Mouse Brain Research Articles

Structure-based discovery of a 4,5-Dihydropyrazole-cored PET ligand for imaging of receptor-interacting serine/threonine-protein kinase 1 (RIPK1) in the brain

Receptor-interacting serine/threonine-protein kinase 1 (RIPK1) regulates programmed cell death and inflammation, contributing to a wide range of human pathologies, including inflammatory disorders, neurodegenerative conditions, and cancer. Despite this, no RIPK1 positron emission tomography (PET) ligand with significant in vivo specificity has been reported to date. In this work, we designed and synthesized a new family of dihydropyrazole-cored ligands suitable for 18F-labeling at the late stage. Among these, WL8 showed a strong binding affinity to RIPK1 (EC50 = 19.9 nM, Kd = 25 nM) and was successfully labeled with 18F in the 6-position of pyridine ring, yielding a high radiochemistry yield of 27.9 % (decay-corrected) and a high molar activity of 18.8–31.2 GBq/μmol. In in vitro autoradiography, [18F]WL8 showed some specific binding in the brain sections of rats and lipopolysaccharide (LPS) model mice. Preliminary PET studies in rat brains revealed that [18F]WL8 could efficiently penetrate the blood-brain barrier and was rapidly washed out. As anticipated, [18F]WL8 exhibited a high initial uptake (brain2min = 4.80 % ID/g) in mouse brains, followed by a rapid washout (brain60min = 0.14 % ID/g), although no clear specific binding to RIPK1 was observed. Moderate in vivo stability was noted for [18F]WL8 in mouse brains with 35.2 % of the parent fraction remaining after 30 min post-administration. Altogether, our work broadens the landscape and offers a new chemotype for RIPK1 PET ligand development.

Serialized on-grid lift-in sectioning for tomography (SOLIST) enables a biopsy at the nanoscale

Cryo-focused ion beam milling has substantially advanced our understanding of molecular processes by opening windows into cells. However, applying this technique to complex samples, such as tissues, has presented considerable technical challenges. Here we introduce an innovative adaptation of the cryo-lift-out technique, serialized on-grid lift-in sectioning for tomography (SOLIST), addressing these limitations. SOLIST enhances throughput, minimizes ice contamination and improves sample stability for cryo-electron tomography. It thereby facilitates the high-resolution imaging of a wide range of specimens. We illustrate these advantages on reconstituted liquid–liquid phase-separated droplets, brain organoids and native tissues from the mouse brain, liver and heart. With SOLIST, cellular processes can now be investigated at molecular resolution directly in native tissue. Furthermore, our method has a throughput high enough to render cryo-lift-out a competitive tool for structural biology. This opens new avenues for unprecedented insights into cellular function and structure in health and disease, a ‘biopsy at the nanoscale’.

Ultra-high-resolution mapping of myelin and g-ratio in a panel of Mbp enhancer-edited mouse strains using microstructural MRI

Non-invasive myelin water fraction (MWF) and g-ratio mapping using microstructural MRI have the potential to offer critical insights into brain microstructure and our overall understanding of neuroplasticity and neuroinflammation. By leveraging a unique panel of variably hypomyelinating mouse strains, we validated a high-resolution, model-free image reconstruction method for whole-brain MWF mapping. Further, by employing a bipolar gradient echo MRI sequence, we achieved high spatial resolution and robust mapping of MWF and g-ratio across the whole mouse brain. Our regional white matter-tract specific analyses demonstrated a graded decrease in MWF in white matter tracts which correlated strongly with myelin basic protein gene (Mbp) mRNA levels. Using these measures, we derived the first sensitive calibrations between MWF and Mbp mRNA in the mouse. Minimal changes in axonal density supported our hypothesis that observed MWF alterations stem from hypomyelination. Overall, our work strongly emphasizes the potential of non-invasive, MRI-derived MWF and g-ratio modeling for both preclinical model validation and ultimately translation to humans.

Factors controlling the organ-specific T1 contrast effect of silica nanoparticles co-doped with both Mn2+ ions and oleate-coated iron oxides.

