Abstract
Non-healing skin ulcers are often resistant to most common therapies. Treatment with growth factors has been demonstrated to improve closure of chronic wounds. Here we investigate whether lyophilized culture supernatant of freshly isolated peripheral blood mononuclear cells (PBMC) is able to enhance wound healing. PBMC from healthy human individuals were prepared and cultured for 24 hours. Supernatants were collected, dialyzed and lyophilized (SECPBMC). Six mm punch biopsy wounds were set on the backs of C57BL/6J-mice and SECPBMC containing emulsion or controls were applied daily for three days. Morphology and neo-angiogenesis were analyzed by H&E-staining and CD31 immuno-staining, respectively. In vitro effects on diverse skin cells were investigated by migration assays, cell cycle analysis, and tube formation assay. Signaling pathways were analyzed by Western blot analysis. Application of SECPBMC on 6 mm punch biopsy wounds significantly enhanced wound closure. H&E staining of the wounds after 6 days revealed that wound healing was more advanced after application of SECPBMC containing emulsion. Furthermore, there was a massive increase in CD31 positive cells, indicating enhanced neo-angiogenesis. In primary human fibroblasts (FB) and keratinocytes (KC) migration but not proliferation was induced. In endothelial cells (EC) SECPBMC induced proliferation and tube-formation in a matrigel-assay. In addition, SECPBMC treatment of skin cells led to the induction of multiple signaling pathways involved in cell migration, proliferation and survival. In summary, we could show that emulsions containing the secretome of PBMC derived from healthy individuals accelerates wound healing in a mouse model and induce wound healing associated mechanisms in human primary skin cells. The formulation and use of such emulsions might therefore represent a possible novel option for the treatment of non-healing skin ulcers.
Highlights
The skin is the largest organ of the human body
supernatants of unstimulated cultured PBMC (SECPBMC) containing emulsions enhance wound healing in mice in vivo To study the capacity of SECPBMC to promote skin wound healing we used the full thickness punch wound model in C57BL/ 6J mice
Microscopic examination revealed that the wound gap distance was smaller in the SECPBMC-treated mice and re-epithelialization was markedly enhanced in these animals (Figure 1C)
Summary
The skin is the largest organ of the human body It covers and protects the underlying organs from ultraviolet radiation, mechanical and chemical damage, invading microorganisms and excessive water loss [1]. Due to this essential function, skin wounds need to be efficiently repaired within a very short time frame. The seminal work by Holzinger et al has shown that autologous transplantation of mononuclear cells effectively initiates and improves granulation and epithelialization of skin ulcers [6]. It has been demonstrated that the efficacy of a stem cell based therapy is dependent on soluble factors produced by these cells, since the secretome of stem cell cultures is sufficient to accelerate cutaneous wound healing [11,12,13]
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