α-Lipoic acid (LA) enantiomers protect SH-SY5Y cells against glutathione depletion

Growing evidence suggests that α-lipoic acid (LA) has neuroprotective effects in various pathological conditions including brain ischemia and neurodegeneration. While anti-oxidative activity has been thought to play a central role in LA-mediated neuroprotection, the precise mechanism and the effect of LA enantiomers (R- and S-LA) are not fully clarified. We, therefore, estimated the neuroprotective effects of LA against different cellular stresses including oxidative stress, endoplasmic reticulum (ER) stress and proteolytic stress using human neuroblastoma SH-SY5Y cells. All types of LAs (racemate, R-LA and S-LA) most effectively prevented cell death induced by buthionine sulfoximine (BSO) which depletes intracellular glutathione. Although direct effects of LA on glutathione depletion or generation of the reactive oxygen species (ROS) were relatively small upon BSO treatment, LA enhanced expressions of anti-oxidative genes such as heme oxygenase-1 (HO-1) and phase II detoxification enzymes such as NAD(P)H:Quinone Oxidoreductase 1 (NQO1). An inhibitor of NQO1, but not that of HO-1, suppressed LA-mediated protection against BSO. Further experiments revealed that all types of LAs activated cell survival-associated kinase Akt, and an inhibitor of PI3K, LY294002, suppressed both LA-induced upregulation of NQO1 and cell protection against BSO. Our results suggest an important role of PI3K/Akt-mediated upregulation of genes including phase II enzymes such as NQO1 in LA-mediated neuroprotection. © 2011 Elsevier B.V

Deletion of N-myc downstream-regulated gene 2 attenuates reactive astrogliosis and inflammatory response in a mouse model of cortical stab injury

N-myc downstream-regulated gene 2 (Ndrg2) is a differentiation- and stress-associated molecule predominantly expressed in astrocytes in the CNS. In this study, we examined the expression and the role of Ndrg2 after cortical stab injury. We observed that Ndrg2 expression was elevated in astrocytes surrounding the wounded area as early as day 1 after injury in wild-type mice. Deletion of Ndrg2 resulted in lower induction of reactive astroglial and microglial markers in the injured cortex. Histological analysis showed reduced levels of hypertrophic changes in astrocytes, accumulation of microglia, and neuronal death in Ndrg2-/- mice after injury. Furthermore, activation of the IL-6/signal transducer and activator of transcription 3 (STAT3) pathway, including the expression of IL-6 family cytokines and phosphorylation of STAT3, was markedly reduced in Ndrg2-/- mice after injury. In a culture system, both of Il6 and Gfap were up-regulated in wild-type astrocytes treated with forskolin. Deletion of Ndrg2 attenuated induction of these genes, but did not alter proliferation or migration of astrocytes. Adenovirus-mediated reexpression of Ndrg2 rescued the reduction of IL-6 expression after forskolin stimulation. These findings suggest that Ndrg2 plays a key role in reactive astrogliosis after cortical stab injury through a mechanism involving the positive regulation of IL-6/STAT3 signaling. © 2014 International Society for Neurochemistry

The effect of Ndrg2 expression on astroglial activation

N-myc downstream-regulated gene 2 (Ndrg2) is a differentiation- and stress-associated molecule predominantly expressed in astrocytes in the central nervous system (CNS). To study the expression and possible role of Ndrg2 in quiescent and activated astrocytes, mice were administrated with 1-methyl-4-phenyl-1,2,3,6-tetrahydropypridine (MPTP), a Parkinson disease (PD)-related neurotoxin which causes both neurodegeneration and glial activation. Immunohistological analysis revealed that Ndrg2 was highly expressed in both types of astrocytes, but less so in astrocytes during the early process of activation. Ndrg2 was also expressed in astrocyte-like cells, but not in neurons, in human brains from PD and Cortico-basal degeneration (CBD) patients. In cultured astrocytes, gene silencing of Ndrg2 significantly enhanced the numbers of 5-bromo-2′-deoxy-uridine (BrdU)-incorporated and proliferating cell nuclear antigen (PCNA)-positive cells, and reduced the length of cell processes and the amount of F-actin. In contrast, adenovirus-mediated overexpression of Ndrg2 significantly reduced the numbers of BrdU-incorporated and PCNA-positive cells, and enhanced the amount of F-actin. Fractionation and immunocytochemical analysis further revealed that Ndrg2 was located in different cellular fractions including the cytosol and cell surface membranes. These results suggest that Ndrg2 may regulate astroglial activation through the suppression of cell proliferation and stabilization of cell morphology. © 2011 Elsevier Ltd. All rights reserved

