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Animal Models of Exfoliation Syndrome, Now and Future

John, Simon W.M. PhD*,†,‡; Harder, Jeffrey M. PhD; Fingert, John H. MD, PhD§; Anderson, Michael G. PhD§,∥,¶

doi: 10.1097/IJG.0000000000000121

At present, no animal models fully embody exfoliation syndrome or exfoliation glaucoma. Both genetic and environmental factors appear critical for disease manifestation, and both must be considered when generating animal models. Because mice provide a powerful mammalian platform for modeling complex disease, this paper focuses on mouse models of exfoliation syndrome and exfoliation glaucoma.

*The Howard Hughes Medical Institute

The Jackson Laboratory, Bar Harbor, ME

Department of Ophthalmology, Tufts University School of Medicine, Boston, MA

Departments of §Ophthalmology and Visual Sciences

Molecular Physiology and Biophysics, University of Iowa

Center for the Prevention and Treatment of Visual Loss, Iowa City VA Health Care System, Iowa City, IA

Disclosure: The authors declare no conflict of interest.

Reprints: Simon W.M. John, PhD, The Jackson Laboratory, 600 Main St., Bar Harbor 04609, Maine (e-mail:

Received August 11, 2014

Accepted August 11, 2014

Exfoliation syndrome (XFS) is the most common identifiable cause of open-angle glaucoma. Despite its prevalence, little is understood about its molecular etiology or about the factors determining susceptibility and progression to exfoliation glaucoma (XFG). It is clear that XFS is a complex, age-related disease affected by both genetic and environmental factors. Nevertheless, both the genetic and environmental factors need better definition.1–4 A strong association between single-nucleotide polymorphisms (SNPs) in the lysyl oxidase–like 1 (LOXL1) gene and XFS was identified in the Swedish and Icelandic populations using a genome-wide association study.5 This association was replicated in other populations.6–22 However, risk alleles of LOXL1 are very common in unaffected controls, and the mechanisms of action of these alleles are not clear.7,23 Risk alleles at contactin-associated protein-like 2 (CNTNAP2) loci also have significant associations between XFS and XFG in some but not all populations.24–26 Mutations in CNTNAP2 cause neurologic disease, and it is not clear if allelic, genetic background or other differences modulate the phenotypic consequences of mutation in this gene. A host of environmental factors have been suggested to influence XFS, but findings are often inconsistent across studies.3,4

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To understand the molecular mechanisms, underlying XFS and progression to XFG tractable animal models are needed. Extremely few publications have studied animal models of XFS, and to our knowledge only 2 animal models have been reported.27,28 There are no reports of models with all of the features of XFS with XFG. The lack of animal models has been suggested to reflect the typical occurrence of XFS at old ages, with the belief that model species do not live long enough to develop the condition. Although possible, we do not feel that the shorter lifespan of model species in absolute years prevents development of XFS. Disease susceptibility in model species increases with age relative to lifespan in a similar manner to that in people.29 For example, within a 2 years life span, mice often develop complex, age-related diseases that occur later in human life. In contrast, differences in environment may be critical and profoundly impact whether or not model species develop XFS. The degree and nature of exposure to light (UV), low temperature, viruses, and caffeine have all been suggested to impact development of XFS.4,30–33 Most laboratory animals are housed in cages with limited UV exposure, constant controlled temperature, limited or no exposure to pathogens, and lack of caffeine and other lifestyle factors. Genetic differences across species may also be important. In humans a high-risk genotype at the LOXL1 locus is typically necessary for manifestation of XFS, as high-risk alleles are present in almost all patients with XFS.23 Thus, the challenge of modeling XFS/XFG in animals is to develop models reflecting these complex genetic and environmental risk factors.

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Porcine Model

The first animal model was reported in pigs.27 The authors fed pigs a high sucrose, high salt diet to induce cataracts. They reasoned that cataracts are common in XFS and that mature cataracts shed exfoliative material. After a few months on the diet the pigs developed cataracts and had an exfoliation-like material that contained crystallins. We are not aware of any follow-up studies. The relevance of this model to the human disease is not yet clear, although it may caution against high salt and sugar intakes.

