Diabetic macro- and microangiopathy is associated with a high risk of vascular complications and in consequence a high morbidity and mortality rate. The diabetic patient exhibits a pathological coagulation state, which is represented by both an increased synthesis of factor VII, thrombin, tissue factor and plasminogen activator inhibitor 1 (PAI-1) as well as by platelet aggregation enhanced due to elevated GP IIb/IIIa expression and altered calcium homeostasis. Former studies showed improved microvascular blood flow of skin and muscle after C-peptide administration in type 1 diabetic patients. Additionally, positive effects of C-peptide application on leukocyte-endothelial-interaction with increased production of nitric oxide and decreased expression of endothelial P-selectin and ICAM-1 have been reported. The aim of this study was therefore to examine whether the administration of C-peptide has anti-thrombotic effects in vivo. In microvessels of mouse cremaster muscle preparations of diabetic and non-diabetic mice, ferric chloride-induced thrombus formation was analyzed using intravital fluorescence microscopy. C-peptide was applied systemically in low (7 nmol/kg) and high (70 nmol/kg) dose, while control groups received equivalent amounts of heat-inactivated C-peptide. Diabetes was induced by application of streptozotocin (40 mg/kg) on 5 consecutive days. The diabetic metabolic state was confirmed by repeated positive testing for hypergly-cemia and glucosuria. Additionally, flow cytometric analysis of human platelet P-selectin and GP IIb/IIIa expression and immunohistochemistry of endothelial PAI-1 expression within the cremaster muscle tissue were conducted. Administration of C-peptide in high, but not in low dose, caused a significant delay in venular and arteriolar thrombus growth in diabetic mice (complete occlusion: 1171 ± 233 s and 1317 ± 218 s vs. control: 496 ± 50 s and 488 ± 50 s; p<0.05) and in non-diabetic mice (complete occlusion: 1063 ± 286 s and 908 ± 289 s vs. control: 367 ± 48 s and 382 ± 59 s; p<0,05). Endothelial PAI-1 expression was found to be reduced in C-peptide-treated diabetic and non-diabetic mice, when compared with control. Additionally, C-peptide induced a slight, but significant, decrease of resting platelet P-selectin and GP IIb/IIIa expression, while no effect of C-peptide on maximal platelet activation upon exposure to TRAP (thrombin receptor activating peptide) was seen. The present study indicates an anti-thrombotic effect of C-peptide in vivo. The role of PAI-1 in this setting is subject of ongoing studies and remains to be elucidated. The application of C-peptide might be a future treatment option to improve rheology and microcirculation in diabetic patients.