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Background: Fenofibrate, a PPARα agonist, reduced amputation events in the Fenofibrate Intervention and Event Lowering in Diabetes study among 9795 patients, the mechanisms of which are unknown. We hypothesised that fenofibrate may attenuate diabetes-related impairment in ischemia-mediated neovascularization. Methods: Hindlimb ischemia was induced in a murine model of streptozotocin induced diabetes mellitus (DM) in wildtype and PPARα receptor knockout mice (KO) with/without fenofibrate (30mg/kg/day in high fat diet). Ischemic recovery was assessed by laser doppler, foot movement and capillary density analysis. Key angiogenic events (tubulogenesis) and expression of thioredoxin-interacting protein (TXNIP) mRNA and protein following normal and high glucose (5 vs 25mM glucose) environments were determined in cultured endothelial cells (ECs), with/without selective PPARα receptor antagonist (10μM MK886) and agonist (10μM WY14643). Results: DM profoundly impaired blood flow recovery following hindlimb ischemia (0.51±0.04[n=5] vs 0.34±0.03[n=5];p Conclusions: Fenofibrate rescues diabetic impairment in ischemia-mediated angiogenesis via a largely PPARα-independent mechanism. Improved neovascularization may therefore explain in part the reduced amputations seen with fenofibrate in type 2 diabetes. These findings provide a novel mechanistic understanding for fenofibrate action and may provide a platform for new drug targets.

Introduction: Endothelial progenitor cell (EPC) participation in neovascularization of ischemic tissue is impaired in diabetes mellitus. Thioredoxin interacting protein (TXNIP) is strongly induced by glucose. TXNIP is overexpressed in tissues of diabetes sufferers and is associated with impaired angiogenesis. We hypothesized that gene-silencing of TXNIP in diabetic EPC would ameliorate impaired EPC angiogenic function. Methods and results: EPC (CD34+/CD45+/KDR+) isolated from young Type 1 diabetes patients were markedly reduced, which was associated with induction of TXNIP and reduced expression of VEGF compared to age- and sex-matched healthy controls. We further found that high glucose (15-25 mmol/L) impaired EPC angiogenic processes in vitro, including cell proliferation, migration and vascular network formation. This was associated with induction of TXNIP and a reduction in VEGF. However, EPC transfected with siRNA to TXNIP were protected from high-glucose mediated impairment of angiogenic processes and had normal VEGF production. Next, diabetic and healthy EPC were treated with TXNIP siRNA and xenotransplanted into a normoglycemic Balb/c nude mouse model of unilateral hindlimb ischemia. Xenotransplanted diabetic EPC were associated with reductions in blood flow recovery (laser Doppler perfusion imaging) and increased tissue ischemia/amputations. However, gene silencing of TXNIP in diabetic EPC restored blood flow recovery, with reductions in tissue ischemia/amputations, to levels seen in mice xenotransplanted with healthy control EPC. Conclusion: Hyperglycemia-mediated induction of TXNIP impairs EPC angiogenic function in vitro and in vivo. Inhibition of TXNIP in diabetic EPC rescues impaired angiogenesis, with implications for therapeutic modulation of diabetic vascular disease.