Endothelial Progenitor Cells(EPC) in Peripheral blood in Diabetic Foot with and without PAD.

Abstract

5130 EPC is not very well defined population. These cells have either angiopoietic or angiostimulating function, the latter seems to be more relevant. It is well known that Mobilization of EPC to peripheral blood is decreased in diabetic rats. From the opposite, ischemia proved to stimulate mobilization. That is why, we decided to study EPC in diabetic patients with and without PAD Patients and methods 40 pts with diabetes(type I- 4, type II-36) were studied. Duration of diabetes -1-46 years. The duration of more than 10 years was in 57% patients. Age- 49-73 years(mean age- 62 years). The patients were divided into two groups- diabetic foot without PAD(25 pts) – Group A, and diabetic foot with PAD- group B. PAD was diagnosed with sonography, duplex sonography and Rx arteriography. Normal volunteers were use as controls(mean age 51.6+2.0) CD34+ and CD133+ cells were numerated with CytoFlow(BD). 5 Day CFU-Hill Colony Assay- non-adherent mononuclear cells (MNCs) was used to study EPC In this method, peripheral blood MNCs are plated on fibronectin-coated dishes (6-well). After a 48 h pre-plating step to deplete the sample of adherent macrophages and mature endothelial cells, the non-adherent cells are removed and re-plated on fibronectin-coated dishes (24-well). Unique colonies that are formed in the 5 Day CFU-Hill Colony Assay are referred to as colony-forming unit-Hill colonies (CFU-Hill colonies) or colony-forming unit-ECs (CFU-ECs). Count the number of colonies per well for each sample. CFU-Hill colonies are defined as a central core of round cells with radiating elongated spindle-like cells at the periphery. Colonies without the CFU-Hill morphology may also be present but are not scored as CFU-Hill colonies. CFU-Hill Colonies are fixed with methanol and stained with a Giemsa solution. Plasma concentration of VEGF was studied by ELISA Results EPC colony forming ability in group A was 4.0+ 0.7, in group B-27.4+3.9. The differences between these two groups is highly significant(p 0.001). Low EPC level in diabetic neuropathy is probably related to decreased mobilization(G.Fadini e.a.2006). Rather unusual was higher EPC level in group B. It was shown that age-related decrease in EPC is due to decline in HIF-1 signaling(M. Hoenig e.a 2008). Probably, diabetic defect of mobilization is partly overcome with ischemia induced up-regulation of HIF-1 and VEGF. EPC in group B patients does not differ from controls(26.1+6.5). There were now correlations between EPC number and the number of either CD34+, CD133+, or KDR+ cells. We also failed to find any correlations between VEGF and EPC Conlusion EPC number in peripheral blood in diabetic patients without PAD is severely decreased. Diabetic patients with PAD have near normal EPC number. Thus, ischemia in diabetic patients is unable adequately mobilize EPC Disclosures No relevant conflicts of interest to declare.