EC-clusters) that contain less than 50 cells, and (4) mature terminally differentiated endothelial cells (EC), which do not divide (Blood, 2004). To determine whether these different populations of EPCs could be identified in the ECs derived from Rhesus EPCs, we performed single cells deposition assays on 1,000 cells. All types of EPCs could be identified in the Rhesus ECs (Table I). Further, ECs derived from the Rhesus EPCs rapidly form chimeric vessels with human ECs derived from adult blood, implying that the molecular mechanisms critical for vessel formation are conserved between the two species. Finally, while the murine model is an animal model widely used for studying EPCs, a similar hierarchy of EPCs could not be established from the peripheral blood of mice. Thus, given the diversity of therapeutic applications of EPCs for treating a variety of human diseases, these studies establish the Rhesus Macaque as an important preclinical model.Percent of 1,000 Single Cells PlatedMature ECEC-ClusterLPP-ECFCHPP-ECFCRhesus ECs85.8±2.14.2±1.17.8±0.51.3±0.5Human ECs80.8±9.68.6±1.412.4±8.10.2±0.2

Endothelial Progenitor Cell Culture for Vascular Regeneration

A certain population of mononuclear cells in the peripheral blood is capable of contributing to new vessel formation by differentiating into endothelial cells. The basis for native as well as therapeutic neovascularization is not restricted to angiogenesis but includes postnatal vasculogenesis. These cells were discovered by Asahara in 1997 and named endothelial progenitor cells (EPCs). Clinical usefulness of EPCs from human peripheral blood is also suggested in the animal experiments. These results indicate that administering EPCs can be a new clinical strategy for treating ischemic disease, diabetic retinopathy, or neoplasm in which the promotion or inhibition of neovascular formation is critical. However, expansion of EPCs ex vivo is not currently suitable for the clinical setting because of the animal products that are necessary for EPC expansion. To approach the autologous cell-based clinical application of EPCs, we established EPC culture using auto serum. In this article, we will discuss the sources that can generate EPCs and show the usefulness and potential of EPC culture using auto serum in the basic and clinical setting of neovascular formation.

Endothelial progenitor cell culture for vascular regeneration.

A certain population of mononuclear cells in the peripheral blood is capable of contributing to new vessel formation by differentiating into endothelial cells. The basis for native as well as therapeutic neovascularization is not restricted to angiogenesis but includes postnatal vasculogenesis. These cells were discovered by Asahara in 1997 and named endothelial progenitor cells (EPCs). Clinical usefulness of EPCs from human peripheral blood is also suggested in the animal experiments. These results indicate that administering EPCs can be a new clinical strategy for treating ischemic disease, diabetic retinopathy, or neoplasm in which the promotion or inhibition of neovascular formation is critical. However, expansion of EPCs ex vivo is not currently suitable for the clinical setting because of the animal products that are necessary for EPC expansion. To approach the autologous cell-based clinical application of EPCs, we established EPC culture using auto serum. In this article, we will discuss the sources that can generate EPCs and show the usefulness and potential of EPC culture using auto serum in the basic and clinical setting of neovascular formation.

Endothelial progenitor cells: mobilization, differentiation, and homing.

Postnatal bone marrow contains a subtype of progenitor cells that have the capacity to migrate to the peripheral circulation and to differentiate into mature endothelial cells. Therefore, these cells have been termed endothelial progenitor cells (EPCs). The isolation of EPCs by adherence culture or magnetic microbeads has been described. In general, EPCs are characterized by the expression of 3 markers, CD133, CD34, and the vascular endothelial growth factor receptor-2. During differentiation, EPCs obviously lose CD133 and start to express CD31, vascular endothelial cadherin, and von Willebrand factor. EPCs seem to participate in endothelial repair and neovascularization of ischemic organs. Clinical studies using EPCs for neovascularization have just been started; however, the mechanisms stimulating or inhibiting the differentiation of EPC in vivo and the signals causing their migration and homing to sites of injured endothelium or extravascular tissue are largely unknown at present. Thus, future studies will help to explore areas of potential basic research and clinical application of EPCs.