Table 1.
Primer sequences and product sizes.
Fig 1.
Wound healing and epithelialization in nondiabetic and diabetic rats in experiment 1.
(a) Macroscopic observation of the healing of full-thickness wounds on the right flanks of nondiabetic (upper panels) and diabetic rats (lower panels) until complete wound closure. Delayed formation of granulation tissue on PWD 4, abundant necrotic tissue on the wound bed on PWD 7 and 10, and delayed epithelialization on PWD 7 were evident in diabetic rats. Scale bar = 1 cm. (b) The wound area measured on PWD 7–9 was significantly greater in diabetic rats (red line) than in nondiabetic rats (blue line). *P < 0.05; **P < 0.01. (c) HE staining showed that the regenerating epidermis was shorter and thicker in diabetic rats than in nondiabetic rats. The histological abnormalities, including invagination of regenerating epidermis into the granulation tissue (arrow), were specific to diabetic rats. Scale bar = 200 μm.
Fig 2.
Immunohistochemistry of the basement membrane components LM5 (a–c), FN (d–f) and Col4 (g–i), and the proliferation marker Ki67 (j–l) in the regenerating epidermis of nondiabetic (a, d, g and j) and diabetic rats (b, e, h and k: flat area; c, f, i and l: invaginating area).
In diabetic rats, immunoreactivity for basement membrane components was decreased compared with that in nondiabetic rats. Structural abnormalities of the basement membrane including fragmentation (arrows) and immaturity (arrowheads) were also observed. Scale bar = 200 μm.
Fig 3.
In vitro analysis of the effects of AHL on keratinocytes.
Foetal rat skin keratinocytes were treated with 10 μM AHL, vehicle only (0.1% DMSO), or were untreated (control). (a) Six hours after treatment, there were no morphological changes in any group. Scale bar = 200 μm. (b) Twenty-four hours after treatment, there were no differences in cellular proliferation among the three groups. (c) Expression analysis of genes related to epidermal basement membrane. (d) Qualitative analysis revealed significant increases in Lm5 expression in AHL-treated cells compared with the vehicle-treated and control groups. Light gray: Control, dark gray: Vehicle, and black: AHL. **P < 0.01.
Fig 4.
Effects of AHL on wound healing and epithelialization in diabetic rats in experiment 2.
(a) Macroscopic observation of healing of full-thickness wounds on both flanks of diabetic rats. The wounds on the right and left flanks were treated with vehicle (upper panels) or AHL (lower panels) on PWD 4. The appearances of both wounds before treatment on PWD 4 were similar. In AHL-treated wounds, by PWD 7, the necrotic tissue had disappeared and regeneration of the epidermis was apparent. Scale bar = 1 cm. (b) The wound area of AHL-treated wounds (red line) on PWD 8, 10 and 12 was significantly smaller than that of vehicle-treated wounds (blue line). *P < 0.05. (c) HE staining showed improvements of the histological abnormalities, including tylosis and invagination (arrow), in AHL-treated wounds but not in vehicle-treated wounds. Scale bar = 200 μm.
Fig 5.
Immunohistochemistry for basement membrane components (a–f) and the proliferation marker Ki67 (g–i) in vehicle-treated (a, c, e and g) and AHL-treated wounds (b, d, f and i).
Scale bar = 200 μm.