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| Priming of mobPBSCs with Angiopoietin-1 enhances engraftment and new vessel formation in the ischemic tissue via Tie2/Ets-1 pathway. |
| 서울대학교 병원 임상의학연구소 심혈관 연구실¹ , 카이스트 생명공학부² , 서울대학교 의과대학 내과학교실³ |
| 이춘수¹, 김민석³ , 허진¹ , 박경우³ , 조현재³ , 이해영³ , 강현재³ , 고규영² , 김효수³ , 오병희³ , 박영배³ |
Background: Limitation of intracoronary infusion of stem cells is the insufficient rate to ischemic tissue (less than 5%). Accordingly, we aimed to find a new method of augmenting neovascularization by overcoming the poor engraftment of peripheral blood stem cells enriched by G-CSF mobilization and apheresis (mobPBSCs) into ischemic tissue and by enhancing its endothelial lineage commitment. Methods and Results: Ang-1 was administered as a priming agent because it was found that mobPBSCs higher express Tie2, Ang-1 receptor than peripheral blood mononuclear cells. When treated with Ang-1, mobPBSCs formed more endothelial lineage cell clusters (EPC clusters). So, we investigated whether the effect of Ang-1 on mobPBSCs is mediated by Ets-1 transcription factor. The gene expression of Ets-1 was increased in a time dependent manner after Ang-1 stimulation in mobPBSCs. Compared with vehicle, treatment with Ang-1 for 4 hours resulted in stronger expression of endothelial markers, such as, CD31 and VE-cadherin, which were significantly reduced with antisense Ets-1 oligodeoxynucleotides (ODN). And, priming of mobPBSCs with Ang-1 increased α4β1 and α5β1 integrins. To correlate these findings with functional changes in mobPBSCs, we performed in vitro adhesion assays. Ang-1 stimulation increased mobPBSC adhesion to HUVECs and fibronectin, while blocking with anti-hTie2 or anti-integrin antibody prior to Ang-1 stimulation abrogated these results. Finally, to investigate the enhancement of in vivo engraftment of mobPBSCs to ischemic target tissue, we developed a novel animal model using radioisotope-tagged mobPBSCs. In the rabbit ear ischemia-reperfusion model, priming of mobPBSCs with Ang-1 resulted in the significant increase of the first pass engraftment to distal vascular bed after intra-arterial delivery. We also confirmed the therapeutic efficacy of primed mobPBSCs to enhance homing and neovascularization in athymic nude mice ischemic hindlimbs. Conclusion: These results suggest that priming mobPBSCs with Ang-1 may enhance the efficacy of transplanted mobPBSCs in terms of increasing neovascularization in ischemic tissue.
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