Participação das selectinas nas interações entre células angiogênicas da medula óssea e endotélio de músculos isquêmicos de membros inferiores em modelo murino.

Abstract

The stimulation of vascular growth by angiogenic stem cells transplantation is a promising alternative for the treatment of PAD. Adhesive mechanisms governing progenitor/stem cell trafficking through the microcirculation of ischemic muscles are broadly unknown. The aim of this work was to evaluate the involvement of selectins on interactions between transplanted bone marrow angiogenic cells and the endothelium of ischemic hindlimbs in mice. Hindlimb ischemia was induced by permanent occlusion of the femoral artery of C57Bl/6 mice. The method of sodic fluorescein diffusion into the bloodstream after plantar cushion injection and the laser Doppler perfusion imaging (LDPI) were used for evidence of ischemia. Bone marrow-derived mononuclear cells (BM-MNC) or hematopoietic lineage negative cells (BM-Lin-) were obtained from GFP transgenic mice and i.v. injected 24h post-ischemia and just before intravital microscopy (IVFM) recording. By using IVM, we observed that the number of circulating cells rolling and adherent to the endothelium of the ischemic muscle was higher at days 1 and 3 post-ischemia when compared with sham-operated animals. After transplantation of the bone marrow-derived angiogenic stem/progenitor cells, we observed a greater number of rolling and adherent BM-MNC and BM-Lin- when compared with sham animals. These events were significantly reduced after i.v. injection of fucoidan, a pan-selectin inhibitor, anti-E-selectin or anti-P-selectin blocking antibodies (10 min before transplantation for all of them) or after platelet depletion, when compared with untreated ischemic animals. Interestingly, after P-selectin blockade, we observed a greater reduction in rolling when compared to E-selectin blockade. Nevertheless, we did not observe any adhesion of transplanted BM-MNC after P-selectin blockade. Finally, the blockade of host selectins or the depletion of platelets before transplantation reduced the reparative revascularization induced by BM-MNC or BM-Lin- as analyzed by histomorphometry, the fluorescein method and/or LDPI. Our data suggest that the blockade of host selectins or the platelet depletion reduces the interactions between transplanted bone marrow angiogenic cells and the endothelium and their recruitment to ischemic hindlimbs. Our data also uncover a major role for P-selectin in these events