Dataset Information

The AMPAR antagonist perampanel protects the neurovascular unit against traumatic injury via regulating Sirt3.

ABSTRACT:

Introduction

Perampanel is a highly selective and noncompetitive ?-amino-3 -hydroxy-5-methyl-4-isoxazole propionate receptor (AMPAR) antagonist, which has been used as an orally administered antiepileptic drug in more than 55 countries. Recently, perampanel was shown to exert neuroprotective effects in hemorrhagic and ischemic stroke models via regulating blood-brain barrier (BBB) function.

Aim

Here, the protective effects of perampanel were investigated in an in vitro neurovascular unit (NVU) system established using a triple cell co-culture model (neurons, astrocytes, and brain microvascular endothelial cells) and in an in vivo traumatic brain injury (TBI) model.

Results

Neurons in the NVU system exhibit a more mature morphological phenotype compared with neurons cultured alone, and the co-culture system mimicked an impermeable barrier in vitro. Perampanel protects the NVU system against traumatic and excitotoxic injury, as evidenced by reduced lactate dehydrogenase (LDH) release and apoptotic rate. Treatment with perampanel attenuated lipid peroxidation and expression of inflammatory cytokines. In addition, perampanel increased Sirt3 protein expression, enhanced the activities of mitochondrial enzyme IDH2 and SOD2, and preserved BBB function in vitro. Knockdown of Sirt3 using specific siRNA (Si-Sirt3) partially reserved the effects of perampanel on neuronal injury and BBB function. Treatment with perampanel in vivo attenuated brain edema, preserved neurological function, inhibited apoptosis and microglia activation after TBI. Furthermore, perampanel increased the expression of Sirt3 and preserved BBB function after TBI. The effect of perampanel on BBB function and brain edema was abolished by knockdown of Sirt3 in vivo.

Conclusion

Our results indicate that the noncompetitive AMPAR antagonist perampanel protects the NVU system and reduces brain damage after TBI via activating the Sirt3 cascades.

SUBMITTER: Chen T

PROVIDER: S-EPMC7804923 | biostudies-literature | 2021 Jan

REPOSITORIES: biostudies-literature

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The AMPAR antagonist perampanel protects the neurovascular unit against traumatic injury via regulating Sirt3.

Chen Tao T Liu Wen-Bo WB Qian Xiao X Xie Ke-Liang KL Wang Yu-Hai YH

CNS neuroscience & therapeutics 20210109 1

<h4>Introduction</h4>Perampanel is a highly selective and noncompetitive α-amino-3 -hydroxy-5-methyl-4-isoxazole propionate receptor (AMPAR) antagonist, which has been used as an orally administered antiepileptic drug in more than 55 countries. Recently, perampanel was shown to exert neuroprotective effects in hemorrhagic and ischemic stroke models via regulating blood-brain barrier (BBB) function.<h4>Aim</h4>Here, the protective effects of perampanel were investigated in an in vitro neurovascul ...[more]

PMID: 33421349

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Project description:Calculus bovis is commonly used for the treatment of stroke in traditional Chinese medicine. Hyodeoxycholic acid (HDCA) is a bioactive compound extracted from calculus bovis. When combined with cholic acid, baicalin and jas-minoidin, HDCA prevents hypoxia-reoxygenation-induced brain injury by suppressing endoplasmic reticulum stress-mediated apoptotic signaling. However, the effects of HDCA in ischemic stroke injury have not yet been studied. Neurovascular unit (NVU) dysfunction occurs in ischemic stroke. Therefore, in this study, we investigated the effects of HDCA on the NVU under ischemic conditions in vitro. We co-cultured primary brain microvascular endothelial cells, neurons and astrocytes using a transwell chamber co-culture system. The NVU was pre-treated with 10.16 or 2.54 μg/mL HDCA for 24 hours before exposure to oxygen-glucose deprivation for 1 hour. The cell counting kit-8 assay was used to detect cell activity. Flow cytometry and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling were used to assess apoptosis. Enzyme-linked immunosorbent assay was used to measure the expression levels of inflammatory cytokines, including interleukin-1β, interleukin-6 and tumor necrosis factor-α, and neurotrophic factors, including brain-derived neurotrophic factor and glial cell line-derived neurotrophic factor. Oxidative stress-related factors, such as superoxide dismutase, nitric oxide, malondialdehyde and γ-glutamyltransferase, were measured using kits. Pretreatment with HDCA significantly decreased blood-brain barrier permeability and neuronal apoptosis, significantly increased transendothelial electrical resistance and γ-glutamyltransferase activity, attenuated oxidative stress damage and the release of inflammatory cytokines, and increased brain-derived neurotrophic factor and glial cell line-derived neurotrophic factor expression. Our findings suggest that HDCA maintains NVU morphological integrity and function by modulating inflammation, oxidation stress, apoptosis, and the expression of neurotrophic factors. Therefore, HDCA may have therapeutic potential in the clinical management of ischemic stroke. This study was approved by the Ethics Committee of Experimental Animals of Beijing University of Chinese Medicine (approval No. BUCM-3-2016040201-2003) in April 2016.

