Abstract-Endothelial progenitor cell (EPC) cultures and colony-forming units (CFUs) have been extensively studied for their therapeutic and diagnostic potential. Recent data suggest a role for EPCs in the release of proangiogenic factors. To identify factors secreted by EPCs, conditioned medium from EPC cultures and CFUs was analyzed using a matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometer combined with offline peptide separation by nanoflow liquid chromatography. Results were verified by RT-PCR and multiplex cytokine assays and complemented by a cellular proteomic analysis of cultured EPCs and CFUs using difference in-gel electrophoresis. This extensive proteomic analysis revealed the presence of the proangiogenic factor thymidine phosphorylase (TP). Functional experiments demonstrated that inhibition of TP by 5-bromo-6-amino-uracil or gene silencing resulted in a significant increase in basal and oxidative stress-induced apoptosis, whereas supplementation with 2-deoxy-D-ribose-1-phosphate (dRP), the enzymatic product of TP, abrogated this effect. Moreover, dRP produced in EPC cultures stimulated endothelial cell migration in a paracrine manner, as demonstrated by gene-silencing experiments in transmigration and wound repair assays. RGD peptides and inhibitory antibodies to integrin ␣v␤3 attenuated the effect of conditioned medium from EPC cultures on endothelial migration. Finally, the effect of TP on angiogenesis was investigated by implantation of Matrigel plugs in mice. In these in vivo experiments, dRP strongly promoted neovascularization. Our data support the concept that EPCs exert their proangiogenic activity in a paracrine manner and demonstrate a key role of TP activity in their survival and proangiogenic potential. Key Words: angiogenesis Ⅲ endothelium Ⅲ progenitor cells Ⅲ proteomics Ⅲ vascular biology H uman endothelial progenitor cells (EPCs) are attracting considerable attention in cardiovascular research, 1,2 but multiple culture methods from peripheral blood mononuclear cells (PB-MNCs) have been described [3][4][5][6][7] and studied for their clinical relevance. 6,8 -12 EPCs are commonly identified by cell surface antigen expression of CD133, CD34, and the vascular endothelial growth factor receptor-2 (VEGFR-2) (KDR). 13 CD34 and VEGFR-2, however, are also expressed in hematopoietic stem cells 14 ; thus, EPCs cannot yet be unambiguously defined. One alternative approach to flow cytometry has used the colony-forming unit (CFU) assay as a surrogate marker for EPCs. 15 This method has been fundamental to many of the clinical studies published on EPCs to date, which predominantly reported low numbers of CFUs to be correlated to cardiovascular disease risk. Nonetheless, recent publications have cast doubts about the origin of CFUs by demonstrating that they may be clonally derived from the hematopoietic system, possess myeloid progenitor cell activity, and differentiate into phagocytic macrophages. 16 Thus, there is an urgent need to provide a mechanistic underpi...

Cardiogenic shock treated by percutaneous coronary intervention is increasing in incidence and remains a condition associated with high mortality and limited treatment options. Intra-aortic balloon pump therapy was not associated with a long-term survival benefit in this cohort and may be associated with increased early morbidity.

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