Abstract TMP118: Alk1 Deficiency Impairs Mural Cell Recruitment During Brain Angiogenesis
Background and Purpose: VEGF stimulation induced dysplasia vessel formation in the Alk1-deficient brain. The dysplastic vessels had less mural cell coverage associated with increased extravasation of blood contents. Platelet-derived growth factor B (PDGFB) signaling is important in mural cell recruitment during angiogenesis. Notch signaling is upstream of PDGFB. We hypothesized that Alk1 deletion impairs pericyte recruitment through downregulation of DLL4/PDGFB signaling. Thalidomide can increase PDGFB expression and was used to test our hypothesis in vivo.
Methods: Human brain microvascular endothelial cells (HBMECs) were infected with a lentiviral vector carrying ALK1 shRNA. Cells with 80% reduction of ALK1 gene expression were treated with vascular endothelial growth factor (VEGF; 0, 10, 50, and 100 ng/ml) for 18 hours. DLL4 and PDGFB mRNA were measured using real-time PCR. Pericyte-recruitment was evaluated by co-culturing pericytes with HBMECs. Cerebrovascular dysplasia was induced by co-injection of an adenoviral vector expressing cre-recombinase and an adeno-associated viral vector expressing VEGF into the basal ganglia of Alk1-floxed mice. Two weeks later, thalidomide (75 mg/kg, i.p.) or DMSO was administered twice per week for six weeks.
Results: Knockdown of ALK1 attenuated the increase of DLL4 and PDGFB in HBMECs following VEGF stimulation. DLL4 and PDGFB expression was highly correlated (R2 = 0.93). ALK1 knockdown reduced the ability of HBMEC to recruit pericytes (P = 0.014). In vivo, thalidomide treatment significantly reduced vascular dysplasia (dysplastic vessels: thalidomide vs. DMSO, 3.4±1.2/200 vessels vs. 1.0±0.5, P = 0.003), without affecting vessel density (P = 0.11). Smooth muscle-negative dysplastic vessels decreased in thalidomide-treated mice, compared to DMSO-treated mice (21%±6 vs 47±9, P = 0.0002). Prussian blue-positive area was reduced in the thalidomide group, compared to the DMSO group (P = 0.0011).
Conclusions: Our findings suggest that Alk1 regulates pericyte recruitment through DLL4/PDGFB signaling during brain angiogenesis.
- © 2012 by American Heart Association, Inc.