Diabetic foot ulcers that lead to amputations are a major health problem affecting ~20% of the 30 million diabetic patients in the US. The current regimen has limited success, and the amputation rates remain high. Therefore, understanding molecular mechanisms for compounds with translational potential is a crucial step toward making a breakthrough in wound care protocols. Endothelial cells (ECs) are indispensable cellular components for wound angiogenesis. However, EC functions are impaired in patients with diabetes. The coformulation of two dietary compounds - Trans-resveratrol (tRES) and hesperetin (HESP) - improves glucose metabolic profile and arterial function in overweight and obese subjects through inducing the gene expression of glyoxalase 1 (GLO1), an enzyme that detoxifies reactive metabolites during glycolysis and protects cells against glycation stress. Our pilot data indicated that tRES+HESP improved wound healing in diabetic animals with an increase in GLO1 expression. However, its effects are likely far beyond inducing GLO1 expression because tRES+HESP treated ECs produced many pro-angiogenic factors, including angiopoietin-1 (ANGPT1) that plays an essential role in angiogenesis. Therefore, it is critical to determine proteins that are regulated by tRES+HESP in angiogenesis and tissue repair. The objective of this project is to fill the knowledge gap of the role of tRES+HESP in rescuing the disrupted angiogenesis in diabetes, and our long-term goal is to develop therapeutic strategies for diabetic wound repair. We hypothesize that tRES+HESP augments angiogenesis and improves diabetic wound healing through enhancing the expression of GLO1 and a potent pro-angiogenic factor, ANGPT1, and through novel changes in additional proteins in pathways critical to diabetic wound repair. Aim 1: Identification of molecular pathways and protein changes induced by tRES+HESP in human dermal microvascular ECs in vitro. Sub-aim 1: Determine to what extent tRES+HESP can rescue diabetic endothelial cell function in vitro. Sub-aim 2: Determine how vital ANGPT1 is in tRES+HESP-induced angiogenesis in vitro. Sub-aim 3: Discover new proteins and pathways responsible for the benefit of tRES+HESP treatment in endothelial cell function in vitro using state- of-the-art proteomics. Aim 2: Determine the therapeutic potential of tRES+HESP and its underlying molecular mechanisms in chronic diabetic wounds in vivo. Sub-aim 1: Determine the efficacy of tRES+HESP on wound healing in a newly developed diabetic chronic wound model in db/db mice. Sub-aim 2: Determine the role of ANGPT1 in the tRES+HESP-induced improvement in wound healing in vivo. Sub-aim 3: Discover new proteins and pathways responsible for the benefit of tRES+HESP treatment in diabetic wound repair in vivo using state- of-the-art proteomics. The outcome of the proposed research will determine the efficacy of topical application of this formula, tRES+HESP, in diabetic wound healing, and will unveil underlying molecular mechanisms for its beneficial effect. Since tRES+HESP has not been approved by the FDA to treat diabetic wound healing yet, these results may facilitate the FDA approval of this coformulation in diabetic wound treatment.

The objective of this project is to fill the knowledge gap of how an FDA-approved oral formula of small molecules, tRES+HESP, can enhance the new blood vessel formation in treating non- healing wounds in diabetic patients as a topical application. We will evaluate the therapeutic potential of tRES+HESP and its underlying molecular mechanisms in human vascular endothelial cells in Aim 1, and in a newly developed diabetic chronic wound animal model in Aim 2, using a combination of cell culture and animal studies (for causal mechanisms in vitro and in vivo) and cutting-edge proteomics studies (for global analysis of cell signaling & unbiased discovery). The outcome of the proposed research will determine the efficacy of topical application of this formula, tRES+HESP, in diabetic wound healing, and will unveil underlying molecular mechanisms for its beneficial effect.