Excess VEGF‐A disrupts tubular heart formation and cardiac blood flow dynamics
Authors: Rugonyi Sandra , Rykiel Graham , Rongish Brenda 
Published: June 2021

Vascular Endothelial Growth Factor (VEGF) plays a critical role during heart development. Clinical evidence shows that maternal conditions that modify VEGF signaling in utero, such as high altitude hypoxia and maternal diabetes, are correlated with an increased risk of congenital heart defects. While VEGF is essential for vasculogenesis and angiogenesis, its role during early heart tube formation is not fully understood. During early embryogenesis, paired endocardial tubes fuse into a single linear heart tube, which ultimately transforms into a four‐chambered heart. To better understand the role of VEGF in heart tube fusion, we studied how an early transient dysregulation of VEGF, produced by adding exogenous VEGF‐A right before tube fusion, affects heart tube formation and the onset of blood flow in the heart, leading to congenital heart defects.To enable longitudinal studies of heart development, and determine effects on early and late cardiac formation events, we developed an in ovo quail model of early transient VEGF dysregulation. We chose quail due to ease of access and monitoring during development. We applied recombinant human VEGF165 (rhVEGF165; VEGF‐A isoform) to quail embryos in ovo at Hamburger‐Hamilton (HH) stage 8 (~29 hours incubation) just before heart tube fusion (500ng in a 5μL drop on top of the embryo). Control embryos (vehicle applied at HH8) were also prepared. Eggs were then resealed and re‐incubated for an additional 13hrs (approximately HH11, early tubular heart); 22hrs (around HH14, early beating tubular heart); or 10 days (HH38, fully formed heart); to study how a short, transient, early exposure to excess VEGF‐A impacts cardiac formation.We found both early and late cardiac deficiencies after transient VEGF exposure at HH8. Around HH11, the newly formed tubular heart presented three distinct phenotypes when exposed to early VEGF excess: 1. Wider and shorter than normal heart tubes; 2. Heart tubes with multiple ridges (abnormal folds and protrusions); and 3. Lack of heart tube. Total number of malformed hearts accounted for 70% (n = 21/30) in VEGF‐treated embryos versus 27% (n = 6/22) in controls. Around HH14, when the heart is beating, we measured blood flow and cardiac wall mechanics. We found that heart tubes that were significantly wider than normal (~30%) after VEGF‐treatment, had about 30% increase in maximum flow velocity (n = 3), and their heart tubes failed to completely close the lumen to avoid backflow. When the heart was fully formed, at HH38, microCT images revealed four malformation phenotypes: 1. Ventricular septal defect (VSD); 2. Right ventricular hypoplasia; 3. Hearts that were both shorter and wider than normal; and 4. Aortic arch anomalies. Total number of malformed hearts accounted for 28% (n = 9/32) of surviving VEGF‐treated embryos; and 14% (4/28) surviving control embryos. Our data highlight that VEGF‐A is involved in shaping the geometrical characteristics of the early heart tube, and thus the onset of blood flow dynamics, together affecting mature heart formation.Support or Funding InformationNIH R01 HL094570; AHA 16GRNT29840002; OHSU Presidential Bridge Fund.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.