An urgent need exists to identify predictive biomarkers to guide management of diabetic foot ulcers (DFUs). The steroid hormone, cortisol, is elevated in chronic inflammatory states and its pathogenic role and/or prognostic role for poor outcomes across a range of diseases has been suggested. However, the role of cortisol in DFUs is not well understood and the significance of local, tissue-specific cortisol regulation in DFUs remains unclear. We performed a multi-center study of 63 well vascularized, superficial neuropathic, non-infected DFU subjects receiving standard care and used wound biopsy obtained at baseline study to quantify expression levels (via immunohistochemistry) of the major cortisol synthesis enzymes CYP11B1 and 11BHSD1, and other inflammatory cytokines. Subjects were predominantly men in their late-fifties (58±10 years) with poor glycemic control (HbA1C 8.9±2.3%); while hemoglobin (12.6±1.8g/dl), creatinine (1.27±0.5mg/dl) and albumin (4.1±0.42g/dl) were within the normal range. We found high cutaneous levels of CYP11B1 strongly linked to failure to heal DFUs at 6 weeks (p≤0.016), while 11BHSD1 did not correlate with healing (p=0.351). In the logistic regression analyses controlling for demographics and disease characteristics, higher levels of CYP11B1 (10-based log-transformed) were predictive of non-healing of DFU at 6 weeks (OR = 6.83, p =.010). High local tissue levels of the pro-inflammatory TNFalpha were also significantly associated with lack of ulcer healing (p≤0.019). This observation is consistent with reports that TNFalpha is essential for local induction of CYP11B1. In turn, local induction of CYP11B1 and subsequent induced tissue cortisol levels activate membranous glucocorticoid receptor triggering PLC/PKC/GSK-3beta/beta-catenin pathway that inhibits healing. Taken together, these data suggest CYP11B1 is a potential predictive biomarker for DFU outcomes, and may serve as a future target for therapeutic interventions. Disclosure L. Vileikyte: None. B. Shen: None. I. Pastar: None. A.J. Boulton: None. R. Kirsner: None. M. Tomic-Canic: None. M. Hardman: None.