Development of a Hypoxia-Induced Zebrafish Choroidal Neovascularization Model - Zebrafish: Methods for Assessing Drug Safety and Toxicity - page 198

(Semenza, 2000; Schwesinger et al., 2001; Smith, 2002; Shih et al., 2003). Diabetic
retinopathy (DR) and age-related macular degeneration (AMD) are the two leading
causes of adult blindness in developed countries and retinopathy of prematurity
(ROP) is a major cause of vision loss in premature infants. DR occurs when diabetes
damages small blood vessels in the retina. As this disease progresses and enters the
proliferative stage, newly formed blood vessels grow along the retina and in the clear,
gel-like vitreous body that fills the interior of the eyes. These blood vessels can bleed,
cloud vision, and destroy the retina. Macular degeneration (MD) targets the central
area of the retina macula. AMD is usually categorized as either “wet” or “dry.” In the
more common “dry” form, fat deposits damage macular cells. In the “wet” form,
abnormally proliferating blood vessels beneath the retina in the choroid rupture and
leak fluid that damages light-sensitive cells in the macula region. This disease can
progress rapidly, eventually leading to vision loss.

16.1.2 Hypoxia and CoCl 2

HIF-1 is a significant hypoxia transcription factor. Under hypoxic conditions,

ubiquitination decreases dramatically, causing HIF-1

a

a accumulation. HIF-1 a di-

merizes with HIF-1 , binds to the hypoxia response element located in the promoter

region of the corresponding genes, and upregulates gene expression. Several genes,
including glucose/energy metabolism genes and vascular development/remodeling
genes, such as VEGF , which promotes progression of AMD and DR, are known to
respond toHIF transcription factor. Increased VEGF has been observed in both human
retinopathy andMD (Rasmussen et al., 2001). Mounting evidence suggests that genes
involved in vascular development/remodeling, including IGF , also play pivotal roles
in pathological angiogenesis (Bouck, 2002). Oxygen level, which correlates with iron
level and heme protein synthesis, can be modulated by iron chelators, and Co 2þ ,
produced by CoCl 2 , has been found to be the most potent iron chelator (Rafii
et al., 2000). In human cells, CoCl 2 has been shown to induce HIF-1

b

expression

a

and inhibit HIF-1 ubiquitination (Jiang et al., 1997; Yuan et al., 2003). CoCl 2 has

also been shown to increase activity in the HIF-1

a

transcriptional domain and
upregulate VEGF (Van Lieshout et al., 2003). CoCl 2 mimics the hypoxia mechanism
and upregulates downstream genes that are reactive to HIF-1.

a

16.1.3 Current Neovascularization Models

Although HIF and VEGF have been shown to be involved in abnormal ocular
neovascularization (Schwesinger et al., 2001; Smith, 2002; Shih et al., 2003), drug
development has been hindered by the lack of suitable animal models. A ROP model
has been generated by initially exposing animals to a high oxygen atmosphere,
simulating hypoxic conditions, followed by return to normal levels (Shih et al., 2003).
Current in vivo CNV rodent models are highly manual and rely on lengthy surgical
procedures performed by well-trained personnel, limiting our understanding of the
underlying mechanisms of CNV and inhibiting drug discovery.