Imbalance in the production of reactive oxygen species predisposes diabetic patients to delayed wound healing, but treatments that directly address this imbalance are lacking. Nrf2 and its natural inhibitor, Keap1, maintain redox homeostasis. However, the exact contribution Nrf2 makes to healing is unclear. Here we demonstrate that Nrf2 loss of function in discreet layers of the epidermis corresponds with substantial oxidative stress burdens and a concomitant delay in healing human tissue. Compared to nondiabetic human epidermal keratinocytes (NHEKs), diabetic human epidermal keratinocytes (DHEKs) exhibit a pro-inflammatory cytokine profile and decreased transcription of downstream antioxidant genes. Silencing Keap1 in DHEKs led to restoration of a cytokine profile conducive to healing (5-fold increase in TGF-b, p=0.0251, and 2-fold reduction in TNFa, p=0.019), a recovery in the expression of antioxidant genes, normalization of oxidative stress loads, and improvement in keratinocyte proliferation compared to siNS-DHEKs. Keap-1 silenced DHEKs recover the ability to migrate in scratch assays with significant improvement over siNS-DHEKs as early as 2 hours post-injury (1255 vs. 1560 mm2, p=0.014). In surgical specimens of wounded nondiabetic skin, Keap1 levels decrease, intranuclear Nrf2 increases, resulting in increased antioxidant gene expression to maintain redox homeostasis. However, in the wounds of diabetic patients, Keap1 remains elevated with a corresponding decrease in active Nrf2. In particular, the basal epidermis of wounded diabetic skin demonstrates a 77% decrease (p=<0.0001) in the expression of Nrf2. We present evidence that human diabetic epidermal Nrf2 dysfunction likely coincides with that of wounded tissue and results in reduced antioxidant gene expression, a rise in reactive oxygen species, and impaired re-epithelization. Exploiting the Nrf2 pathway may show great potential as a therapeutic strategy to minimize pathologic wound burden in the diabetic population.
J. Kuhn: None. J.A. David: None. A.P. Villarreal Ponce: None. S. Abdou: None. D. Sultan: None. J. Kwong: None. P.S. Rabbani: None. D. Ceradini: None.
American Diabetes Association/Pathway to Stop Diabetes (1-16-ACE-08 to D.C.)