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DYSREGULATION OF THE (IMMUNO)PROTEASOME PATHWAY IN MCD
Introduction
The proteasome is an evolutionarily conserved multicatalytic proteinase complex rep- resenting a major protein degradation system, present in the nucleus and cytoplasm of eukaryotic cells, that regulates diverse biological processes essential for cell survival 1-4. The proteolytic complex of the proteasome is represented by a catalytic 20S core parti- cle, a barrel-shaped complex consisting of four heptameric rings, composed of non-iden- tical α or β subunits 2, 5, 6. The catalytic activity is restricted to three of the beta subunits β1, β2 and β5, which have specific cleavage preferences, and are constitutively expressed in most tissues. Under certain conditions associated with the release of immune-mod- ulatory cytokines (i.e. interferon- γ, IFN-γ), a specialized type of proteasome called the immunoproteasome can be generated by the incorporation of the inducible subunits, β1i (PSMB9; LMP2, low molecular weight protein 2), β2i (PSMB10; LMP10 MECL-1, mul- ticatalytic endopeptidase complex-like1), and β5i (PSMB8; LMP7, low molecular weight protein 7) 7, 8. Increasing evidence supports a key role of the immunoproteasome in the regulation of immune cell function, including both the adaptive and the innate immune response 9-11. A deregulation of the immunoproteasome system, with induction of β1i and β5i subunits in neurons and/or glial cells, has been reported in neurodegenerative diseases (12-14 for review see 15, 16). In particular, recent studies have pointed to the role of immunoproteasome in glial cells, suggesting an novel interaction between immuno- proteasome and glia-mediated inflammatory response, resulting in a pro-inflammatory environment 14, 16. Interestingly, induction of β1i and β5i subunits has been also observed in specimens of patients with pharmaco-resistant mesial temporal lobe epilepsy (MTLE; 17). Moreover, recent experimental data support a role for the β5i subunit in modulating seizure generation in epileptic tissue and interestingly this subunit was not up-regulated in rats exposed to pilocarpine but not developing SE and spontaneous seizures 18.
In the present study, we investigated the expression and cellular distribution of both constitutive (β1, β5) and immunoproteasome (β1i, β5i) subunits using immunohis- tochemistry in a large cohort of patients with malformations of cortical development (MCD; including focal cortical dysplasia, FCD type II and Tuberous Sclerosis Complex, TSC cortical tubers), evaluating a possible relationship between changes in expression of these subunits and the clinical course of epilepsy. To provide better insights into the mechanisms underlying the astroglial regulation of immunoproteasome subunits, we studied their expression in response to IL-1β stimulation in both human fetal astrocytes and FCD-derived cells. Since both FCD II and TSC are associated with constitutive acti- vation of the mammalian target of rapamycin pathway (mTOR pathway) 19, 20 we fur- ther evaluated the effect of rapamycin (inhibitor of the mammalian target of rapamycin, mTOR pathway, mTOR) in FCD II derived cell cultures.
Materials and Methods
Subjects
The cases included in this study were obtained from the archives of the departments of neuropathology of the Academic Medical Center (AMC, University of Amsterdam, The Netherlands), the University Medical Center Utrecht (UMCU, The Netherlands) and the Medical University Vienna (MUV, Austria). A total of 23 brain tissue specimens, removed from patients undergoing surgery for intractable epilepsy, were examined. Tissue was
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