Defining human endothelial progenitor cells

Summary. Thereisnospeciﬁcmarkertoidentifyanendothelialprogenitorcell(EPC)andthisdeﬁciencyisrestrictingtheabilityof an entire ﬁeld of research in deﬁning these cells. We willreviewcurrentmethodstodeﬁneEPCinthehumansystemandsuggestapproachestodeﬁnebetterthecellpopulationsinvolvedinneoangiogenesis.PubMedwasusedtoidentifyarticlesviathesearch term endothelial progenitor cell and those articlesfocused on deﬁning the term were evaluated. The only humancells expressing the characteristics of an EPC, as originallyproposed, are endothelial colony forming cells. A variety ofhematopoietic cells including stem and progenitors, participatein initiating and modulating neoangiogenesis. Future studiesmust focus on deﬁning the speciﬁc hematopoietic subsets thatare involved in activating, recruiting, and remodeling thevascular networks formed by the endothelial colony formingcells.Keywords: endothelial colony forming cells, endothelial pro-genitor cells, hematopoietic progenitor cells, neoangiogenesis.Deﬁning human endothelial progenitor cellsThe term endothelial progenitor cell (EPC) remains difﬁcult todeﬁne due to the lack of a speciﬁc and unique cell surface ormoleculardeterminantthatpermitsprospectiveisolationofthiscell. In this overview, we will discuss the most commonmethods for EPCidentiﬁcationandhighlightsomepitfalls thatmay be encountered with each approach. We will discuss someof the limitations to the ﬁeld that will need to be overcome forthe development of a consensus deﬁnition of a humancirculating EPC. It is important that the ﬁeld continues tostrive to identify accurately which cells form the EPCpopulation in order to assure future clinical trials of using themost approriate cell therapy.In the human system, EPC are deﬁned using three generalapproaches. First, peripheral blood mononuclear cells areisolated and plated on ﬁbronectin coated tissue culture plateswith a variety ofendothelial growth factors. After several days,the non-adherent cells are removed and the adherent cellpopulation remaining, which express the ability to ingestacetylated low density lipoprotein and to bind certain plantlectins, is considered to represent EPC. An inverse correlationbetween the circulating concentration of these putative EPCand an increased risk for developing coronary arterial diseasehas been reported [1,2]. However, this deﬁnition fails torecognize that ﬁbronectin coated tissue culture wells have beenused to isolate human blood monocytes [3]. Human peripheralblood monocytes can be cultured in serum containing mediumwith a variety of growth factors that promote expression ofnumerous proteins normally expressed by primary endothelialcells [4–6] and such modulating culture conditions may lead topathways of alternative macrophage activation that supportenhanced wound healing and angiogenic activities [7,8]. Thus,use of the short term adhesion and culture method for putativeEPC isolation is fraught with complexities in identifying whichcell may be playing a functional role attributable to thiscultured population.A second approach to putative EPC identiﬁcation utilizesmonoclonal antibodies and ﬂuorescence activated cell sorting(FACS) analysis to enumerate speciﬁc cell populations.Asahara et al. [9] reasoned that putative EPC may express cellsurface markers shared by hematopoietic stem cells (HSC)since endothelial and blood cells share a similar mesodermalorigin during embryonic development. Thus, Asahara et al.cultured CD34