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Sephin1 Protects Neurons against Excitotoxicity Independently of the Integrated Stress Response.

ABSTRACT: Sephin1 is a derivative of guanabenz that inhibits the dephosphorylation of the eukaryotic initiation factor 2 alpha (eIF2?) and therefore may enhance the integrated stress response (ISR), an adaptive mechanism against different cellular stresses, such as accumulation of misfolded proteins. Unlike guanabenz, Sephin1 provides neuroprotection without adverse effects on the ?2-adrenergic system and therefore it is considered a promising pharmacological therapeutic tool. Here, we have studied the effects of Sephin1 on N-methyl-D-aspartic acid (NMDA) receptor signaling which may modulate the ISR and contribute to excitotoxic neuronal loss in several neurodegenerative conditions. Time-course analysis of peIF2? levels after NMDA receptor overactivation showed a delayed dephosphorylation that occurred in the absence of activating transcription factor 4 (ATF4) and therefore independently of the ISR, in contrast to that observed during endoplasmic reticulum (ER) stress induced by tunicamycin and thapsigargin. Similar to guanabenz, Sephin1 completely blocked NMDA-induced neuronal death and was ineffective against AMPA-induced excitotoxicity, whereas it did not protect from experimental ER stress. Interestingly, both guanabenz and Sephin1 partially but significantly reduced NMDA-induced cytosolic Ca2+ increase, leading to a complete inhibition of subsequent calpain activation. We conclude that Sephin1 and guanabenz share common strong anti-excitotoxic properties with therapeutic potential unrelated to the ISR.

SUBMITTER: Ruiz A

PROVIDER: S-EPMC7504470 | biostudies-literature | 2020 Aug

REPOSITORIES: biostudies-literature

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Sephin1 Protects Neurons against Excitotoxicity Independently of the Integrated Stress Response.

Ruiz Asier A Zuazo Jone J Ortiz-Sanz Carolina C Luchena Celia C Matute Carlos C Alberdi Elena E

International journal of molecular sciences 20200824 17

Sephin1 is a derivative of guanabenz that inhibits the dephosphorylation of the eukaryotic initiation factor 2 alpha (eIF2α) and therefore may enhance the integrated stress response (ISR), an adaptive mechanism against different cellular stresses, such as accumulation of misfolded proteins. Unlike guanabenz, Sephin1 provides neuroprotection without adverse effects on the α2-adrenergic system and therefore it is considered a promising pharmacological therapeutic tool. Here, we have studied the ef ...[more]

PMID: 32846985

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Project description:Multiple sclerosis is a chronic autoimmune demyelinating disorder of the CNS. Immune-mediated oligodendrocyte cell loss contributes to multiple sclerosis pathogenesis, such that oligodendrocyte-protective strategies represent a promising therapeutic approach. The integrated stress response, which is an innate cellular protective signalling pathway, reduces the cytotoxic impact of inflammation on oligodendrocytes. This response is initiated by phosphorylation of eIF2? to diminish global protein translation and selectively allow for the synthesis of protective proteins. The integrated stress response is terminated by dephosphorylation of eIF2?. The small molecule Sephin1 inhibits eIF2? dephosphorylation, thereby prolonging the protective response. Herein, we tested the effectiveness of Sephin1 in shielding oligodendrocytes against inflammatory stress. We confirmed that Sephin1 prolonged eIF2? phosphorylation in stressed primary oligodendrocyte cultures. Moreover, by using a mouse model of multiple sclerosis, experimental autoimmune encephalomyelitis, we demonstrated that Sephin1 delayed the onset of clinical symptoms, which correlated with a prolonged integrated stress response, reduced oligodendrocyte and axon loss, as well as diminished T cell presence in the CNS. Sephin1 is reportedly a selective inhibitor of GADD34 (PPP1R15A), which is a stress-induced regulatory subunit of protein phosphatase 1 complex that dephosphorylates eIF2?. Consistent with this possibility, GADD34 mutant mice presented with a similar ameliorated experimental autoimmune encephalomyelitis phenotype as Sephin1-treated mice, and Sephin1 did not provide additional therapeutic benefit to the GADD34 mutant animals. Results presented from the adoptive transfer of encephalitogenic T cells between wild-type and GADD34 mutant mice further indicate that the beneficial effects of Sephin1 are mediated through a direct protective effect on the CNS. Of particular therapeutic relevance, Sephin1 provided additive therapeutic benefit when combined with the first line multiple sclerosis drug, interferon ?. Together, our results suggest that a neuroprotective treatment based on the enhancement of the integrated stress response would likely have significant therapeutic value for multiple sclerosis patients.

Project description:BACKGROUND: Neuroprotective and neurotrophic properties of leukemia inhibitory factor (LIF) have been widely reported. In the central nervous system (CNS), astrocytes are the major source for LIF, expression of which is enhanced following disturbances leading to neuronal damage. How astrocytic LIF expression is regulated, however, has remained an unanswered question. Since neuronal stress is associated with production of extracellular adenosine, we investigated whether LIF expression in astrocytes was mediated through adenosine receptor signaling. METHODS: Mouse cortical neuronal and astrocyte cultures from wild-type and adenosine A(2B) receptor knock-out animals, as well as adenosine receptor agonists/antagonists and various enzymatic inhibitors, were used to study LIF expression and release in astrocytes. When needed, a one-way analysis of variance (ANOVA) followed by Bonferroni post-hoc test was used for statistical analysis. RESULTS: We show here that glutamate-stressed cortical neurons induce LIF expression through activation of adenosine A(2B) receptor subtype in cultured astrocytes and require signaling of protein kinase C (PKC), mitogen-activated protein kinases (MAPKs: p38 and ERK1/2), and the nuclear transcription factor (NF)-?B. Moreover, LIF concentration in the supernatant in response to 5'-N-ethylcarboxamide (NECA) stimulation was directly correlated to de novo protein synthesis, suggesting that LIF release did not occur through a regulated release pathway. Immunocytochemistry experiments show that LIF-containing vesicles co-localize with clathrin and Rab11, but not with pHogrin, Chromogranin (Cg)A?and?CgB, suggesting that LIF might be secreted through recycling endosomes. We further show that pre-treatment with supernatants from NECA-treated astrocytes increased survival of cultured cortical neurons against glutamate, which was absent when the supernatants were pre-treated with an anti-LIF neutralizing antibody. CONCLUSIONS: Adenosine from glutamate-stressed neurons induces rapid LIF release in astrocytes. This rapid release of LIF promotes the survival of cortical neurons against excitotoxicity.

Dataset Information

Sephin1 Protects Neurons against Excitotoxicity Independently of the Integrated Stress Response.

Publications

Sephin1 Protects Neurons against Excitotoxicity Independently of the Integrated Stress Response.

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