Reduction of glutamate-induced excitotoxicity in murine primary neurons involving calpain inhibition

Gold, Maike; Koczulla, A. Rembert; Mengel, David; Koepke, Janine; Dodel, Richard; Dontcheva, Guergana; Habib, Pardes; Bach, Jan-Philipp

doi:10.1016/j.jns.2015.11.016

Cited by 14 publications

(21 citation statements)

References 29 publications

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“…We therefore hypothesize that A1AT effects on NLRP3 upregulation in primary astrocytes are mainly triggered by an inhibition of calcium and calpain. This hypothesis needs further evaluation, since other reports also demonstrate that A1AT is able to reduce glutamate-induced toxicity in murine primary neurons [ 45 ]. Since astrocytes release glutamate in response to Aβ 1–42 -stimulation, this could represent another way how A1AT prevents deleterious Aβ 1–42 -induced inflammatory cascades in microglia, neurons, and astrocytes.…”

Section: Discussionmentioning

confidence: 99%

“…Conveniently, A1AT is therapeutically used in patients with A1AT-deficiency and therefore well-established as a pharmaceutical agent. Recently, we demonstrated that A1AT also protected neurons from glutamate-induced toxicity [ 45 ] and reduced Aβ 1–42 -induced inflammation in microglial cells [ 15 ]. In addition, we found that A1AT inhibited calpain and stabilized calcium-homeostasis [ 15 ].…”

Section: Introductionmentioning

confidence: 99%

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α1-antitrypsin mitigates NLRP3-inflammasome activation in amyloid β1–42-stimulated murine astrocytes

Ebrahimi

Rust

Kaiser

et al. 2018

J Neuroinflammation

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BackgroundNeuroinflammation has an essential impact on the pathogenesis and progression of Alzheimer’s disease (AD). Mostly mediated by microglia and astrocytes, inflammatory processes lead to degeneration of neuronal cells. The NLRP3-inflammasome (NOD-like receptor family, pyrin domain containing 3) is a key component of the innate immune system and its activation results in secretion of the proinflammatory effectors interleukin-1β (IL-1β) and interleukin-18 (IL-18). Under physiological conditions, cytosolic NLRP3-inflammsome is maintained in an inactive form, not able to oligomerize. Amyloid β1–42 (Aβ1–42) triggers activation of NLRP3-inflammasome in microglia and astrocytes, inducing oligomerization and thus recruitment of proinflammatory proteases. NLRP3-inflammasome was found highly expressed in human brains diagnosed with AD. Moreover, NLRP3-deficient mice carrying mutations associated with familial AD were partially protected from deficits associated with AD.The endogenous protease inhibitor α1-antitrypsin (A1AT) is known for its anti-inflammatory and anti-apoptotic properties and thus could serve as therapeutic agent for NLRP3-inhibition. A1AT protects neurons from glutamate-induced toxicity and reduces Aβ1–42-induced inflammation in microglial cells. In this study, we investigated the effect of Aβ1–42-induced NLRP3-inflammasome upregulation in primary murine astrocytes and its regulation by A1AT.MethodsPrimary cortical astrocytes from BALB/c mice were stimulated with Aβ1–42 and treated with A1AT. Regulation of NLRP3-inflammasome was examined by immunocytochemistry, PCR, western blot and ELISA. Our studies included an inhibitor of NLRP3 to elucidate direct interactions between A1AT and NLRP3-inflammasome components.ResultsOur study revealed that A1AT reduces Aβ1–42-dependent upregulation of NLRP3 at the mRNA and protein levels. Furthermore, A1AT time-dependently mitigated the expression of caspase 1 and its cleavage product IL-1β in Aβ1–42-stimulated astrocytes.ConclusionWe conclude that Aβ1–42-stimulation results in an upregulation of NLRP3, caspase 1, and its cleavage products in astrocytes. A1AT time-dependently hampers neuroinflammation by downregulation of Aβ1–42-mediated NLRP3-inflammasome expression and thus may serve as a pharmaceutical opportunity for the treatment of Alzheimer’s disease.Electronic supplementary materialThe online version of this article (10.1186/s12974-018-1319-x) contains supplementary material, which is available to authorized users.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

α1-antitrypsin mitigates NLRP3-inflammasome activation in amyloid β1–42-stimulated murine astrocytes

Ebrahimi

Rust

Kaiser

et al. 2018

J Neuroinflammation

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“…Interestingly, proteases outside the serine-protease family are also inhibited by hAAT, albeit to a lesser extent. These include metalloproteases [e.g., MMP-9 ( 15 – 17 ), ADAMTS-4 ( 18 ), and cysteine-proteases (e.g., caspase-3) ( 19 , 20 )], suggesting that some functions of the molecule may extend beyond the specificity conferred by the primary sequence of the RCL. As such, it has been proposed that the globular surface of hAAT may contain significant functional attributes.…”

