Characterization of the retinal degeneration and glial activation of neuronal ceroid lipofuscinosis mouse models Cln3Δex7-8 and Cln6nclf and the beneficial effects of dietary supplementation

Microglia are specialized macrophages of the central nervous system involved in immune regulation, tissue development, homeostasis and wound repair. In healthy tissue, microglia are found in a ramified state which is tightly controlled by neurons. Upon insult, microglia undergo morphological changes, resembling active phagocytic macrophages and are highly motile. In neurodegenerative diseases, such are Alzheimer's or multiple sclerosis and in most inherited retinal dystrophies, microglia cells are found in a permanent pro-inflammatory active state which exacerbates cell death and disease progression. Similarly, astrocytes in the brain or Müller cells in the retina, also become reactive in diseased tissue and may contribute to disease progression through overshoot release of protective factors such as VEGF (vascular endothelial growth factor), massive proliferation and release of inflammatory intermediates which recruit microglia to site of injury. Along with active microglia, reactive astrocytes/Müller cells are also found in virtually all neurodegenerative and retinal dystrophies. Microglia and Müller cell status has not been well defined in retinal degeneration of neurodegenerative disease neuronal ceroid lipofuscinosis (NCL). The NCLs are early onset lysosomal storage disorders characterized by vision loss, mental and motor deficits, and spontaneous seizures. Massive accumulation of auto-fluorescent material in neurons lead to progressive neuronal degeneration and cell death Furthermore, neuropathological analyses of autopsy material from NCL patients and animal models revealed brain atrophy closely associated with microglial and astrocyte activity. However, it is unclear whether this phenomenon is mainly confined to the brain or also occurs in the retina Two mouse models of NCL, Cln3Δex7-8 and Cln6nclf, were selected for detailed characterization of retinal degeneration with focus on microglia and Müller cell activity. Studies showed that Cln3Δex7-8 mice had a late onset of retinal degeneration which started at 9 months and progressed until severe retinal impairment at 18 months. Reactive Müller cells were present starting at six months with no significant microglial activation until late disease stage. Furthermore, degeneration originated from inner retinal cells as seen through electroretinograms with little photoreceptor death. In contrast, temporal studies on Cln6nclf mice showed an early progressive retinal degenerative phenotype which started before one month of age resulting in loss of vision and retinal function by eight months. Full retinal degeneration was accompanied by reactive Müller cells already present at one month of age and fully active microglia at four months. Mixed population of ramified and active microglia were seen at one month. Transcript profiling of the Cln6ncl mouse retina also showed up-regulated expression of apoptotic, inflammatory and active microglia markers. Based on the strong reactivity of microglia, Cln6nclf mice were supplemented with immuno-modulatory compounds curcumin, luteolin or DHA in order to ameliorate microgliosis and reduce retinal degeneration. Studies showed Cln6nclf mice supplemented with 0.6% curcumin, 0.6% luteolin or 5% DHA had improved retinal function, better visual acuity and DHA-supplementation had the greatest preservation of photoreceptors. All supplements were able to attenuate microglia activation as seen by the presence of microglial ramification and intact network. However, curcumin and luteolin supplementation did not have any effect on retinal transcriptom whilst DHA-supplementation reduced expression of inflammatory and apoptotic markers. Taken together, microglial reactivity accompanies disease progression in the Cln6nclf retina but not Cln3Δex7-8. Immuno-modulating compounds were able to attenuate disease progression and reactive microglia to a certain extent in Cln6nclf mice, indicating modulation of microglial activity could be helpful in preserving vision and may ameliorate neuronal degeneration in NCL. Finally, inspection of the retinal function may also allow to monitor general disease progression and could be used to assess the efficacy of therapeutic interventions for NCL patients