Generating and Reversing Chronic Wounds in Diabetic Mice by Manipulating Wound Redox Parameters

Figure 1

db/db mouse wounds have increased oxidative stress and delayed healing: time course of wound closure in C57BL/6 mice (a) and in db/db mice (b). Wound areas were traced and analyzed using Image J and show delayed closure as compared to C57BL/6. (c) SOD activity was measured using tetrazolium salt that converts into a formazan dye detectable at 450 nm. SOD activity was significantly elevated in the db/db wounds. (d) H₂O₂ measurements were based on the peroxidase-catalyzed oxidation by H₂O₂ and fluorescent product resorufin read fluorometrically at 530 nm/605 nm. H₂O₂ levels were significantly higher in the db/db wounds, confirming the elevated SOD activity in the early hours after wounding. (e) Catalase activity was measured by an enzymatic reaction spectrophotometrically detected with the chromogen purpald at 540 nm and showed reduced activity in the db/db wounds, suggesting a buildup in H₂O₂. (f) GPx activity was measured by a coupled reaction with glutathione reductase where GPx activity was rate limiting and absorbance was read at 340 nm per 1 min intervals. GPx activity showed significantly lower levels at 4 hrs and 48 hrs after wounding. These levels confirm improper detoxification of H₂O₂ leading to redox stress. Time zero represents unwounded skin. . All data are mean ± SD. , , . for each of the studies unless indicated differently.

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