High glucose-induced changes in hyaloid-retinal vessels during early ocular development of zebrafish: a short-term animal model of diabetic retinopathy.
OBJECTIVE
Although a variety of animal models have been used to test drug candidates and examine the pathogenesis of diabetic retinopathy, time-saving and inexpensive models are still needed to evaluate the increasing number of therapeutic approaches.


METHODS
We developed a model for diabetic retinopathy using the early stage of transgenic zebrafish (flk:EGFP) by treating embryos with 130 mM glucose, from 3-6 days post fertilisation (high-glucose model). On day 6, lenses from zebrafish larvae were isolated and treated with 3% trypsin, and changes in hyaloid-retinal vessels were analysed using fluorescent stereomicroscopy. In addition, expression of tight junction proteins (such as zonula occludens-1), effects of hyperosmolar solutions and of hypoxia, and Vegf expression were assessed by RT -PCR. NO production was assessed with a fluorescent substrate. Effects of inhibitors of the VEGF receptor, NO synthesis and a VEGF antibody (ranibizumab) were also measured.


RESULTS
In this high-glucose model, dilation of hyaloid-retinal vessels, on day 6, was accompanied by morphological lesions with disruption of tight junction proteins, overproduction of Vegf mRNA and increased NO production. Treatment of this high-glucose model with an inhibitor of VEGF receptor tyrosine kinase or an inhibitor of NO synthase or ranibizumab decreased dilation of hyaloid-retinal vessels.


CONCLUSIONS
These findings suggest that short-term exposure of zebrafish larvae to high-glucose conditions could be used for screening and drug discovery for diabetic retinopathy and particularly for disorders of retinal vessels related to disruption of tight junction proteins and excessive VEGF and NO production.
Korean Medicine Convergence Research Division, Korea Institute of Oriental Medicine (KIOM), Daejeon, 305-811, Korea.
Seung-Hyun
SH
Jung
Korean Medicine Convergence Research Division, Korea Institute of Oriental Medicine (KIOM), Daejeon, 305-811, Korea.
Young Sook
YS
Kim
Korean Medicine Convergence Research Division, Korea Institute of Oriental Medicine (KIOM), Daejeon, 305-811, Korea.
Yu-Ri
YR
Lee
Korean Medicine Convergence Research Division, Korea Institute of Oriental Medicine (KIOM), Daejeon, 305-811, Korea.
Jin Sook
JS
Kim
D000818:Animals; D030801:Animals, Genetically Modified; D003930:Diabetic Retinopathy / Q000139:chemically induced / Q000378:metabolism*; D004195:Disease Models, Animal; D005947:Glucose / Q000009:adverse effects*; D007814:Larva / Q000187:drug effects / Q000378:metabolism; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009569:Nitric Oxide / Q000096:biosynthesis; D000069579:Ranibizumab / Q000494:pharmacology; D040262:Receptors, Vascular Endothelial Growth Factor / Q000378:metabolism; D012171:Retinal Vessels / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D062725:Tight Junction Proteins / Q000378:metabolism; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis; D015027:Zebrafish / Q000254:growth & development / Q000378:metabolism*; D029961:Zebrafish Proteins / Q000096:biosynthesis
D062725:Tight Junction Proteins; D042461:Vascular Endothelial Growth Factor A; C583604:Vegfaa protein, zebrafish; D029961:Zebrafish Proteins; D009569:Nitric Oxide; D040262:Receptors, Vascular Endothelial Growth Factor; D005947:Glucose; D019331:NG-Nitroarginine Methyl Ester; D000069579:Ranibizumab