A Chemokine Receptor CXCR4 Plays a Crucial Role for Repairing Ischemic Tissue through Regulation of HIF-2a

Endothelial progenitor cells (EPCs) are potential therapeutic tools for treatment of ischemic tissues. We previously divided EPCs into two populations on the basis of aldehyde dehydrogenase (ALDH) activities and demonstrated that EPCs with low ALDH activity (Alde-Low EPCs) possess a greater ability for repairing ischemic tissue than those with high ALDH activity by using mouse dorsal skin flap model (Nagano et al., Blood, 2007). To explore molecular basis of the function of Alde-Low EPCs, we transduced small interfering RNAs (siRNAs) into Alde-Low EPCs to reduce the expression of hypoxia inducible transcription factors (HIF-1a or HIF-2a) and examined the ability to repair ischemic tissues by using mouse skin flap model. Injections of Alde-Low EPCs or EPCs with HIF-1a siRNAs via tail vein significantly reduced the necrotic area (4.4±4.5 and 13.2±2.8% of total ischemic area, respectively, n=3 in each group) compared to control PBS injection (30.5±3.2%, n=4). In contrast, EPCs with HIF-2a siRNA showed no positive effect on reducing necrotic area (34.1± 5.2%, n=4). These data suggest that HIF- 2a plays a major role for repairing ischemic tissues in Alde-Low EPCs. We then analyzed the expression of hypoxia-inducible genes in these cells. While the VEGF mRNA level was 30% reduced in both HIF-1a and HIF-2a siRNA treated cells, the expression of a chemokine receptor CXCR4 was reduced in HIF-2a, but not HIF-1a siRNA transduced cells. Moreover, transduction of CXCR4 siRNA into Alde-Low EPCs significantly lost the ability to repair ischemic regions in a mouse flap model (35.5± 3.5%, n=3). Collectively, these results indicate that CXCR4 plays an essential role for repairing ischemic tissue through regulation of HIF-2a.