Cellular and vascular changes in the retina of neonatal rats after an acute exposure to hypoxia.
OBJECTIVE
This study was undertaken to examine the effects of an acute hypoxic exposure on the retinal cells and production of vascular factors such as vascular endothelial growth factor (VEGF) and nitric oxide (NO), which may affect vascular permeability in the developing retina.


METHODS
Retinas of 1-day-old rats were examined at 3 hours to 14 days after hypoxic exposure. The mRNA and protein expression of hypoxia-inducible factor-1alpha (HIF-1alpha), VEGF, endothelial nitric oxide synthase (eNOS), neuronal NOS (nNOS), and inducible NOS (iNOS) were determined by real-time RT-PCR, Western blot analysis, and immunohistochemistry. Electron microscopy was used to examine the structural alterations in retinal cells, and rhodamine isothiocyanate (RhIC) or horseradish peroxidase (HRP) was administered intraperitoneally or intravenously to determine vascular permeability.


RESULTS
The mRNA and protein expression of HIF-1alpha, VEGF, eNOS, nNOS, and iNOS, along with VEGF concentration and NO production, were increased in response to hypoxia. Swollen Müller cell processes, apoptotic and necrotic cells in the inner nuclear layer, and changes in ganglion cells such as swollen and disrupted mitochondria were observed in hypoxic animals. Increased leakage of RhIC and HRP from retinal and hyaloid vessels was seen after hypoxic exposure.


CONCLUSIONS
The authors suggest that increased VEGF and NO production in hypoxia resulted in increased vascular permeability, leading to changes in Müller cells and degeneration of neural cells. Melatonin administration reduced VEGF and NO production, diminished leakage of RhIC and HRP, and promoted cell proliferation, suggesting this as a potential therapeutic agent in reducing hypoxia-associated damage in the developing retina.
Kaur
Charanjit
C
Department of Anatomy, Yong Loo Lin School of Medicine, National University of Singapore, Singapore. antkaurc@nus.edu.sg
Sivakumar
Viswanathan
V
Foulds
Wallace S
WS
Luu
Chi D
CD
Ling
Eng-Ang
EA
D000818:Animals; D000831:Animals, Newborn; D000975:Antioxidants / Q000008:administration & dosage; D017209:Apoptosis; D015153:Blotting, Western; D002199:Capillary Permeability; D019084:Fluorescent Antibody Technique, Indirect; D000860:Hypoxia / Q000378:metabolism*; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000235:genetics / Q000378:metabolism; D007150:Immunohistochemistry; D007274:Injections, Intraperitoneal; D008550:Melatonin / Q000008:administration & dosage; D009457:Neuroglia / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism*; D052248:Nitric Oxide Synthase Type I / Q000235:genetics / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II / Q000235:genetics / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D015427:Reperfusion Injury / Q000378:metabolism* / Q000473:pathology / Q000517:prevention & control; D012164:Retinal Diseases / Q000378:metabolism* / Q000473:pathology / Q000517:prevention & control; D012171:Retinal Vessels / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000378:metabolism*
D000975:Antioxidants; C497444:Hif1a protein, rat; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D012333:RNA, Messenger; D042461:Vascular Endothelial Growth Factor A; C467498:vascular endothelial growth factor A, rat; D009569:Nitric Oxide; D052248:Nitric Oxide Synthase Type I; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III; C496318:Nos2 protein, rat; C496316:Nos3 protein, rat; D008550:Melatonin