*2.4.1 Peroxisome proliferator-activated receptor gamma coactivator – 1 alpha (Pgc1α)/Nrf2 DKO mice*

As stated previously, NRF2 is a transcription factor that regulates the expression of detoxifying and antioxidant enzymes. PGC-1α regulates angiogenesis and oxidative stress, among other functions. Single *Nrf2*−/− mice present signs of AMD, such as degenerated RPE and photoreceptor outer segment atrophy. When fed a HFD, *Nrf 2*−/− mice exhibited exacerbated signs of AMD such as RPE hyper/hypopigmentation. On a similar HFD, *Pgc-1α*+/− mice also develop increased lipofuscin accumulation, another indication of AMD [26].

*PGC-1α*/*Nrf2* DKO mice exhibit AMD signs and dysfunctional photoreceptors by 12 months of age. Autophagy and oxidative damage were greater in the DKO compared to single knockout mice [26]. This polygenic model may better represent the multifactorial nature of AMD. Environmental factors such as cigarette smoke or HFD exposure have not yet been tested in this model, which may be useful for future studies.

#### *2.4.2 Systems genetics and the BXD family of mice*

In recent years, the use of systems genetics to uncover genes underlying the pathological mechanisms of retinal diseases has become an invaluable tool in the study of multiple human diseases. Specifically, applying this approach to the BXD family of mice has elucidated novel genes associated with ocular hypertension and optic nerve necrosis in glaucoma [35, 36]. As a brief background, the BXD family of mice were generated by breeding the standard C57B/6 J mouse with the DBA/2 J strain from the Jackson Labs. Offspring were then inbred for 20 or more generations to allow for a homologous recombination-induced variety of genetic backgrounds [37]. Currently, studies are underway to use the BXD family of mice to generate a more accurate model of AMD in the mouse. This is being done by delving into the genomes of each strain of BXD mouse to find different combinations of haplotypes in AMD-associated genes which contain SNPs similar to those found in humans, or that result in an altered protein function as observed in humans. Currently, multiple strains have shown promise for not just AMD, but other retinal diseases as well.

#### **2.5 Rabbit models**

Although rodents are the most studied, the larger eye size of rabbits make them advantageous for certain pharmacological and pathological studies. While rabbits are more expensive than rodents, they are less expensive than non-human primates (NHP). Rabbits are also relatively easy to handle and breed. Furthermore, the size of the rabbits allow for easy administration of subretinal injections and vectors for

gene therapy [38]. Rabbits possess a visual streak where rods and cones are dense, but they do not have a macula which, like mice, can present a caveat for direct translation to human AMD [38, 39].

Promoting a wet AMD phenotype using conventional methods of inducing CNV in rodents and primates, such as laser-induced damage of BrM and injection of proangiogenic factors, have not worked for rabbit models; however, Qui *et al*. created an exudative AMD model in rabbits by injecting Matrigel, a membrane matrix mixture that includes growth factors like basic fibroblast growth factor and endothelial growth factor. Growth factors from the Matrigel are slowly released for up to 9 weeks, with subsequent production of CNV lesions and BrM disruption that are reminiscent of AMD. This shows promise of a way to test new therapies for exudative AMD on an inexpensive and reproducible model with similar pathological features of AMD [39].

#### **2.6 Non-human primate models**

Although primates and humans have the most similar anatomy, primates are disadvantageous as models because they are difficult to genetically manipulate, expensive, and their disease course is relatively long [17]. Furthermore, they are difficult to handle and breed [38]. Historically, there were limitations to exudative AMD models, however new techniques have generated some exudative AMD models, discussed below.

Because AMD affects the cone dense macula, a major limitation of animal models such as rodents, canines, and felines is the lack of a macula [40]. NHPs are the only pre-clinical animals that have a macula and a similar organization of photoreceptors within the macula like humans [17, 41]. Another advantage of NHP models is their shared similarity in organization of the visual pathway. The macula only receives nutrients and removes waste from the choroidal circulation. Furthermore, since the macula is responsible for high acuity central vision, copious amounts of light are focused on the macula subjecting it to high levels of ROS. These details may explain why the macula is affected in AMD in both humans and NHP [17].

Genetic risk factors are also suspected in NHPs with AMD. Polymorphisms in age-related maculopathy susceptibility 2 (*ARMS2*) and high-temperature requirement factor A1 (*HTRA1*) were linked to significantly higher rates of drusen formation in both humans and rhesus monkeys. These findings suggest shared genetic risk factors and can further infer that humans and rhesus monkeys have commonality in pathophysiology [17].

Another shared risk factor between humans and NHPs is diet. Rhesus monkeys with a diet without lutein or zeaxanthin formed drusen earlier than monkeys fed a standard diet. In another study, monkeys without carotenoids and omega-3 fatty acids developed some RPE atrophy [17]. In fact, the AREDS2 study found that human subjects in the bottom quartile of nutrition benefited the most from vitamin supplementation [42].

#### *2.6.1 Primate models for early onset drusen*

Some NHP species such as rhesus macaque monkeys spontaneously develop drusen and early to intermediate AMD. The amount of drusen increased with age [43]. Drusen analyzed in these monkeys were found by immunohistochemical analysis to have similar location and composition as human drusen, sharing compounds such as apolipoprotein E, amyloid P component, complement components, immunoglobulins, vitronectin, membrane cofactor protein, annexins, and crystallins [17, 28].

#### *An Overview of Age-Related Macular Degeneration: Clinical, Pre-Clinical Animal Models… DOI: http://dx.doi.org/10.5772/intechopen.96601*

Interestingly, a group of cynomolgus macaques and Japanese macaques were found to have early-onset drusen in the macula and periphery at around 1–2 years of age. The drusen in these groups of monkeys were also similar in composition to human drusen. This syndrome exhibited a dominant inheritance which along with early onset drusen may serve as a useful animal model for future studies [17].