The ATF6β-calreticulin axis promotes neuronal survival under endoplasmic reticulum stress and excitotoxicity

While ATF6α plays a central role in the endoplasmic reticulum (ER) stress response, the function of its paralogue ATF6β remains elusive, especially in the central nervous system (CNS). Here, we demonstrate that ATF6β is highly expressed in the hippocampus of the brain, and specifically regulates the expression of calreticulin (CRT), a molecular chaperone in the ER with a high Ca2+-binding capacity. CRT expression was reduced to ~ 50% in the CNS of Atf6b−/− mice under both normal and ER stress conditions. Analysis using cultured hippocampal neurons revealed that ATF6β deficiency reduced Ca2+ stores in the ER and enhanced ER stress-induced death. The higher levels of death in Atf6b−/− neurons were recovered by ATF6β and CRT overexpressions, or by treatment with Ca2+-modulating reagents such as BAPTA-AM and 2-APB, and with an ER stress inhibitor salubrinal. In vivo, kainate-induced neuronal death was enhanced in the hippocampi of Atf6b−/− and Calr+/− mice, and restored by administration of 2-APB and salubrinal. These results suggest that the ATF6β-CRT axis promotes neuronal survival under ER stress and excitotoxity by improving intracellular Ca2+ homeostasis

The ATF6β-calreticulin axis promotes neuronal survival under endoplasmic reticulum stress and excitotoxicity

神経細胞死を抑制する新たな分子を発見 --脳卒中やアルツハイマー病への応用に期待--. 京都大学プレスリリース. 2021-06-30.While ATF6α plays a central role in the endoplasmic reticulum (ER) stress response, the function of its paralogue ATF6β remains elusive, especially in the central nervous system (CNS). Here, we demonstrate that ATF6β is highly expressed in the hippocampus of the brain, and specifically regulates the expression of calreticulin (CRT), a molecular chaperone in the ER with a high Ca²⁺-binding capacity. CRT expression was reduced to ~ 50% in the CNS of Atf6b⁻/⁻ mice under both normal and ER stress conditions. Analysis using cultured hippocampal neurons revealed that ATF6β deficiency reduced Ca²⁺ stores in the ER and enhanced ER stress-induced death. The higher levels of death in Atf6b⁻/⁻ neurons were recovered by ATF6β and CRT overexpressions, or by treatment with Ca²⁺-modulating reagents such as BAPTA-AM and 2-APB, and with an ER stress inhibitor salubrinal. In vivo, kainate-induced neuronal death was enhanced in the hippocampi of Atf6b⁻/⁻ and Calr⁺/⁻ mice, and restored by administration of 2-APB and salubrinal. These results suggest that the ATF6β-CRT axis promotes neuronal survival under ER stress and excitotoxity by improving intracellular Ca²⁺ homeostasis

Deletion of Herpud1 Enhances Heme Oxygenase-1 Expression in a Mouse Model of Parkinson’s Disease

Herp is an endoplasmic reticulum- (ER-) resident membrane protein that plays a role in ER-associated degradation. We studied the expression of Herp and its effect on neurodegeneration in a mouse model of Parkinson’s disease (PD), in which both the oxidative stress and the ER stress are evoked. Eight hours after administering a PD-related neurotoxin, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), to mice, the expression of Herp increased at both the mRNA and the protein levels. Experiments using Herpud1+/+ and Herpud1−/− mice revealed that the status of acute degeneration of nigrostriatal neurons and reactive astrogliosis was comparable between two genotypes after MPTP injection. However, the expression of a potent antioxidant, heme oxygenase-1 (HO-1), was detected to a higher degree in the astrocytes of Herpud1−/− mice than in the astrocytes of Herpud1+/+ mice 24 h after MPTP administration. Further experiments using cultured astrocytes revealed that the stress response against MPP+, an active form of MPTP, and hydrogen peroxide, both of which cause oxidative stress, was comparable between the two genotypes. These results suggest that deletion of Herpud1 may cause a slightly higher level of initial damage in the nigrastrial neurons after MPTP administration but is compensated for by higher induction of antioxidants such as HO-1 in astrocytes