7,4'-dimethoxy-3-hydroxyflavone, a protease-activated receptor 4 (PAR4) inhibitor with antioxidant activity,...

Hyperglycaemia-induced protease-activated receptor 4 (PAR4) has been suggested to be linked with vascular complications in patients with diabetes mellitus. In the present study, we investigated if 7,4'-dimethoxy-3-hydroxyflavone (DMF-OH), a flavonoid-derived PAR4 antagonist, can ameliorate hyperglycaemia-associated endothelial dysfunction, which is a key early event in the pathogenesis of diabetic vasculopathy.

The effects of hyperglycaemia on PAR4 expression and endothelial dysfunction were examined in a streptozotocin-induced diabetic mouse model and in a high glucose-treated human endothelial cell model. The contribution of PAR4 was studied by using selective antagonists and shRNA-mediated knockdown.

In diabetic mice, up-regulation of vascular PAR4 was associated with endothelial dysfunction, defined by increases in proinflammatory cytokines, adhesive molecules, macrophage infiltration, oxidative stress and the procoagulant tissue factor. These abnormalities were ameliorated by oral administration of 7,4'-dimethoxy-3-hydroxyflavone, a flavonoid-derived PAR4 antagonist. In cultured endothelial cells, high glucose exposure induced PAR4 expression and enhanced Ca²⁺ responses to PAR4-activating peptide and thrombin, leading to exaggeration of proinflammatory and procoagulant states. The tissue factor-thrombin-PAR4 loop was blockaded by selective PAR4 antagonists and, to a greater extent, by 7,4'-dimethoxy-3-hydroxyflavone. The extra benefit of the latter may be attributed to inhibition of reactive oxygen species (ROS)-dependent NF-κB activation, which is required for hyperglycaemia-induced PAR4 and proinflammatory factors.

Our results indicate an important role for hyperglycaemia-induced PAR4 in aggravating diabetic endothelial dysfunction and suggest a novel strategy targeting PAR4 and ROS for treating vascular complications in diabetes.