Abstract
Purpose: :
To assess endothelin expression over time in the trVEGF029 model of RNV
Methods: :
Total RNA from 10 week old transgenic (TG) and wild-type (wt) mouse posterior eye cups was hybridised to Affymetrix Mouse Genome 430 GeneChips®. Levels of endothelin (Edn1, Edn2, Edn3) and endothelin receptor (EdnRa, EdnRb) gene expression were investigated in TG and wt mice by real-time PCR (RT-PCR) from 10 days to 27 weeks. Edn2 protein expression was localised in TG retinae by immunohistochemistry.
Results: :
Effect of VEGF over-expression in 10 week-old TG mice led to upregulation of 11 genes. When compared to wt animals, Edn2 was the most significantly upregulated gene in TG mice [6-fold change (FC) by microarray; 13.5 FC; RT-PCR]. At 10 days postnatal, all endothelins and their receptors were significantly upregulated compared to wt animals (approx. 5.5-FC; RT- PCR]. From 3 to 27 weeks postnatal, there was no change in expression levels of Edn1, Edn3, EdnRa and EdnRb compared to wt. However, Edn2 expression remained high (>4.6-FC; RT- PCR). Edn2 protein was localised in the retinal pigment epithelial (RPE) layer in the TG mice.
Conclusions: :
Endothelins are a family of vasoactive peptides and gene expression of Edn2 was significantly increased in response to VEGF upregulation at all timepoints examined in the trVEGF029 mouse model of RNV. Edn2 expression remained high through to 27 weeks whilst Edn1, Edn3, EnRa and EdnRb expression was unchanged compared to wt animals from 3 weeks onwards. Edn1 and Edn3 may play a role in early postnatal angiogenic development in the eye, but may not be involved in the RNV response to VEGF upregulation in these animals. Interestingly, Edn2 has not been directly linked to VEGF and RNV. Our novel finding suggested that an early spike of VEGF expression in the retinae of trVEGF029 was sufficient to induce significant long-term Edn2 expression, mostly in the RPE layer. In conclusion, Edn2 may be a contributing factor in RNV and might be a good target to reduce neovascularisation in the eye.
Keywords: retinal neovascularization • diabetic retinopathy • candidate gene analysis