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Mutations of the SPG4 ( SPAST ) gene encoding for spastin protein are the main causes of hereditary spastic paraplegia. Spastin binds to microtubules and severs them through the enzymatic activity of its AAA domain. Several missense mutations located in this domain lead to stable, nonsevering spastins that decorate a subset of microtubules, suggesting a possible negative gain-of-function mechanism for these mutants. Of the two main isoforms of spastin, only mutations of the long isoform, M1, are supposed to be involved in the onset of the pathology, leaving the role of the ubiquitously expressed shorter one, M87, not fully investigated and understood. Here, we show that two isoforms of spastin harboring the same missense mutation bind and bundle different subsets of microtubules in HeLa cells, and likely stabilize them by increasing the level of acetylated tubulin. However, only mutated M1 has the ability to interact with wild-type M1, and decorates a subset of perinuclear microtubules associated with the endoplasmic reticulum that display higher resistance to microtubule depolymerization and increased intracellular ionic strength, compared with those decorated by mutated M87. We further show that only mutated M1 decorates microtubules of proximal axons and dendrites, and strongly impairs axonal transport in cortical neurons through a mechanism likely independent of the microtubule-severing activity of this protein., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2018. Published by The Company of Biologists Ltd.)

Purpose: The role of cerebrovascular dysfunction in seizure disorders is recognized. Blood-brain barrier (BBB) damage in epilepsy has been linked to endothelial and glial pathophysiological changes. Little is known about the involvement of pericytes, a cell type that contributes to BBB function., Methods: NG2DsRed mice were used to visualize cerebrovascular pericytes. The pattern of vascular and parenchymal distributions of platelet-derived growth factor receptor beta (PDGFRβ) cells was evaluated by immunohistochemistry. Status epilepticus was induced in NG2DsRed or C57BL/6J mice by intraperitoneal kainic acid (KA). Animals were perfused intracardially using FITC-Dextran or FITC-Albumin to visualize the cerebrovasculature. Colocalization was performed between NG2DsRed, PDGFRβ and microglia IBA-1. Confocal 3D vessel reconstruction was used to visualize changes in cell morphology and position. PDGFRβ expression was also evaluated in vitro using organotypic hippocampal cultures (OHC) treated with kainic acid to induce seizure-like activity. Co-localization of PDGFRβ with the vascular marker RECA-1 and NG2 was performed. Finally, we assessed the expression of PDGFRβ in brain specimens obtained from a cohort of patients affected by drug resistant epilepsy compared to available autoptic brain., Results: In vivo, severe status epilepticus (SE) altered NG2DsRed vascular coverage. We found dishomogenous NG2DsRed perivascular ramifications after SE and compared to control. Concomitantly, PDGFRβ(+) cells re-distributed towards the cerebrovasculature after severe SE. Cerebrovascular NG2DsRed partially colocalized with PDGFRβ(+) while parenchymal PDGFRβ(+) cells did not colocalize with IBA-1(+) microglia. Using in vitro OHC we found decreased NG2 vascular staining and increased PDGFRβ(+) ramifications associated with RECA-1(+) microvessels after seizure-like activity. Cellular PDGFRβ and NG2(+) colocalization was observed in the parenchyma. Finally, analysis of human TLE brains revealed perivascular and parenchymal PDGFRβ(+) cell distributions resembling the murine in vivo and in vitro results. PDGFRβ(+) cells at the cerebrovasculature were more frequent in TLE brain tissues as compared to the autoptic control., Conclusions: The rearrangement of PDGFRβ(+) and vascular NG2DsRed cells after SE suggests a possible involvement of pericytes in the cerebrovascular modifications observed in epilepsy. The functional role of vascular-parenchymal PDGFRβ(+) cell redistribution and the relevance of a pericyte response to SE remain to be fully elucidated., (Copyright © 2014 Elsevier Inc. All rights reserved.)