Abstract 18963: Over-expression of Angiopoietin-2 in Human Patients With Continuous-flow Left Ventricular Assist Devices Induces Abnormal Angiogenesis and Predicts Gastrointestinal Bleeding Events

Abstract

Arteriovenous malformations are common in patients with Continuous-Flow Left Ventricular Assist Devices (LVAD) and may reflect deregulation of angiogenesis. Angiopoietin-2 (Ang-2) is a stimulator of angiogenesis produced by endothelial cells in response to thrombin. In this study, we evaluate Ang-2 expression in patients with LVAD compared with stable heart failure (HF) or orthotopic heart transplant (OHT) and test the hypothesis that elevated Ang-2 in patients with LVAD leads to increased angiogenesis. Blood samples were taken from 79 patients (n= 21 HF, 41 LVAD, 17 OHT) during routine heart catheterization. Ang-2 was measured by ELISA and thrombin was measured by Western Blot. Cultured human umbilical vein endothelial cells (HUVECs) were incubated with plasma from each patient and Ang-2 expression was measured by RT-PCR. HUVECs were then incubated on Matrigel with serum from each patient in the presence or absence of an Ang-2 blocking antibody and tube formation was measured by microscopy. Also, vena caval endothelial cells were isolated from discarded guide wires used during catheterization and Ang-2 expression was measured by quantitative immunofluorescence. In patients with LVAD vs. HF or OHT, Ang-2 was elevated in both serum (12.3±9.6, 4.5±2.1, and 4.9±2.0 ng/mL respectively, p<0.01) and endothelial cells (56.3±14.1, 31.3±4.2, 35.0±11.2 AU respectively, p<0.05). On Matrigel, serum from patients with LVAD vs. HF or OHT induced increased tube formation (57.1±3.2, 39.2±7.1, 34.7±5.1 tubes per low power field respectively, p<0.05) and this elevation normalized with Ang-2 blocking antibody. Plasma from patients with LVAD vs. HF or OHT induced Ang-2 gene over-expression in HUVECs (1.6±0.4, 1.0±0.3, 0.6±0.2 RQ respectively, p<0.01) and was associated with higher plasma thrombin. In patients with LVAD, serum Ang-2 >10 ng/mL was associated with a higher 3 month risk of GI bleeding vs. Ang-2 <10 ng/mL (30% vs. 0%, p<0.01). Therefore, we conclude that over-expression of Ang-2 in patients with LVAD induces increased angiogenesis and predicts GI bleeding events. Thrombin is increased in patients with LVAD and their plasma induces Ang-2 over-expression in vitro . Therefore, Ang-2 overexpression may contribute to AVM formation in patients with LVAD.