Objective The immunogenicity of tissue engineered skins is still vague, though it has been appl ied cl inically for several years. To observe the evidence of immunologic rejection of tissue engineered skins transplanted to severe combined immunodeficiency (SCID) mice, which are implanted human splenic lymphocytes to construct human immunesystem. Methods Tissue engineered skins and acellular dermic matrix were constructed in vitro. Twenty SCID mice, aging4-6 weeks and weighing 16-17 g, were randomly divided into four groups equally (n=5). The tissue engineered skins, human foreskins from circumcision and acellular dermic matrix were transplanted to groups A, B, and C, respectively; group D was used as a control. After 2 weeks of transplanting, 3 × 107 human splenic lymphocytes were injected into every SCID mouse intraperitoneally. After 4 weeks, the morphology, histology, immunohistochemistry and human IgG immunofluorescence were used to observe immunologic rejection. Results Group A showed that transplanted tissue engineered skins had the bilayer structure of dermis and epidermis, which was similar to the normal human skin structure. Group B showed that the transplanted human foreskins still retained normal structure of human skin. Group C showed that acellular dermic matrix were located in situ and had no sign of degradation. After injecting human splenic lymphocytes into the SCID mice, no inflammatory cells infil itration were observed basically in groups A, C, and D; the inflammatory cells infil itration of group B were significantly higher than that of other 3 groups (P lt; 0.05). The results of anti human keratin 14 monoclonal antibody (mAb) staining and anti human type IV collagen mAb staining were positive in group A; no positive cells for CD3, CD4, and CD8 were observed in groups A, C, and D; and many positive cells for CD3, CD4, and CD8 were observed in group B. The results of IgG immunofluorescence staining was negative in group A, C, and D, and positive in the great vessel wells of group B. Conclusion The immunogenicity of tissue engineered skins is very weak, and tissue engineered skins would not be rejected by host immune system after transplantation.
To investigate the effect of hepatocyte growth factor (HGF) on prol iferation of cultured human eccrine sweat gland epithel ial cells (hESGc) and the involvement of phosphorylation of ERK1/2. Methods hESGc were cultured in keratinocyte serum free medium (KSFM) and the first generation of hESGc was harvested. The expression of C-met was detected by immunocytochemistry. MTT assay was used to detect the effect of HGF on the prol iferation of hESGc. The cells were divided into blank group, control group and experimental group. The culture density was 2 × 103 cells/hole in control group and experimental group. Two hundred μL KSFM with HGF in different levels was added to every hole. hESGcwere cultured in KSFM with HGF at different levels (2, 20, 40 and 80 ng/mL) in experimental group, in KSFM without HGF incontrol group, and in KSFM without HGF and no hESGc in blank group. The cell prol iferation was observed in xperimental group 2 and 4 days later. Western blot was used to detect the expression of phosphorylated ERK1/2 at 40 ng/mL HGF after 0, 5, 30, 90 and 120 minutes. Results The results were positive for anti-C-met staining in the cytoplasm. HGF (40 ng/mL and 80 ng/mL) significantly improved the prol iferation of hESGc (P lt; 0.05). When cultured in the KSFM with 40 ng/mL HGF, the cell prol iferation rate and the absorbance were 74.2%, 0.239 3 ± 0.070 9 at 2 days and 74.8%, 0.287 8 ± 0.074 3 at 4 days; showing significant differences when compared with control group (P lt; 0.05). When cultured in KSFM with 80 ng/mL HGF, the cell prol iferation rate and the absorbance were 54.5%, 0.212 3 ± 0.059 2 at 2 days and 40.3%, 0.231 0 ± 0.056 7 at 4 days; showing significant differences when compared with control group (P lt; 0.05). The expression of p-ERK1/2 reached to the maximum after stimulation of 40 ng/mL HGF for 5 minutes, and relative integral absorbance (RIA) was 0.593 2 ± 0.192 2, increased 8.1 times compared with instant stimulation (P lt; 0.01). Conclusion HGF could induce the prol iferation of hESGc and activate the phosphorylation of ERK1/2 protein.