Automated analysis of gray matter damage in aged mice reveals impaired remyelination in the cuprizone model.

Multiple sclerosis is a chronic autoimmune disease of the central nervous system characterized by myelin loss, axonal damage, and glial scar formation. Still, the underlying processes remain unclear, as numerous pathways and factors have been found to be involved in the development and progression of the disease. Therefore, it is of great importance to find suitable animal models as well as reliable methods for their precise and reproducible analysis. Here, we describe the impact of demyelination on clinically relevant gray matter regions of the hippocampus and cerebral cortex, using the previously established cuprizone model for aged mice. We could show that bioinformatic image analysis methods are not only suitable for quantification of cell populations, but also for the assessment of de- and remyelination processes, as numerous objective parameters can be considered for reproducible measurements. After cuprizone-induced demyelination, subsequent remyelination proceeded slowly and remained incomplete in all gray matter areas studied. There were regional differences in the number of mature oligodendrocytes during remyelination suggesting region-specific differences in the factors accounting for remyelination failure, as, even in the presence of oligodendrocytes, remyelination in the cortex was found to be impaired. Upon cuprizone administration, synaptic density and dendritic volume in the gray matter of aged mice decreased. The intensity of synaptophysin staining gradually restored during the subsequent remyelination phase, however the expression of MAP2 did not fully recover. Microgliosis persisted in the gray matter of aged animals throughout the remyelination period, whereas extensive astrogliosis was of short duration as compared to white matter structures. In conclusion, we demonstrate that the application of the cuprizone model in aged mice mimics the impaired regeneration ability seen in human pathogenesis more accurately than commonly used protocols with young mice and therefore provides an urgently needed animal model for the investigation of remyelination failure and remyelination-enhancing therapies.

Delayed Demyelination and Impaired Remyelination in Aged Mice in the Cuprizone Model.

To unravel the failure of remyelination in multiple sclerosis (MS) and to test promising remyelinating treatments, suitable animal models like the well-established cuprizone model are required. However, this model is only standardized in young mice. This does not represent the typical age of MS patients. Furthermore, remyelination is very fast in young mice, hindering the examination of effects of remyelination-promoting agents. Thus, there is the need for a better animal model to study remyelination. We therefore aimed to establish the cuprizone model in aged mice. 6-month-old C57BL6 mice were fed with different concentrations of cuprizone (0.2-0.6%) for 5-6.5 weeks. De- and remyelination in the medial and lateral parts of the corpus callosum were analyzed by immunohistochemistry. Feeding aged mice 0.4% cuprizone for 6.5 weeks resulted in the best and most reliable administration scheme with virtually complete demyelination of the corpus callosum. This was accompanied by a strong accumulation of microglia and near absolute loss of mature oligodendrocytes. Subsequent remyelination was initially robust but remained incomplete. The remyelination process in mature adult mice better represents the age of MS patients and offers a better model for the examination of regenerative therapies.

FoxP3 deficiency causes no inflammation or neurodegeneration in the murine brain.

Regulatory T cells (Treg) maintain immunological self-tolerance and their functional or numerical deficits are associated with progression of several neurological diseases. We examined the effects of Treg absence on the structure and integrity of the unchallenged murine brain. When compared to control, Treg-deficient FoxP3sf mutant mice showed no differences in brain size, myelin amount and oligodendrocyte numbers. FoxP3sf strain displayed no variations in quantity of neurons and astrocytes, whereas microglia numbers were slightly reduced. We demonstrate lack of neuroinflammation and parenchymal responses in the brains of Treg-deficient mice, suggesting a minor Treg role in absence of blood-brain barrier breakdown.