OBJECTIVE Uridine adenosine tetraphosphate(Up4A),a dinucleotide,contains both purine and pyrimidine moieties,and exerts its vascular influence via activation of purinergic receptors.Here,we aimed to investigate the effects of Up4 A on angiogenesis and the putative purinergic receptors(PR)involved in this process.METHODS Tubule formation assay was performed in 3D matrix system.In this assay,human umbilical vein endothelial cells(HUVECs)were co-cultured with pericytes with various Up4 A doses(0,1,2.5,5,10 and 20μmol·L-1)in the absence and presence of P2Y6 R antagonist MRS2578(10μmol·L-1)for 5d.Expression profile of PR subtypes and angiogenic factors was assessed in HUVECs by q-PCR with and without P2Y6 R antagonist.RESULTS No difference in initial tubule formation was detected between Up4 A stimulation and control conditions at day 2.In contrast,a significant increase in vascular density in response to Up4 A was observed at day 5.Up4 A at a dose of 2.5and 5μmol·L-1 promoted total tubule length(by-1.89 fold and-2.23fold),number of tubules(by-1.71 fold and-1.89fold)as well as number of junctions(by-2.24 fold and-2.80fold),all of which were inhibited by MRS2578.Further increase in Up4 A dose to10 and 20μmol·L-1 did not induce an increase in these vascular parameters as compared to non-treated controls.Moreover,Up4 A increased mRNA level of P2YRs(P2Y2R,P2Y4 R and P2Y6R)but not P2XR(P2X4R and P2X7R)or P1R(A2AR and A2BR),while Up4 A upregulated VEGFA and ANGPT1 but not VEGFR2,ANGPT2,Tie1 and Tie2at mRNA level.Transcriptional upregulation of P2 YRs and angiogenic factors by Up4 A was inhibited by MRS2578.CONCLUSION Up4 A is functionally capable of promoting tubule formation in vitro co-culture system.This process is likely mediated by activation of pyrimidine-favored P2 YRs but not P2 XR or P1 Rs,and involves stimulation of well known angiogenic factors. OBJECTIVE Uridine adenosine tetraphosphate (Up4A), a dinucleotide, containing both purine and pyrimidine moieties, and exerts its vascular influence via activation of purinergic receptors. Here, we aimed to investigate the effects of Up4 A on angiogenesis and the putative purinergic receptors (PR) involved in this process. METHODS Tubule formation assay was performed in 3D matrix system. in this assay, human umbilical vein endothelial cells (HUVECs) were co-cultured with pericytes with various Up4 A doses (0,1,2.5,5,10 and 20 μmol·L-1) in the absence and presence of P2Y6 R antagonist MRS2578 (10 μmol·L-1) for 5d. Expression profile of PR subtypes and angiogenic factors was assessed in HUVECs by q-PCR with and without P2Y6 R antagonist. No difference in initial tubule formation was detected between Up4 A stimulation and control conditions at day 2. In contrast, a significant increase in vascular density in response to Up4 A was observed at day 5. Up4 A at a dose of 2.5 and 5 μmol·L -1 Proposed total t ubule length (by-1.89 fold and-2.23 fold), number of tubules (by-1.71 fold and-1.89 fold) as well as number of junctions (by-2.24 fold and-2.80 fold), all of which were inhibited by MRS 2578 .Further increase in Up4 A dose to10 and 20 μmol·L-1 did not induce an increase in these vascular parameters as compared to non-treated controls. Moreover, Up4 A increased mRNA level of P2YRs (P2Y2R, P2Y4 R and P2Y6R) but not P2XR (P2X4R and P2X7R) or P1R (A2AR and A2BR) while Up4 A upregulated VEGFA and ANGPT1 but not VEGFR2, ANGPT2, Tie1 and Tie2 at mRNA level. Transcriptional upregulation of P2 YRs and angiogenic factors by Up4 A was inhibited by MRS 2578. CONCLUSION Up4 A is functionally capable of promoting tubule formation in vitro co-culture system. This process is likely mediated by activation of pyrimidine-favored P2 YRs but not P2 XR or P1 Rs, and involves stimulation of well known angiogenic factors.