Neutralizing VEGF decreases tortuosity and alters endothelial cell division orientation in arterioles and veins in rat model of ROP: Relevance to plus disease
M. Elizabeth Hartnett 1*,
David J Martiniuk 2,
Grace E Byfield 2,
Pete Geisen 2,
Gefei Zeng 3,
and
Victoria L Bautch 3
1 Ophthalmology, University of North Carolina, 103 Mason Farm Road, Chapel Hill, North Carolina, 27599-7041, United States
2 Ophthalmology, University of North Carolina, Chapel Hill, North Carolina, United States
3 Dept. of Biology and Carolina Cardiovascular Biology Center, University of North Carolina, Chapel Hill, North Carolina, United States
* To whom correspondence should be addressed. E-mail: hartnet{at}med.unc.edu.
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Abstract |
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Purpose: To study the effects of vascular endothelial growth factor (VEGF) on endothelial nitric oxide synthetase (eNOS) and retinal vascular tortuosity and cleavage planes in a rat model of retinopathy of prematurity (ROP).
Methods: Within 4 hours of birth, pups and mothers were cycled between 50% and 10% oxygen daily. At postnatal (p)12, pups received either intravitreous anti-rat neutralizing antibody to VEGF or control non-immune rat IgG in one eye and returned to oxygen cycling until p14 when they were placed into room air (RA) for 4 days (50/10 oxygen-induced retinopathy [50/10OIR]). Tortuosity indices and endothelial cleavage plane angles relative to the long axes of the major retinal vessels during anaphase were calculated from phosphohistone and Alexa-isolectin stained retinal flat mounts. Some retinas were processed for eNOS protein or phosphorylated/total eNOS.
Results: Retinas from 50/10 OIR had increased tortuosity over time with peaks at p12 and p14 (p<0.001 vs. RA) prior to the development of intravitreous neovascularization, which peaks at p18. Compared to RA, eNOS/actin in 50/10 OIR retinas was increased at p12 (p=0.0003) and p14 (p=0.047). Inhibition of VEGF with a neutralizing antibody decreased tortuosity and caused endothelial mitosis cleavage planes to orient in favor of vessel elongation but did not affect eNOS protein or activation.
Conclusions: In the 50/10 OIR model, a model with relevance to ROP, arteriolar tortuosity and venous dilation are increased through VEGF, which influences the orientation of endothelial cell cleavage in major arterioles and veins, independent of eNOS.
Key Words:
retinopathy of prematurity, growth factors, retinal vasculature, retinal development, VEGF, mitosis planes
