3.2.2. Models of diabetic retinopathy
Rodent models are the most commonly used animal models to study diabetic retinopathy. Traditionally, the mouse model of retinopathy of prematurity (ROP) has been used as a surrogate model to study the formation of new malformed and leaky vessels, a hallmark of diabetic retinopathy. In this model, pups with their mothers are placed in an oxygen chamber (75% O2 ± 5% and 2% CO2 for 22h/day) at postnatal day 7 until postnatal day 11, after which they are returned to room air until day 18. The early exposure to elevated oxygen causes a central obliteration of the vasculature, whilst the return to room air mediates an hypoxia-driven increase in VEGF via HIF-1a resulting vascular neogenesis. Using this model, we have shown that oxidative stress is a key driver of neovascularisation[123], and that interventions like ebselen to bolster specific antioxidants such glutathione peroxidase-1[124] leads to significant attenuation of vaso-obliteration and neovascularisation. In addition, by targeting the master regulator of oxidative stress, namely the transcription factor Nrf2 via the bardoxolone-methyl mimetic dh404, we additionally demonstrated that a global approach to bolstering antioxidant defence improved ROP in our mouse models[125].
To study diabetic retinopathy per se mouse or rat models can be used after exposure to STZ. Rat models (a single tail vein injection of STZ at 55mg/kg with daily insulin) enable robust analysis of vascular leakage after 10-weeks of diabetes. In most experimental models, Sprague-Dawley rats are used for diabetic retinopathy studies. Gene and protein expression analysis of diabetic retinae enable the study of the molecular pathways involved, with emphasis on VEGF since this growth factor is the main causal factor in the vascular permeability that accompanies diabetic retinopathy. Similarly, the use of STZ has enabled the long-term study of diabetic retinopathy in mice. As mentioned earlier, female mice are known to be resistant to STZ. Recently, a study by Saadane et al have reported that the resistance of female C57BL/6J mice to STZ-induced diabetes can be overcome by injecting higher doses of STZ[26]. Female mice administered a dose of 75mg/kg of body weight over 5 consecutive days, developed equivalent levels of hyperglycaemia as male counterparts administered 55mg/kg of body weight over 5 consecutive days, which was maintained for at least 8months, enabling a robust evaluation of diabetic retinopathy in this model. Importantly, both male and female diabetic mice developed similar abnormalities in retinal oxidative stress as measured by the level of superoxide in the retinae. This was accompanied by significantly increased expression of pro-inflammatory proteins, leucocyte mediated cytotoxicity and similarities in the number of degenerated capillaries in both male and female mice[26]. Thus, varying STZ dosage is an easy adjustment to enable the inclusion of both sexes in the exploration of therapeutic agents for complications such as diabetic retinopathy.
Inflammatory processes play a key role in driving diabetic retinopathy. To directly assess a role for inflammatory cytokines in the progression of diabetic retinopathy, Mugisho et al have developed a novel animal model that incorporates both hyperglycaemia and inflammation via single intravitreal injections of the pro-inflammatory cytokines, IL-1ß and TNF-α, in non-obese diabetic (NOD) mice[126]. The injection of intravitreal cytokines evokes inflammation in the surrounding ocular tissue and exhibits retinal oedema, increased astrogliosis and microglia upregulation. In addition, pericyte loss observed by advanced vessel dilation and vessel beading, endothelial cell death and blood-retina barrier breakdown, are prominent[126]. The model may provide a new opportunity to further study the pathology of diabetic retinopathy, although its proneness to autoimmunity due to a genetic defect causing cytokine dysregulation could be a limitation. However, the effectiveness of this model was further highlighted in a follow-up study where connexcin43, an inhibitor of inflammation and inflammasome activation, was shown to significantly attenuate diabetic retinopathy[127].