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Lyst Mutant Mouse Model

The other model is an inherited mouse model, which shares several features with human XFS. In addition to the accumulation of fibrillar material in the eye, patients with XFS have characteristic saw-tooth morphology of the iris pigment epithelium.34,35 This results in iris transillumination defects characterized by a specific concentric, circular transillumination pattern.36 Similar to human patients, Lyst mutant mice have microscopically detectable deposits of fibrillin 1-positive material in the eye.28 They also replicate the saw-tooth morphology of the iris pigment epithelium and have the same pattern of transillumination defects as human patients (Fig. 1). In human XFS, increased susceptibility to oxidative stress has been suggested to contribute to the pathology37,38 and, as in other glaucomas, levels of transforming growth factor-β(TGF-β) superfamily members are elevated.39–42 The Lyst mutant iris disease involves oxidative damage43 and TGF-β levels in Lyst mutant eyes remain to be tested. Thus, the mice have some XFS-like phenotypes but lack clinically obvious XFS deposits or glaucoma. Although the degree of relevance of this mouse to human XFS is not yet clear, it is currently the most similar available model. Further evaluation of this model will be important. Similarly, evaluation of the LYST gene and functionally related genes in human patients is worthwhile.



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Future Models

The housing environment may critically interact with genetics to determine if model species develop XFS. Thus, environment should be considered and manipulated when working to produce new models of XFS. Among other factors, the degree and nature of exposure to light (UV), low temperature, presence of specific viruses/microbes, dietary composition, and caffeine intake may impact the development of XFS.4,30–33,44 Using animal models, the importance of specific environmental factors could be clearly determined. It is now possible to abrogate or greatly decrease the amount of gene product produced by mutating genes in various species and a variety of species may contribute to improved understanding of XFS. Because of their small size, high fecundity, relatively low cost, and the most powerful array of available genetic resources and tools for dissecting complex diseases and humanizing their genomes, mice will remain a very important model species.45,46 Mice are likely to add substantially to our mechanistic understanding of XFS (Fig. 2).



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As the LOXL1 genotype is critically important in the development of XFS, an important step in producing a new mouse model of XFS is to make mice with variant forms of the Loxl1 gene. Mice with a completely nonfunctional allele of Loxl1 have an elastic tissue disease47 but do not develop XFS.48,49 Although it is worth aging and assessing these mice in different environments, it remains possible that a null allele will not cause XFS. As the specific DNA change(s) that render susceptibility to XFS are not clearly defined and may be intronic, making mice that are transgenic for a human bacterial artificial chromosome (BAC) containing a high-risk human allele of LOXL1 is a high priority. If multiple high-risk alleles are defined from different populations, mice can be made with alleles conferring different degrees of risk with initial bias toward those with the strongest effect. Further, it will be important to evaluate multiple transgenic mouse lines with different expression levels.

LOXL1 genotype alone is unlikely to be sufficient to induce XFS, as many individuals with a high-risk allele do not have the disease. Thus, it will be important both to combine the human LOXL1 transgene with mutations in other genes (such as Lyst) and to assess the effects of different environments. Decreased antioxidant capacity and increased propensity to inflammation may be important in XFS and XFG.38,50,51 Mutations that alter these systems can be introduced into the transgenic mice by breeding. Altered TGF-β signaling has been implicated in XFS, but transgenic or knockout mice affecting this pathway have not been demonstrated to develop XFS. Combining human LOXL1 transgenes with mutations affecting TGF-β pathways may prove valuable, especially for the development of high IOP and glaucoma.52–55 Although less clearly important, mutations in CNTNAP2 are also implicated in XFS and it is worth assessing mice with CNTNAP2 mutations and humanizing mice with human high-risk alleles.

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Various efforts to identify XFS genes are underway, including genome-wide association studies and genetic studies in human families. Because of the complexity of the disease these studies are not easy, but once more genes are identified they can be assessed in mice using the approaches discussed above. Genetic studies in families with many affected individuals may have the highest likelihood of success and are very important. Mutagenesis screens in mice are another powerful approach for providing animal models and discovering disease mechanism.46,56–59 With adequate aging and attention to environment, such screens may provide key new models of XFS and XFG. One important strategy for discovering XFS genes and pathways would be to perform a mutagenesis screen using mice that are transgenic for human high-risk alleles of LOXL1. Modifier screens to identify genetic differences that alter phenotypes caused by the LOXL1 transgene (or other human alleles) between mouse strains may prove valuable.60–63

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exfoliation syndrome; animal models; knockout mice; transgenic mice; alleles

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