Project description:In addition to neurons, all components of the neurovascular unit (NVU), such as glial, endothelial, and basal membranes, are destroyed during traumatic brain injury (TBI). Previous studies have shown that excessive stimulation of calpain is crucial for cerebral injury after traumatic insult. The objective of this study was to investigate whether calpain activation participated in NVU disruption and edema formation in a mouse model of controlled cortical impact (CCI).One hundred and eight mice were divided into three groups: the sham group, the control group, and the MDL28170 group. MDL28170 (20 mg/kg), an efficient calpain inhibitor, was administered intraperitoneally at 5 min, 3 h, and 6 h after experimental CCI. We then measured neurobehavioral deficits, calpain activity, inflammatory mediator levels, blood-brain barrier (BBB) disruption, and NVU deficits using electron microscopy and histopathological analysis at 6 h and 24 h after CCI.The MDL28170 treatment significantly reduced the extent of both cerebral contusion (MDL28170 vs. vehicle group, 16.90 ± 1.01 mm? and 17.20 ± 1.17 mm? vs. 9.30 ± 1.05 mm? and 9.90 ± 1.17 mm?, both P < 0.001) and edema (MDL28170 vs. vehicle group, 80.76 ± 1.25% and 82.00 ± 1.84% vs. 82.55 ± 1.32% and 83.64 ± 1.25%, both P < 0.05), improved neurological scores (MDL28170 vs. vehicle group, 7.50 ± 0.45 and 6.33 ± 0.38 vs. 12.33 ± 0.48 and 11.67 ± 0.48, both P < 0.001), and attenuated NVU damage resulting (including tight junction (TJ), basement membrane, BBB, and neuron) from CCI at 6 h and 24 h. Moreover, MDL28170 markedly downregulated nuclear factor-?B-related inflammation (tumor necrosis factor-? [TNF-?]: MDL28170 vs. vehicle group, 1.15 ± 0.07 and 1.62 ± 0.08 vs. 1.59 ± 0.10 and 2.18 ± 0.10, both P < 0.001; inducible nitric oxide synthase: MDL28170 vs. vehicle group, 4.51 ± 0.23 vs. 6.23 ± 0.12, P < 0.001 at 24 h; intracellular adhesion molecule-1: MDL28170 vs. vehicle group, 1.45 ± 0.13 vs. 1.70 ± 0.12, P < 0.01 at 24 h) and lessened both myeloperoxidase activity (MDL28170 vs. vehicle group, 0.016 ± 0.001 and 0.016 ± 0.001 vs. 0.024 ± 0.001 and 0.023 ± 0.001, P < 0.001 and 0.01, respectively) and matrix metalloproteinase-9 (MMP-9) levels (MDL28170 vs. vehicle group, 0.87 ± 0.13 and 1.10 ± 0.10 vs. 1.17 ± 0.13 and 1.25 ± 0.12, P < 0.001 and 0.05, respectively) at 6 h and 24 h after CCI.These findings demonstrate that MDL28170 can protect the structure of the NVU by inhibiting the inflammatory cascade, reducing the expression of MMP-9, and supporting the integrity of TJ during acute TBI.

Project description:BackgroundThe brain vasculature plays a pivotal role in the inflammatory process by modulating the interaction between blood cells and the neurovascular unit. Argonaute-2 (Ago2) has been suggested as essential for endothelial survival but its role in the brain vasculature or in the endothelial-glial crosstalk has not been addressed. Thus, our aim was to clarify the significance of Ago2 in the inflammatory responses elicited by these cell types.MethodsMouse primary cultures of brain endothelial cells, astrocytes and microglia were used to evaluate cellular responses to the modulation of Ago2. Exposure of microglia to endothelial cell-conditioned media was used to assess the potential for in vivo studies. Adult mice were injected intraperitoneally with lipopolysaccharide (LPS) (2 mg/kg) followed by three daily intraperitoneal injections of Ago2 (0.4 nM) to assess markers of endothelial disruption, glial reactivity and neuronal function.ResultsHerein, we demonstrated that LPS activation disturbed the integrity of adherens junctions and downregulated Ago2 in primary brain endothelial cells. Exogenous treatment recovered intracellular Ago2 above control levels and recuperated vascular endothelial-cadherin expression, while downregulating LPS-induced nitric oxide release. Primary astrocytes did not show a significant change in Ago2 levels or response to the modulation of the Ago2 system, although endogenous Ago2 was shown to be critical in the maintenance of tumor necrosis factor-α basal levels. LPS-activated primary microglia overexpressed Ago2, and Ago2 silencing contained the inflammatory response to some extent, preventing interleukin-6 and nitric oxide release. Moreover, the secretome of Ago2-modulated brain endothelial cells had a protective effect over microglia. The intraperitoneal injection of LPS impaired blood-brain barrier and neuronal function, while triggering inflammation, and the subsequent systemic administration of Ago2 reduced or normalized endothelial, glial and neuronal markers of LPS damage. This outcome likely resulted from the direct action of Ago2 over the brain endothelium, which reestablished glial and neuronal function.ConclusionsAgo2 could be regarded as a putative therapeutic agent, or target, in the recuperation of the neurovascular unit in inflammatory conditions.

Dataset Information

The AMPAR antagonist perampanel protects the neurovascular unit against traumatic injury via regulating Sirt3.

Introduction

Aim

Results

Conclusion

Publications

The AMPAR antagonist perampanel protects the neurovascular unit against traumatic injury via regulating Sirt3.

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