Integration of Blood Outgrowth Endothelial Cells in Dermal Fibroblast Sheets Promotes Full Thickness Wound Healing

Abstract

Vascularization is the cornerstone of wound healing. We introduced human blood outgrowth endothelial cells (hBOEC) in a self-assembled human dermal fibroblast sheet (hDFS), intended as a tissue-engineered dermal substitute with inherent vascular potential. hBOEC were functionally and molecularly different from early endothelial progenitor cells and human umbilical vein endothelial cells (HUVEC). hBOEC alone, unlike HUVEC, efficiently revascularized and re-oxygenated the wound bed, both by active incorporation into new vessels and by trophic stimulation of host angiogenesis in a dose-dependent manner. Furthermore, hBOEC alone, but not HUVEC, accelerated epithelial coverage and matrix organization of the wound bed. In addition, integration of hBOEC in hDFS not only further improved vascularization, epithelial coverage and matrix organization but also prevented excessive wound contraction. In vitro analyses with hBOEC, fibroblasts and keratinocytes revealed that these effects were both due to growth factor crosstalk and to short cutting hypoxia. Among multiple growth factors secreted by hBOEC, placental growth factor mediated at least in part the beneficial effects on keratinocyte migration and proliferation. Overall, this combined tissue engineering approach paves the way for clinical development of a fully autologous vascularized dermal substitute for patients with large skin defects that do not heal properly.

PlGF: A Multitasking Cytokine with Disease-Restricted Activity

“…This finding is corroborated by results that PlGF gene or protein delivery in the skin stimulates angiogenesis, macrophage infiltration, granulation tissue formation, keratinocyte migration and healing of skin wounds in normal mice as well as in diabetic mice, which suffer impaired wound healing (Carmeliet et al 2001;Cianfarani et al 2006;Odorisio et al 2006). Wound vascularization and healing are improved after treatment of skin wounds with dermal fibroblasts containing endothelial progenitor cells, presumably because these cells release PlGF (Hendrickx et al 2010).…”

Neovascularization Potential of Blood Outgrowth Endothelial Cells From Patients With Stable Ischemic Heart Failure Is Preserved

“…Engraftment, studied in 3 versus 3 mice, showed 4±2 incorporated ICMP‐BOECs/4.5 mm 2 versus 3±2 incorporated CON‐BOECs/4.5 mm 2 . This limited incorporation after injection of 250 000 BOECs/mouse suggests a concomitant paracrine contribution of the transplanted cell population to their documented neovascularization potential 7, 8…”

“…In patients with ischemic heart disease, refractory to optimal medical treatment and percutaneous or surgical revascularization, progenitor cell transfer may constitute a promising biological alternative (“biological bypass”) 6. Autologous BOECs are able to induce neovascularization not only by incorporation into functional vessels, but also through paracrine modulation 7, 8…”

Cell-Based Vascularization Strategies for Skin Tissue Engineering

“…31 HUVEC are the most widely used standardized EC line and serve as a reference in many vascularization studies (e.g., refs. 9,32 ). Unfortunately, in the absence of cell banking, these cells can only be used in an allogeneic setting (Table 1).…”

“…6 However, the more layers are combined, the more likely the cell layers by further from the wound surface will die due to hypoxia. Although acute hypoxia initially triggers fibroblast 7 and keratinocyte 8 healing responses, sustained hypoxia leads to fibroblast dysfunction, 7 decreased keratinocyte migration and proliferation, 9 and tissue loss. 10 Hence, a major threat for clinical use of dermal substitutes is insufficient vascularization leading to loosening, infection or partial necrosis of the substitute.…”