Section: Introductionmentioning

confidence: 99%

Point Mutation of a Non-Elastase-Binding Site in Human α1-Antitrypsin Alters Its Anti-Inflammatory Properties

et al. 2018

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IntroductionHuman α1-antitrypsin (hAAT) is a 394-amino acid long anti-inflammatory, neutrophil elastase inhibitor, which binds elastase via a sequence-specific molecular protrusion (reactive center loop, RCL; positions 357–366). hAAT formulations that lack protease inhibition were shown to maintain their anti-inflammatory activities, suggesting that some attributes of the molecule may reside in extra-RCL segments. Here, we compare the protease-inhibitory and anti-inflammatory profiles of an extra-RCL mutation (cys232pro) and two intra-RCL mutations (pro357cys, pro357ala), to naïve [wild-type (WT)] recombinant hAAT, in vitro, and in vivo.MethodsHis-tag recombinant point-mutated hAAT constructs were expressed in HEK-293F cells. Purified proteins were evaluated for elastase inhibition, and their anti-inflammatory activities were assessed using several cell-types: RAW264.7 cells, mouse bone marrow-derived macrophages, and primary peritoneal macrophages. The pharmacokinetics of the recombinant variants and their effect on LPS-induced peritonitis were determined in vivo.ResultsCompared to WT and to RCL-mutated hAAT variants, cys232pro exhibited superior anti-inflammatory activities, as well as a longer circulating half-life, despite all three mutated forms of hAAT lacking anti-elastase activity. TNFα expression and its proteolytic membranal shedding were differently affected by the variants; specifically, cys232pro and pro357cys altered supernatant and serum TNFα dynamics without suppressing transcription or shedding.ConclusionOur data suggest that the anti-inflammatory profile of hAAT extends beyond direct RCL regions. Such regions might be relevant for the elaboration of hAAT formulations, as well as hAAT-based drugs, with enhanced anti-inflammatory attributes.

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“…Moreover, we demonstrated that the regulation of the calcium influx exerted by HPTOPE results in the significant prevention of overactivation of the calcium-dependent protease calpain. Since the uncontrolled activation of calpain is lethal to neurons, as well as to other cell types [22,43], we can hypothesize that the preservation of neuronal viability by cell treatment with HPTOPE could be related to a preservation of physiological calpain activity. In this context, it is possible to find a relationship between calpain activity and neuronal cell death by evaluating the occurrence of a specific degradation pattern of nNOS [39,40,44].…”

Section: Discussionmentioning

confidence: 99%

“…Moreover, when the calcium homeostasis is compromised, the persistent activation of the proteolytic calcium-dependent protease calpain, known to be involved in the development of neurodegeneration, leads to the uncontrolled degradation of its substrates and, finally, to cell death [20]. For this reason, the inhibition of calpain is considered an interesting target to prevent neurodegenerative disease associated with neuronal loss [21][22][23][24].…”

Section: Introductionmentioning

confidence: 99%

A Bioactive Olive Pomace Extract Prevents the Death of Murine Cortical Neurons Triggered by NMDAR Over-Activation

et al. 2020

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We have recently demonstrated that bioactive molecules, extracted by high pressure and temperature from olive pomace, counteract calcium-induced cell damage to different cell lines. Here, our aim was to study the effect of the same extract on murine cortical neurons, since the preservation of the intracellular Ca2+-homeostasis is essential for neuronal function and survival. Accordingly, we treated neurons with different stimuli in order to evoke cytotoxic glutamatergic activation. In these conditions, the high-pressure and temperature extract from olive pomace (HPTOPE) only abolished the effects of N-methyl-d-aspartate (NMDA). Particularly, we observed that HPTOPE was able to promote the neuron rescue from NMDA-induced cell death. Moreover, we demonstrated that HPTOPE is endowed with the ability to maintain the intracellular Ca2+-homeostasis following NMDA receptor overactivation, protecting neurons from Ca2+-induced adverse effects, including aberrant calpain proteolytic activity. Moreover, we highlight the importance of the extraction conditions used that, without producing toxic molecules, allow us to obtain protecting molecules belonging to proanthocyanidin derivatives like procyanidin B2. In conclusion, we can hypothesize that HPTOPE, due to its functional and nontoxic properties on neuronal primary culture, can be utilized for future therapeutic interventions for neurodegeneration.

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Reduction of glutamate-induced excitotoxicity in murine primary neurons involving calpain inhibition

Cited by 14 publications

References 29 publications

α1-antitrypsin mitigates NLRP3-inflammasome activation in amyloid β1–42-stimulated murine astrocytes

α1-antitrypsin mitigates NLRP3-inflammasome activation in amyloid β1–42-stimulated murine astrocytes

Point Mutation of a Non-Elastase-Binding Site in Human α1-Antitrypsin Alters Its Anti-Inflammatory Properties

A Bioactive Olive Pomace Extract Prevents the Death of Murine Cortical Neurons Triggered by NMDAR Over-Activation

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