CONICET | Buscador de Institutos y Recursos Humanos

Tissue-engineered skin represents a useful strategy for the treatment of deep skin injuries. However, its correct vascularization remains a major challenge. We have shown that the presence of dermal papilla cells (DPCs) in these constructs favors the vascularization process, resulting in a better wound healing and graft take. We also have seen DPC-spheres culture increase the expression of angiogenic genes as VEGF, angiogenin and FGF. Since angiogenesis in scaffolds is essential for grafts to survive and integrate with existing host tissue, our aim was to develop two in vitro models to quantify angiogenesis stimulation of monolayer and spheres cultures of DPC.The first model was a migration assay, in which DPC or dermal fibroblasts (DF) were co-cultured as monolayers with Human Umbilical Vein Endothelial Cells (HUVEC) in a transwell system. We observed more migrated HUVEC in the coculture with DPC than in the DF or medium controls.The second model was an endothelial cell tube formation assay where HUVEC, resuspended in DPC-conditioned medium and seeded on Matrigel, showed an increase in the total tubule?s length and in the number of segments and joints, compared to the medium control.To be able to compare the inductive molecules secretion between monolayer and spheres -DPC cultures, we looked for the condition in which both systems contained the same amount of metabolically active cells. In our culture conditions, 55 spheres were metabolically equivalent to 104 cells/cm2 monolayer seeded cells. Using this equivalence, we are evaluating the effects that spheroid culture conditions have on the angiogenic abilities of DPCs.We conclude that these two angiogenic models are useful to measure, in vitro, the stimulation of angiogenesis of cell cultures to be used as dermal components in skin substitutes. In this way, the use of DPC in skin substitutes could favor the vascularization of the grafts, and thus the closure of the wound and the graft take.