[Repair of swine full-thickness cutaneous deficiency by autogenic BMSCs compounded with collagen membrane].

OBJECTIVE:To supply references to tissue-engineered skin clinical applications with autogenic BMSCs composited collagen membrane to repair swine full-thickness cutaneous deficiency. METHODS:Twenty mL bone marrow were obtained respectively from 4 swine, autogenic BMSCs were cultured and passed to the 3rd passage. The fresh bovine tendon treated by means of chemically cross-linked was made 5 cm diameter collagen I (Col I) membrane. The 2 x 10(7)/mL P3 swine autogenic BMSCs labeled DAPI were planted to sterile Col I membrane for 24 hours incubation, then the tissue-engineered skin was constructed. The five full-thickness skin defect of 5 cm diameter was excised to the muscle from forward to backward on the back midline two sides of swine. The tissue-engineered skin were implanted in the experimental group, while Col I membrane was implanted in control group. After 3 and 8 weeks of implantation, the two swine wound surface healing circumstance was observed and further evaluated with histology analysis and TEM. After 3 weeks of implantation, the experimental group were observed with fluorescence microscopy and staining for glycogen. RESULTS:After 3 weeks of implantation, the wound surface of control group were observed nigrescene, scab and putrescence, and after 8 weeks of implantation, also evident putrescence and scar. The wound surface of experiment group was alive after 3 weeks implantation, appearance was leveled off and flexible without evident scar. The wound surface recovered well after 8 weeks of implantation, wound surface healing rate was significantly difference between the two groups (P < 0.01). After 3 weeks of implantation, control group were observed acestoma hyperplasia and no epidermal coverage by histology analysis. The experimental group was showed integrity epidermis and dermis structure. The basal layer was crimson and continuously positive with glycogen staining. After 8 weeks of implantation, the experimental group and control group were emerged normal skin structure. After 3 weeks of implantation in control group, a lot of neutrophilic granulocytes and fibroblasts were noticed, but no epidermal structure was observed under TEM. In the experimental group, a lot of epidermal cells were observed, dermatome connection among epidermal cells and hemidesmosome connection between basilar membrane cells and basal membrane were observed in epidermis. In the dermis experimental group, blood capillary endothelial cells were noticed. Furthermore, considerable collagen fiber deposit was found in the surrounding tissue of fibroblasts. After 3 weeks of implantation, BMSCs labeled with DAPI were located reconstructed epidermal basement membrane and dermis by fluorescence microscopy. CONCLUSION:Tissue-engineered skin which is composited with autogenic BMSCs as seed cells and collagen membrane were potential prospects in application of repairing swine full-thickness cutaneous deficiency.