Use of cultured human epithelium for coverage : a defect of radial forearm free flap donor site

The radial forearm free flap has been popular in many areas of reconstructive surgery. Despite the many attributes of this flap in maxillofacial reconstruction, one of the disadvantages has been the morbidity of the donor site. Allogeneic cultured epidermis has been successfully applied on large second degree burns and on chronic leg ulcers. Autologous human keratinocytes and fibroblast equivalents can be cultured in-vitro from a small skin sample in order to produce a sufficient amount of epithelial autografts to cover the large defects of third-degree burn wounds. Interestingly, transplanted cultured epidermis retains characteristics of the original donor site. We report a case of a patient who underwent skin replacement by cultured epithelial autograft after wound breakdown occurred in the forearm donor site during the early postoperative period. This method could represent an auspicious alternative to conventional grafting methods for forearm free flap reconstruction. To the best of our knowledge, skin replacement by cultured epithelial autografts in this region has not been extensively described in the literature

Cooperation by fibroblasts and bone marrow-mesenchymal stem cells to improve pancreatic rat-to-mouse islet xenotransplantation.

Experimental and clinical experiences highlight the need to review some aspects of islet transplantation, especially with regard to site of grafting and control of the immune response. The subcutaneous space could be a good alternative to liver but its sparse vasculature is its main limitation. Induction of graft tolerance by using cells with immunoregulatory properties is a promising approach to avoid graft rejection. Both Fibroblasts and Mesenchymal Stem Cells (MSCs) have shown pro-angiogenic and immunomodulatory properties. Transplantation of islets into the subcutaneous space using plasma as scaffold and supplemented with fibroblasts and/or Bone Marrow-MSCs could be a promising strategy to achieve a functional extra-hepatic islet graft, without using immunosuppressive drugs. Xenogenic rat islets, autologous fibroblasts and/or allogenic BM-MSCs, were mixed with plasma, and coagulation was induced to constitute a Plasma-based Scaffold containing Islets (PSI), which was transplanted subcutaneously both in immunodeficient and immunocompetent diabetic mice. In immunodeficient diabetic mice, PSI itself allowed hyperglycemia reversion temporarily, but the presence of pro-angiogenic cells (fibroblasts or BM-MSCs) within PSI was necessary to improve graft re-vascularization and, thus, consistently maintain normoglycemia. In immunocompetent diabetic mice, only PSI containing BM-MSCs, but not those containing fibroblasts, normalized glycemia lasting up to one week after transplantation. Interestingly, when PSI contained both fibroblasts and BM-MSCs, the normoglycemia period showed an increase of 4-times with a physiological-like response in functional tests. Histology of immunocompetent mice showed an attenuation of the immune response in those grafts with BM-MSCs, which was improved by co-transplantation with fibroblasts, since they increased BM-MSC survival. In summary, fibroblasts and BM-MSCs showed similar pro-angiogenic properties in this model of islet xenotransplantation, whereas only BM-MSCs exerted an immunomodulatory effect, which was improved by the presence of fibroblasts. These results suggest that cooperation of different cell types with islets will be required to achieve a long-term functional graft

Evolution of bioluminescent cells embedded in PSI and subcutaneously transplanted in both immunocompetent and immunodeficient mice.

<p>A) <i>Immunocompetent mice</i>. Relative luminescence signal of 10⁶ allogenic Luc[+]-BM-MSCs transplanted alone into PSI (PSI-5M_luc) or co-transplanted with 10⁶ autologous Luc[-]-fibroblasts (PSI-5FM_luc), as well as representative IVIS® lumina images of previously described mice, 12 days after transplantation. B) <i>Immunodeficient mice</i>. Relative luminescence signal of 10⁶ allogenic Luc[+]-BM-MSCs transplanted alone into PSI (PSI-M_luc) or co-transplanted with 10⁶ autologous Luc[-]-fibroblasts (PSI-FM_luc), as well as luminescence signal of 10⁶ allogenic Luc[+]-fibroblasts transplanted alone into PSI (PSI-F_luc). Representative IVIS® lumina images of previously described mice, 12 days after transplantation. Values related to day 0 (100%) are represented as mean ± SEM. For all groups n=5. (*) p<0.05; (**) p<0.01; (***) p<0.001.</p

Histology of subcutaneous islet xenografts in immunocompetent diabetic mice.

<p>A) Representative images of subcutaneously transplanted islets alone (ISC), PSI without cells and PSI containing: 10⁶ autologous fibroblasts (PSI-5F), 10⁶ allogenic BM-MSCs (PSI-5M), or 10⁶ of both autologous fibroblasts and allogenic BM-MSCs (PSI-5FM). Islet grafts were retrieved seven days after transplantation in diabetic mice. Samples were stained with H and E, or labelled with anti-insulin or anti-MPO antibodies. Black arrows point to islets. B) Quantification of the number of MPO-positive cells per field present on the leukocyte infiltration of the islet graft. C) Quantification of the insulin-positive area per section present on the islet graft. (**) p<0.01; (***) p<0.001.</p

Assessment of Optimal Virus-Mediated Growth Factor Gene Delivery for Human Cutaneous Wound Healing Enhancement

Using a recently described skin-humanized model based on the engraftment of human bioengineered skin equivalents onto immunodeficient mice, we compared the efficacy of different in vivo gene transfer strategies aimed at delivering growth factors to promote skin wound healing. The approaches involving transient delivery of keratinocyte growth factor (KGF) to wounds performed in the engrafted human skin included (1) KGF gene transfer by intradermal adenoviral injection; (2) KGF gene transfer by adenoviral vector immobilized in a fibrin carrier; and (3) KGF-adenoviral gene-transferred human fibroblasts embedded in a fibrin matrix. All delivery systems achieved KGF protein overproduction at the wound site, with a concomitant re-epithelialization enhancement. However, although direct gene delivery strategies exhibited variability in terms of the number of successfully transduced humanized mice, the use of genetically modified fibroblast-containing matrix as an in situ protein bioreactor was highly reproducible, leading to a significant improvement of the overall healing process. This latter approach appeared to be the most reliable means to deliver growth factors to wounds and also avoided the potential danger of scoring cases of faulty administration as therapeutic failures and direct exposure to viral vectors. The combined use of cell and gene therapy appears a robust tool to aid healing in a clinical context