Immunomodulation with minocycline rescues retinal degeneration in juvenile neuronal ceroid lipofuscinosis mice highly susceptible to light damage

ABSTRACT Juvenile neuronal ceroid lipofuscinosis (jNCL) is a rare but fatal inherited lysosomal storage disorder mainly affecting children. The disease is caused by mutations in the CLN3 gene that lead to the accumulation of storage material in many tissues, prominent immune responses and neuronal degeneration. One of the first symptoms is vision loss followed by motor dysfunction and mental decline. The established Cln3Δex7/8 mouse model mimics many pathological features of the human disease except the retinal phenotype, which is very mild and occurs only very late in these mice. Here, we first carefully analyzed the retinal structure and microglia responses in these animals. While prominent autofluorescent spots were present in the fundus, only a moderate reduction of retinal thickness and no prominent microgliosis was seen in young CLN3-deficient mice. We next genetically introduced a light-sensitive RPE65 variant and established a light-damage paradigm that showed a high susceptibility of young Cln3Δex7/8 mice after exposure to 10,000 lux bright light for 30 min. Under these ‘low light’ conditions, CLN3-deficient mice showed a strong retinal degeneration, microglial activation, deposition of autofluorescent material and transcriptomic changes compared to wild-type animals. Finally, we treated the light-exposed Cln3Δex7/8 animals with the immunomodulatory compound minocycline, and thereby rescued the retinal phenotype and diminished microgliosis. Our findings indicate that exposure to specific light conditions accelerates CLN3-dependent retinal degeneration, and that immunomodulation by minocycline could be a possible treatment option to delay vision loss in jNCL patients. This article has an associated First Person interview with the first author of the paper.

Juvenile neuronal ceroid lipofuscinosis ( jNCL) is a rare but very severe disease affecting children at school age. Beginning with blindness, those kids lose previously acquired skills, exhibit seizures and become bedridden until they die in their late 20s. The reason for these symptoms is a malfunction of CLN3, a protein of so far unknown function leading to neuronal cell death in the retina and brain. Neuronal cell death is usually accompanied by a strong activation of the immune system, leading to additional neuronal damage. The most frequently used jNCL mouse model, carrying the same defect as found in patients, develops the neuronal phenotype relatively late in age. Therefore, we exposed the mice to bright white light to accelerate the retinal immune response. Our results show that microglia, the resident cells of the immune system in the CNS, are more aggressive when CLN3 is deficient, leading to massive cell loss. Our findings also show that treating those light-exposed CLN3deficient mice using an immunomodulatory compound can reduce microglial activity and thus reduce retinal cell death.
What are the potential implications of these results for your field of research?
Our modified jNCL mouse model is useful to analyze the retinal pathomechanisms of jNCL and to find possible treatment strategies in young jNCL mice. Furthermore, our results show that microglia are a possible target to modulate the immune response in neurodegenerative diseases. Therefore, the results indicate that the extent of microglial damage can also be limited in other neurodegenerative diseases of the brain and retina by administering immunomodulatory compounds.
What are the main advantages and drawbacks of the model system you have used as it relates to the disease you are investigating?
For our research we used a well-established mouse model, carrying the same genetic defect as most jNCL patients. Despite the relatively late occurrence of the retinal immune response and neuronal cell death, the hallmarks of the human pathology are shown to occur in the mouse model. Although the exact role of the CLN3 protein is unclear so far, experiments with other model organisms suggest a complex function and interaction of CLN3 with other proteins. Therefore, a highly organized tissue like the mouse retina is most suitable to analyze influenced cell morphology, function and cellto-cell communication. The major drawback of the jNCL mouse model is that, in contrast to human patients, the retina is affected relatively late in age, with a comparably lower severity of microglial activation. Additionally, mice are complex organisms and, just like humans, the extent of a pathology can vary more widely compared to simple model organisms.

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we used a well-established mouse model, carrying the same genetic defect as most jNCL patients. Despite the relatively late occurrence of the retinal immune response and neuronal cell death, the hallmarks of the human pathology are shown to occur in the mouse model." Describe what you think is the most significant challenge impacting your research at this time and how will this be addressed over the next 10 years?
Definitively the most significant challenge of jNCL research is to find an appropriate antibody against CLN3. So far, no specific commercial CLN3 antibody is available, leading to the exclusion of a lot of experiments necessary to analyze the localization and detailed function of the CLN3 protein. Hopefully, feasible strategies to generate antibodies against hydrophobic proteins that are only present in small amounts will arise. A great company with decades of immunization and antibody generation experience, like the Belgian company Eurogentec, might be able to provide such a much-needed antibody and heal the world from jNCL for good. Furthermore, there is little financial support for jNCL-related research projects. Luckily, I was very lucky to have had a very supportive PI, giving me the chance to conduct a lot of autonomous work and projects. I've learned a lot about writing, collaborative projects, presenting my findings and networking. If embedded in a good graduate school, students at the PhD level can effectively acquire important soft skills useful for their future careers. At some universities, the structure of graduate schools is in the process of reorganization, resulting in maximum support and preparation for early-career scientists.