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DYSREGULATION OF THE (IMMUNO)PROTEASOME PATHWAY IN MCD
included patients with mild degree of cortical dysplasia (mMCDs; 33). Age at surgery, seizure duration and seizure frequency were not statistically different between patients with FCD II and mMCD in this cohort, as well as between FCD IIa and FCD IIb cases included in our cohort. Accordingly to the international consensus classification sys- tem of FCD 21, FCD II represents isolated focal lesions with architectural and dysmorphic abnormalities (FCD IIa with dysmorphic neurons only; FCD IIb with dysmorphic neurons and balloon cells; Fig. 1-4 C-E). TSC patients were younger compared to mMCD and FCD patients. All six TSC-tubers displayed similar histopathological features, including loss of lamination, astrogliosis, dysmorphic neurons and giant cells with pale eosinophilic cyto- plasm (34; Fig. 1-4 F-G).
Proteasome subunit expression in FCD and cortical tubers
Expression of β1 β1i, β5 and β5i was observed in FCD, cortical tubers and mMCD speci- mens (Fig. 1-4; supplementary Fig. 3). We observed differences in the expression level, as well as in the cell-specific and subcellular distribution of the different subunits (Table 3).
Constitutive proteasome catalytic subunit β1 and β5
Moderate expression of β1 and β5 subunit was observed in human control cortical spec- imens (Fig. 1 A-B; Fig. 3 A-B; Table 3). Nuclear neuronal expression was detected for β5 in MCD specimens (Table 3; supplementary Fig. 3 E; whereas only cytoplasm expression was detected in specimens from patients with Alzheimer’s disease for both subunits (supple- mentary Fig. 3 B, F). Increased expression of both constitutive subunits was observed in FCD and TSC specimens (Fig. 1 C-G; Fig. 3 C-G; Table 2). In the large majority of FCD and TSC cases β1 IR was detected in cytoplasm and nucleus of neuronal and glial cells (Fig. 1 C-G; Table 3). β1 was also detected in balloon (FCD IIb; Fig. 1 E) and giant cells (TSC; Fig. 1G). FCD and TSC specimens displayed also strong β5 IR with prominent nuclear expression in both neuronal and glial cells, as well as in balloon (FCD IIb) and giant cells (TSC; Fig. 3 C-G; Table 3). A similar pattern was detected in the post-mortem TSC case; double-labeling experiments confirmed the co-localization with astroglial and neuronal markers within the dysplastic area for both subunits in FCD and TSC specimens (Fig. 1 F-G; Fig. 3 G).
Immunoproteasome subunits β1i and β5i
In the large majority of control (Fig. 2 A-B; Fig. 4 A-B) and mMCD (supplementary Fig. 3 C, G) specimens the immunoproteasome subunits β1i and β5i were under the detection levels in both neuronal and glial cells (Table 3). β1i and β5i were consistently high in FCD and TSC specimens (Table 3) with strong cytoplasmic and nuclear IR in neuronal and glial cells, in both surgical post-mortem TSC specimens (whereas only cytoplas- mic expression was detected in glial cells in specimens from patients with Alzheimer’s disease for both β1i and β5i subunits supplementary Fig. 3 D, H). A similar pattern with strong expression in FCD and TSC specimens was observed using in situ hybridization (supplementary Fig. 4). Double-labeling experiments confirmed the co-localization with astroglial and neuronal markers, as well as with major histocompatibility complex (MHC) class I (HLA-I; in few balloon/giant cells and in dysmorphic neurons) within the dysplas- tic area for both subunits in FCD and TSC specimens (Fig. 2 and 4 F-G). In regions with
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