Seladin-1/DHCR24 Is Neuroprotective by Associating EAAT2 Glutamate Transporter to Lipid Rafts in Experimental Stroke

Abstract

Background and Purpose-3β-Hydroxysteroid-Δ24 reductase (DHCR24) or selective alzheimer disease indicator 1 (seladin-1), an enzyme of cholesterol biosynthetic pathway, has been implicated in neuroprotection, oxidative stress, and inflammation. However, its role in ischemic stroke remains unexplored. The aim of this study was to characterize the effect of seladin-1/DHCR24 using an experimental stroke model in mice. Methods-Dhcr24+/− and wild-type (WT) mice were subjected to permanent middle cerebral artery occlusion. In another set of experiments, WT mice were treated intraperitoneally either with vehicle or U18666A (seladin-1/DHCR24 inhibitor, 10 mg/kg) 30 minutes after middle cerebral artery occlusion. Brains were removed 48 h after middle cerebral artery occlusion for infarct volume determination. For protein expression determination, peri-infarct region was obtained 24 h after ischemia, and Western blot or cytometric bead array was performed. Results-Dhcr24+/− mice displayed larger infarct volumes after middle cerebral artery occlusion than their WT littermates. Treatment of WT mice with the seladin-1/DHCR24 inhibitor U18666A also increased ischemic lesion. Inflammationrelated mediators were increased after ischemia in Dhcr24 +/− mice compared with WT counterparts. Consistent with a role of cholesterol in proper function of glutamate transporter EAAT2 in membrane lipid rafts, we found a decreased association of EAAT2 with lipid rafts after ischemia when DHCR24 is genetically deleted or pharmacologically inhibited. Accordingly, treatment with U18666A decreases

DHCR24 exerts neuroprotection upon inflammation-induced neuronal death

“…Finally, elucidation of the expression profile of TNFα, BDNF, HMOX1, and NQO1 did not reveal alterations in mice overexpressing DHCR24 as compared to control mice, suggesting that DHCR24 is able to locally mitigate the ischemia-induced damage in a mouse model of transient focal cerebral ischemia without significantly affecting the inflammatory, neurotrophic, or oxidative stress responses. Collectively, the findings in the present study are in line with a previous study showing that DHCR24 exerts cholesterol-dependent neuroprotection in an experimental stroke model in mice, in which DHCR24 was genetically downregulated [ 49 ]. More specifically, it was suggested that the underlying molecular mechanism of DHCR24 is linked to the maintenance of lipid raft integrity in astrocytes by assuring the EAAT2-mediated uptake of glutamate excess upon ischemic stress [ 49 ].…”

“…Collectively, the findings in the present study are in line with a previous study showing that DHCR24 exerts cholesterol-dependent neuroprotection in an experimental stroke model in mice, in which DHCR24 was genetically downregulated [ 49 ]. More specifically, it was suggested that the underlying molecular mechanism of DHCR24 is linked to the maintenance of lipid raft integrity in astrocytes by assuring the EAAT2-mediated uptake of glutamate excess upon ischemic stress [ 49 ]. This is an interesting mechanistic finding also in the context of the present study, as we observed that the number of astrocytes significantly increased after the addition of BV2 microglial cells to the neuronal cultures before the induction of neuroinflammation.…”

Molecular analysis of NPAS3 functional domains and variants

“…The rare psychiatric illness-associated p.Val304Ile is localized between the PAS domains of NPAS3 and has been shown to be associated with aggregation, further, it can be hypothesized to affect protein interactivity and function (Fig. 1 a, [ 40 ]). Although our assays were not designed to assess protein aggregation, we did not observe alterations in expression, localization, deficits in interactivity with ARNT or transactivation function due to this variant in our assays (Figs.…”

Aging Increases Hippocampal DUSP2 by a Membrane Cholesterol Loss-Mediated RTK/p38MAPK Activation Mechanism

“…There are also several examples where brain inflammation, in which p38MAPK has a preponderant role, has been associated to the loss of neuronal cholesterol that occurs both in conditions of acute (e.g., stroke) and chronic (aging) inflammation (21, 22). Therefore, we decided to investigate the relationship between p38MAPK increase and neuronal cholesterol loss.…”