ATP promotes resident CD34⁺ cell migration mainly through P2Y2-Stim1-ERK/p38 pathway

Extracellular adenosine triphosphate (ATP) is one of the most abundant biochemical constitutes within the stem cell microenvironment and is postulated to play critical roles in cell migration. However, it is unclear whether ATP regulates the cell migration of CD34⁺ vascular wall-resident stem/progenitor cells (VW-SCs) and participates in angiogenesis. Therefore, the biological mechanisms of cell migration mediated by ATP was determined by in vivo subcutaneous matrigel plug assay, ex vivo aortic ring assay, in vitro transwell migration assay, and other molecular methods. In the present study, ATP dose-dependently promoted CD34⁺ VW-SCs migration, which was more obviously attenuated by inhibiting or knocking down P2Y2 than P2Y6. Furthermore, it was confirmed that ATP potently promoted the migration of resident CD34⁺ cells from cultured aortic artery rings and differentiation into endothelial cells in matrigel plugs by using inducible lineage tracing Cd34-CreER^T2; R26-tdTomato mice, whereas P2Y2 and P2Y6 blocker greatly inhibited the effect of ATP. In addition, ATP enhanced the protein expression of stromal interaction molecule 1 (STIM1) on cell membrane, blocking the calcium release-activated calcium (CRAC) channel with shSTIM1 or BTP2 apparently inhibited ATP-evoked intracellular Ca²⁺ elevation and channel opening, thereby suppressing ATP-driven cell migration. Moreover, extracellular signal-regulated protein kinase (ERK) inhibitor PD98059 and p38 inhibitor SB203580 remarkably inhibited ERK and p38 phosphorylation, cytoskeleton rearrangement, and subsequent cell migration. Unexpectedly, it was found that knocking down STIM1 greatly inhibited ATP-triggered ERK/p38 activation. Taken together, it was suggested that P2Y2 signaled through the CRAC channel mediated Ca²⁺ influx and ERK/p38 pathway to reorganize the cytoskeleton and promoted the migration of CD34⁺ VW-SCs.NEW & NOTEWORTHY In this study, we observed that the purinergic receptor P2Y2 is critical in the regulation of vascular wall-resident CD34⁺ cells' migration. ATP could activate STIM1-mediated extracellular Ca²⁺ entry by triggering STIM1 translocation to the plasma membrane, and knockdown of STIM1 prevented ERK/p38 activation-mediated cytoskeleton rearrangement and cell migration.