Abstract 838: Ablation of AngiotensinIV Receptor Attenuates Hypofibrinolysis in Acute and Chronic Thrombosis Models via Plasminogen Activator type 1 (PAI-1) Down Regulation.
Reduced fibrinolytic activity is associated with adverse cardiovascular events. Although Insulin-Regulated Aminopepeidase (IRAP) was recently identified as angiotensin (Ang) IV receptor (AT4R), the extent to which Ang IV-IRAP signaling affects PAI-1 activation in the fibrinolytic process remains unknown. To test hypothesis that AngIV would inhibit fibrinolysis via PAI-1 activation, we evaluated the degree of fibrinolysis in thrombosis models using IRAP knockout mice (IRAP−/−).
Methods and Results: IRAP−/− showed normal glucose levels (68±6 mg/dl, mean±SD), and their visceral fat weight did not differ from that of wild type mice (C57BL6/J; WT). We tested the levels of PAI-1 mRNA expression in endothelial cells (ECs) isolated from WT and IRAP−/− (n=3 each). In ECs from WT, AngIV treatment (10-9 to 10-6 M) as well as AngII (10-9 to 10-6 M) increased PAI-1 mRNA expression in a dose dependent manner, while AngII (but not AngIV) increased PAI-1 mRNA in ECs from IRAP−/−. To assess fibrinolysis in an acute thrombosis model, renal glomerular fibrin deposition was counted after lipopolysaccharide injection (0.2mg/kg, n=5). Fibrin deposition was lower in IRAP−/− than in WT at 4 hrs (12±3% of total gromeluli vs. 27±4%, p<0.012). The levels of plasma active PAI-1 antigen as measured by ELISA were also lower in IRAP−/− than in WT (120±31 vs. 62±12 ng/mL, p<0.001). NF-kB activation confirmed by phosphorylation of p65 and nuclear translocation of Rel A in the livers was suppressed in IRAP−/− by 43% and 32%, respectively. For a chronic thrombosis model, thrombus formation was observed over 7 days after carotid artery ligation and cuff placement. Intraluminal thrombus occlusion was observed in the arteries of WT by 100%, while 67% in IRAP−/− (p<0.01, n=12). The length of thrombi from the ligated point was shorter in IRAP−/− compared to WT(0.89±0.2 vs 1.9±0.3mm, p<0.016).Consistently, historlogical analysis demonstrated that mononuclear cell infiltration and the organization of thrombi were suppressed in the arteries of IRAP−/−.
Conclusions: In acute and chronic thrombosis models, disruption of AngIV-IRAP signaling leads to accelerated fibrinolysis with decreased PAI-1 expression, suggesting that AngIV pathway is a novel therapeutic target against hypofibrinolysis.