This article describes the first (to our knowledge) tetracycline-inducible regulatory system that demonstrates that the tetracycline repressor (tetR) alone, rather than tetR-mammalian cell transcription factor fusion derivatives, can function as a potent trans-modulator to regulate gene expression in mammalian cells. With proper positioning of tetracycline operators downstream of the TATA element and of human epidermal growth factor (hEGF) as a reporter, we show that gene expression from the tetracycline operator-bearing hCMV major immediate-early enhancer-promoter (pcmvtetO) can be regulated by tetR over three orders of magnitude in response to tetracycline when (1) the reporter was cotransfected with tetR-expressing plasmid in transient expression assays, and (2) the reporter unit was stably integrated into the chromosome of a tetR-expressing cell line. This level of tetR-mediated inducible gene regulation is significantly higher than that of other repression-based mammalian cell transcription switch systems. In an in vivo porcine wound model, close to 60-fold tetR-mediated regulatory effects were detected and it was reversed when tetracycline was administered. Collectively, this study provides a direct implementation of this tetracycline-inducible regulatory switch for controlling gene expression in vitro, in vivo, and in gene therapy.

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Diabetic wounds result in significant morbidity, prolonged hospitalization, and enormous health-care expenses. Pigs have been shown to have wound healing resembling that in humans. The aim of this study was to develop a large-animal model for diabetic wound healing. Diabetes was induced by streptozotocin injection in Yorkshire pigs. Full-thickness wounds were created and dressed with a sealed chamber. Nondiabetic pigs with or without high glucose wound fluid concentration served as controls. Glucose concentration in serum and wound fluid was measured and collected. Wound contraction was monitored, and biopsies were obtained for measurement of reepithelialization. Wound fluid was analyzed for insulin-like growth factor-1 (IGF-1), platelet-derived growth factor, and transforming growth factor. Glucose concentration in wound fluid initially followed serum levels and then decreased to undetectable on day 9. Reepithelialization was significantly delayed in diabetic pigs. In nondiabetic pigs, wounds treated in a local hyperglycemic environment, and thus excluding the effects of systemic hyperglycemia, showed no difference in wound closure compared with controls. This suggests that delayed wound healing in diabetes is not induced by local highglucose concentration itself. Analysis of growth factor expression showed a marked reduction in IGF-1 in the diabetic wounds. Diabetic pigs have impaired healing that is accompanied by a reduction of IGF-1 in the healing wound and is not due to the local hyperglycemia condition itself.Approximately 5 million patients in the United States suffer from chronic wounds. 1 With the increased longevity, obesity, and diabetes, the problem of chronic wounds has increased, resulting in significant morbidity, lost time from work, and enormous health-care expenses. According to the American Diabetes Association, 25% of people with diabetes will suffer from a wound problem during their lifetime, and approximately 82,000 limb amputations for nontraumatic wounds were performed in people with diabetes in 2002. 2 The Agency for Health Care Policy and Research reports that wound care for pressure ulcers uses $200 billion a year for hospitalization, durable medical goods, nursing home care, physicians, and transportation. 3 Surgical treatment of diabetic wounds remains difficult and often insufficient, leading to high morbidity among those patients. 4 We need better ways to treat diabetic wounds and relevant preclinical models are needed to develop new therapeutic strategies.Numerous diabetic wound healing models have been described. 5,6 Small mammals, such as rats, rabbits, guinea pigs, and mice, are frequently used in wound healing studies because of cost and ease of handling. However, the anatomy and physiology of small mammals differ from those of humans in many ways. 7 Pig physiology and wound healing has been found to be significantly more similar to humans. 8,9 In wound healing models investigating basic fibroblast growth factor (bFGF), wounds in genetically diabetic db/db mice treate...

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