OBJECTIVE: To review the recent progresses on tissue engineering of skin. METHODS: Recent original articles about tissue engineering of skin were extensively reviewed, which focused on the progresses and major problems concerning the epidermal substitutes, dermal substitutes, cultured-epidermal composite skin graft. RESULTS: Most investigators had come to conclusion that the optimal skin substitute should provide for immediate reconstruction of both the lost epidermis and dermis. The research was mainly focused on how to transplant epidermal cells immediately, preserve their activity and function, and develop the extracellular matrix which could effectively accelerate the function of transplanted cells, induce vascular growth from the wound bed, could be biodegradable, no toxicity and no danger of carrying pathogen. CONCLUSION: The major research trends of tissue engineering of skin should be focused on the study of immediate transplantation of epidermal cells, accelerate wound healing and developing extracellular matrix of dermis.
OBJECTIVE The human epidermal cells were bred on a kind of bio-membrane, the bio-brane, in engineering a kind of new epidermal substitute, the bio-membrane bred cell graft. METHODS Fresh and frozen grafts of biomembrane bred epidermal cells were transplanted into the full-thickness wounds of nude mice and those received simple Bio-brane were served as control. The wounds of the two groups were observed daily and biopsy was taken on the 3, 5, 7, 10, 21 and 35 days respectively. RESULTS Epidermal cells could be cultured in vitro on the bio-membrane reaching the sub-saturated state of 60 to 70 percents. The bio-membrane after being grafted the epidermal cells continued to proliferate and differentiate to form a layer of new epidermis. There was no difference between the fresh and the frozen bio-membranes. CONCLUSION Bio-membrane bred with epidermal cells could be a kind of ideal epidermal substitute.
ObjectiveTo investigate the application of autologous keratinocyte suspension transplantation in skin reconstruction. MethodsForty adult Sprague Dawley rats (weighing, 210-230 g) were randomly divided into 4 groups (n=10): high density keratinocyte suspension transplantation (1×106 cells/cm2) group (group A), middle density (1×105 cells/cm2) group (group B), low density (1×104 cells/cm2) group (group C), and control group (group D). Skin samples were harvested from the rats in groups A, B, C for the isolation of keratinocytes. The model of anti-contracture of full thickness skin wound was made in rats and autologous keratinocyte suspension was transplanted into the wound. The wound was covered by the allogeneic skin (allogeneic skin derived from Wistar rats). The survival of rats was observed after operation. The survival of allogeneic skin was observed at 7, 14, and 21 days after operation, and wound healing rate was calculated after allogeneic skin dropped off. Histological staining and immunohistochemical staining were carried out at 21 days after operation. ResultsAll the rats survived to the end of the experiment. The allograft skins survived in all groups, dried and dropped off. The epithelium sheet could be seen in groups A and B at 21 days, a few very thin epithelium in group C, and no epithelization in group D. The wound healing rate of groups A (62.9%±9.6%) and B (64.2%±9.1%) were significantly higher than that of groups C (38.5%±5.7%) and D (22.7%±5.5%) (P<0.05), and significant difference was found between groups C and D (P<0.05), but there was no significant difference between groups A and B (P>0.05). The results of histological observation showed that squamous epithelial cells were observed in groups A, B, and C, but not in group D; obvious layers of epidermis were observed in groups A and B, thin epidermis and inflammatory cell infiltration in group C, and granulation tissue in group D. The immunohistochemistry staining showed that the expressions of collagen type IV and collagen type VII in groups A, B, and C; the percentage of collagen type IV and collagen type VII positive cells in groups A and B were significantly higher than that of group C (P<0.05), but there was no significant difference between groups A and B (P>0.05). The expressions of collagen type IV and collagen type VII in group D were negative. ConclusionThe repair of full thickness skin wound with graft of autologous keratinocyte suspension can achieve reconstruction of the skin. The appropriate density of keratinocyte suspension for wound healing is 1×105 cells/cm2.
Objective To investigate the possibility of enhancing the inducing rate of adipose-derived stem cells (ASCs) into epidermal cells in the medium containing all-trans retinoic acid (ATRA) by supplementing with HaCaT condition medium. Methods ASCs were isolated and identified by detecting the expression of CD34, CD45, CD73, CD90, and CD105 with flow cytometry and differentiating into adipose and osteoblast lineage in the induction medium. The air-liquid interface cell culture model was established with the Transwell Room. The induction medium A contained ATRA, epidermal growth factor (EGF), and keratinocyte growth factor (KGF), while the induction medium B contained ATRA, EGF, KGF, and HaCaT condition medium. Experiment was divided into three groups cultured for 12 days: induction medium A (group A), induction medium B (group B), basic medium (group C). The epidermal cell surface markers: cytokeratin (CK) 14, 15, 16, 19 (Pan-CK) were detected by flow cytometry and CK14 were identified by immunofluorescence stain. Results After induction for 12 days, flow cytometry showed that the positive rate of Pan-CK in group B [(22.0±3.5)%] was higher than that in group A [(11.9±2.7)%], which were both higher than that in group C [(1.1±0.3)%], and the differences were statistical significantly (P<0.01). Immunofluorescence stain showed that the positive rate of CK14 in group B was higher than that in group A [(19.5±7.0)%vs. (10.8±5.7)%, P<0.01], and the expression of CK14 was negative in group C. Conclusion HaCaT condition medium can enhance the ability of ASCs differentiation into epidermal cells in the culture medium containing ATRA.