[Explorative study of the immobilizing effect of full-thickness skin subcutaneous grafting on allogeneic full-thickness skin graft in rats].

Objective: To investigate the immobilizing effect of full-thickness skin subcutaneous grafting on allogeneic full-thickness skin graft in rats. Methods: The experimental research method was used. The inbred male Brown-Norway rats (n=10) and Lewis rats (n=10) were used as donors and recipients respectively. After subcutaneously full-thickness separation of a 2.2 cm×2.2 cm area on the nape of the recipient rat, a full-thickness skin of 2.0 cm×2.0 cm taken from the abdomen of the donor rat was subcutaneously grafted, and the donor site was pulled together and sutured. The autologous skin over the allograft in the recipient rat was excised 5-6 d after grafting, and the stitches were removed 7 d after excision. Within 2 months after grafting, the feeding, activity, and survival of the donor and recipient rats, behavior of tearing and scratching the wounds of the recipient rats, the wound condition after autologous skin excision in recipient rats, and the survival and hair growth of the grafted allogeneic skin were observed. Results: Within 2 months after grafting, the donor and recipient rats all ate normally and could move freely with no abnormal death. No tearing or scratching of the wounds occurred in recipient rats. There was a small amount of exudation and partial epidermal desquamation after autologous skin excision in recipient rats. All transplanted allografts survived, which were free of infection and necrosis, with new hairs growing out smoothly. Conclusions: The immobilizing method of full-thickness skin subcutaneous grafting of allogeneic full-thickness skin graft in rats is simple and time-saving without postoperative dressing change, with reliable pressure fixation and high survival rate of skin grafts, which can be promoted for animal skin grafting models.

Use of genetically modified allograft to deliver local immunomodulatory molecule with minimal systemic toxicity in a rat model of allogeneic skin flap transplantation.

The effects of OX40-OX40 ligand (OX40L) costimulatory pathway blockade to prevent T-cell-mediated acute rejection were investigated in a rat model of allogeneic superficial inferior epigastric artery flap transplantation. An ex vivo gene transfer technique was used to modify allografts to locally deliver an immunomodulatory molecule. The flaps were separated from donors, perfused with an adenoviral vector expressing the OX40 immunoglobulin (AdOX40Ig) for 1 hour, and incubated at 37°C for 2 hours. Before transplantation, the flaps were flushed with phosphate-buffered saline solution to remove unincorporated viral particles. Recipients were randomly divided into 5 groups, and treated with topical OX40Ig gene transfer, a single low dose of rapamycin alone, or a combination of agents. Graft survival was assessed using histopathologic classification of skin rejection. All animals in the untreated group (n = 9) or the group treated with adenovirus expressing green fluorescence protein (n = 9) developed grade 3 clinical rejection by postoperative day 7. No significant difference was observed in graft survival between the locally treated AdOX40Ig groups (mean [SD], 8.1 [0.7] days) and the untreated groups (7.7 [1.2] days) could be observed (P > .05, t test). Graft survival in the locally treated AdOX40Ig groups was extended to 18.7 (1.2) days when transduction was combined with a low dose of rapamycin, a significant improvement over survival with rapamycin treatment alone (13.2 [0.6] days) (P < .01). These results demonstrated that local immunomodulation by the allograft itself and low-dose rapamycin treatment promote graft acceptance. This protocol may enable reduction of the dosage of immunosuppressive drugs needed for successful inhibition of acute rejection in the early postoperative period.

Troglitazone suppresses transforming growth factor-beta1-induced collagen type I expression in keloid fibroblasts.

BACKGROUND: Peroxisome proliferator-activated receptor (PPAR)-gamma agonists are increasingly used in patients with diabetes and some studies have suggested a beneficial effect on organ fibrosis. However their effects on dermal fibrosis in keloids are unknown. OBJECTIVE: To investigate the effect of the PPAR-gamma agonist troglitazone on transforming growth factor (TGF)-beta1-induced collagen type I expression in keloid fibroblasts. METHODS: Keloid fibroblasts were cultured and exposed to different concentrations of troglitazone in the presence of TGF-beta1. The mRNA expression of PPAR-gamma was determined by semiquantitative reverse transcriptase-polymerase chain reaction. The protein of PPAR-gamma, Smad2, Smad3, phoshpo-Smad2/3 and collagen type I was determined by Western blotting and collagen synthesis was evaluated by measuring (3)H-proline incorporation. The effect of troglitazone on cell viability was evaluated by the colorimetric conversion of 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide. RESULTS: PPAR-gamma was expressed at a moderate level in keloid fibroblasts. Troglitazone depressed TGF-beta1-stimulated collagen type I expression and collagen synthesis in keloid fibroblasts in a concentration-dependent manner. Moreover, troglitazone inhibited expression and phosphorylation of TGF-beta1-induced Smad2/3. Cell viability was unaffected. These inhibitory effects of troglitazone were reversed by the PPAR-gamma-specific antagonist GW9662. CONCLUSIONS: Our data suggest that PPAR-gamma is present in keloid fibroblasts and PPAR-gamma activation inhibits TGF-beta1-induced collagen type I expression at least in part by decreasing collagen synthesis. PPAR-gamma may be a promising therapeutic target for keloids.

VEGF gene therapy for the survival of transplanted fat tissue in nude mice.

The objective of this study was to determine the effects of adenovirus-mediated vascular endothelial growth factor (Ad-VEGF) on the angiogenesis and survival of free-fat tissue transplantation in nude mice. Thirty 6-week-old CD-1 nude male mice were injected with 1ml fat tissue (harvested by suction-assisted lipectomy from the breast of humans) in the subcutaneous of scalp and were randomised into three groups of 10 animals each. Group 1 was the study group, in which Ad-VEGF was mixed with transplanted fat tissue and injected into mice. In group 2, adenovirus-mediated green fluorescent protein (Ad-GFP) gene was mixed with transplanted fat tissue and injected into the mice. In group 3, normal saline alone was used. Both group 2 and group 3 are control groups. The animals were euthanised 15 weeks after the procedure. The fat survival weight and volume of the study group were significantly greater than those of two control groups (p<0.05). Light microscopical examination of haematoxylin and eosin-stained slides of the dissected fat 15 weeks after injection was performed in group 1 and group 2. Less cyst formation and fibrosis, indicating improved quality of the injected fat, can be obtained by the addition of Ad-VEGF. Vascular density was evaluated at the microvascular level through the use of light microscopic sections of the central part of the fat tissue at 15 weeks after injection by von Willebrand factor staining. Histological evaluation showed that capillary density increased markedly in the study group mice. Mice of the study group disclosed significantly higher VEGF protein levels detected by ELISA assay of plasma samples obtained from the mice after the fat injection (day 1, 4, 7 and 28; p<0.01) at each time point than the mice of the two control groups. The findings reported in this study indicate that the VEGF gene therapy can enhance the survival and the quality of grafted fat tissue, which may be due to induction of angiogenesis.