Abstract 908: Functional CXCR4 Induction by Hypoxia Promotes Mobilization of Cardiosphere-derived Cells
Introduction: The CXC chemokine SDF-1α and its receptor CXCR4 have been identified as critical mediator for the ischemia-specific recruitment of progenitor cells. Recent studies have identified the resident cardiac progenitor cells (CPCs) in the adult myocardium, yet the level of CXCR4 expression in CPCs is quite low in normal conditions. The ability to functional CXCR4 induction on CPCs has become increasingly important as the role for CPCs in the strategy of stem cell homing continue to expand. In this study, we evaluate whether hypoxia could induce functional CXCR4 expression in CPCs and the molecular mechanisms.
Methods and Results: c-kit(+) or Sca-1(+) CPCs were enriched using the magnetic activated cell sorting (MACS) from mouse cardiosphere-derived cells. To clarify whether hypoxia could induce CXCR4 expression in CPCs, we investigated the CXCR4 expression of CPCs in response to hypoxia, Western blotting revealed a marked up-regulation of CXCR4 protein content following hypoxia induction, and flow cytometry revealed 4-fold more CXCR4 cells than normoxia cultures. We next investigate the transcriptional activity of the CXCR4 promoter under hypoxia by transfecting CPCs with pCXCR4/LacZ vector, in which the LacZ reporter gene was driven by the 279-bp CXCR4 promoter, the transgene expression activation was determined by X-gal staining. We observed the CXCR4 promoter exhibited the enhanced LacZ activity in CPCs treated with hypoxia while very low activity in CPCs with normoxia. Furthermore, siRNA-mediated knock-down of hypoxia-inducible factor-1α (HIF-1α) gene expression in CPCs resulted in significant inhibition of early induction of CXCR4 by hypoxia in the CPCs, indicating that increased levels of HIF-1α under hypoxia may be associated with the high inducibility of CXCR4 promoter. More importantly, in the Transwell system, the hypoxia-treated CPCs had a higher percentage of transmigration toward the CXCR4-specific ligand SDF-1α as compared to the normoxia-treated CPCs (49.68±2.27% vs 7.19%±1.22%, p<0.001), confirming the function of induced CXCR4 receptor.
Conclusions: Hypoxia can induce functional CXCR4 expression in CPCs via direct regulation by HIF-1α. Hypoxia pretreatment may be used as a novel strategy for cardiac stem cell homing.