Studio del ruolo di CDKL5 nella mielinizzazione della corteccia cerebrale mediante modelli di topo.

CDKL5 deficiency disorder (CDD) is a rare X-linked neurodevelopmental disorder, caused by mutations in the cyclin-dependent kinase-like 5 (CDKL5) gene. This disease affects mostly girls, with an incidence of 1.40.000/60.000 live births and it is characterized by a high heterogeneity of symptoms such as early onset epilepsy, severe cognitive deficits, cortical visual impairment and motor disabilities. CDKL5 is a serine-threonine kinase, highly expressed in neurons where it acts as a modulator of epigenetic factors and regulates cytoskeletal dynamics, dendritic and axonal outgrowth, and synaptogenesis. Although CDKL5 is also expressed by glial cell lineages, including myelinating oligodendroglial cells (OLs) very little is known on the role of this gene in these cellular populations. OLs are crucial for the process of myelination in the central nervous system, and emerging data are suggesting that grey matter myelination is important for the correct function and plasticity of neural circuits. Although the function of myelin in the grey matter is still under active investigation, its importance is highlighted by data showing that defects of myelination are often present in many neurological diseases including several neurodevelopmental disorders. In the context of CDD, MRI analyses showed evidences of cortical atrophy and white matter lesions in CDD patients that could arise from altered myelinating conditions. Despite this, it is still not known whether and how the lack of CDKL5 can impact the formation of the myelin sheath. To address this issue, I used Sudan Black B to stain myelin and investigate its organization in the brain of Cdkl5 mutant mice. This analysis revealed a decreased intensity in staining of most areas (i.e. corpus callosum, cerebral cortex and striatum) of both young and adult mutants compared to WT mice. Next, I evaluated the expression of myelin basic protein (MBP) using immunofluorescence and confocal microscopy, and the phosphorylation of neuro filaments (NF) by western blotting in the cerebral cortex. The results from these analyses pointed out a robust reduction of both MBP and NF phosphorylation in P15 and P56 Cdkl5-/y mice compared to their littermate controls. Finally, to understand if myelin/axonal abnormalities could arise from cell-autonomous or non-cell autonomous effects, I examined the expression of MBP in forebrain neuron-specific conditional Cdkl5-/y mice . The relative analysis showed that these conditional mutants largely recapitulate the decreased MBP expression observed in total Cdkl5 null mice, thus suggesting an important role of CDKL5 in neuronal-OLs communication that is crucial for myelination. In conclusion, this work provides evidence of aberrant myelination in primary cortical areas of mutant mice, revealing a novel role of CDKL5 in the myelination process. Furthermore, these data provide novel insight into CDD pathophysiology and support the putative role of myelination as a novel CDD biomarker.