The present work introduces the synergistic effect of co-doping of both oleate-coated superparamagnetic iron oxide nanoparticles (SPIONs) and Mn(NO3)2 into silica nanoparticles (SNs) on the T1-relaxivity of Mn2+ ions. The observed synergism can be attributed to the limited oxidation of Mn2+ ions when they are doped into the outer layer of SNs doped with SPIONs, despite the alkaline synthesis conditions. The electrochemical behaviour of the manganese ions inside co-doped SNs corroborates the predominance of their oxidation state +2. Moreover, the T1 relaxivities of co-doped SNs have been determined to be 20.0mM-1s-1 and 30.0mM-1s-1 at 0.47T. The T2 relaxivity of co-doped SNs can be tuned by incorporating 6 or 13nm SPIONs with different saturation magnetizations, which allows the T2/T1 relaxation ratios to be limited to 0.8–5.9. The incorporation of amino groups on the surface of co-doped SNs by substituting silanol groups with propylamino groups reduces T1 relaxivity to 9.0mM-1s-1, which is nevertheless sufficient to provide a brightening of the abdominal organs and mouse brain in magnetic resonance imaging at 11.7T. The preferential localisation of co-doped SNs in the kidneys and intestines compared to the liver is a consequence of the specificity of amino-substituted SNs compared to bare SNs.

Targeting ceramide-induced microglial pyroptosis: Icariin is a promising therapeutic therapy for Alzheimer's disease

Alzheimer’s disease (AD), a progressive dementia, is one of the most common neurodegenerative diseases. Clinical trial results of amyloid-β (Aβ) and tau regulators based on the pretext of straightforward amyloid and tau immunotherapy were disappointing. There are currently no effective strategies for slowing the progression of AD. Herein, we spotlight the dysregulation of lipid metabolism, particularly the elevation of ceramides (Cers), as a critical yet underexplored facet of AD pathogenesis. Our study delineates the role of Cers in promoting microglial pyroptosis, a form of programmed cell death distinct from apoptosis and necroptosis, characterized by cellular swelling, and membrane rupture mediated by the NLRP3 inflammasome pathway. Utilizing both in vivo experiments with APP/PS1 transgenic mice and in vitro assays with BV-2 microglial cells, we investigate the activation of microglial pyroptosis by Cers and its inhibition by Icariin (ICA), a flavonoid with known antioxidant and anti-inflammatory properties. Our findings reveal a significant increase in Cers levels and pyroptosis markers (NLRP3, apoptosis-associated speck-like protein containing a caspase recruitment domain, caspase-1, gasdermin D, and interleukin-18) in the brains of AD model mice, indicating a direct involvement of Cers in AD pathology through the induction of microglial pyroptosis. Conversely, ICA treatment effectively reduces these pyroptotic markers and Cer levels, thereby attenuating microglial pyroptosis and suggesting a novel therapeutic mechanism of action against AD. This study not only advances our understanding of the pathogenic role of Cers in AD but also introduces ICA as a promising candidate for AD therapy, capable of mitigating neuroinflammation and pyroptosis through the COX2-NLRP3 inflammasome-GSDMD axis. Our results pave the way for further exploration of Cer metabolism disorders in neurodegenerative diseases and highlight the therapeutic potential of targeting microglial pyroptosis in AD.

Distinct functional diversity of branched oligosaccharides as chaperones and inhibitory-binding partners of amyloid beta-protein and its aggregates

Aggregation and deposition of amyloid beta-protein 1–42 (Aβ42) in the brain, primarily owing to hydrophobic interactions between Aβ42 chains, is a common pathology in all forms of Alzheimer's disease (AD). Hydrophilic oligosaccharides are widely present in the extracellular matrix and on the cytoplasmic membrane. To determine if oligosaccharides bind to Aβ42 or its aggregates and consequently affect their aggregation and cellular function, this study examined the interaction of typical functional oligosaccharides with Aβ42 or its aggregates. Isomaltooligosaccharides (IMOs), particularly isomaltotriose, panose, and isomaltotetraose, functioned as molecular chaperones for Aβ42 by binding directly to Aβ42, preserving Aβ42's active conformation and cytotrophic activity. Oral IMOs reduced total plasma Aβ level and indirectly caused a slight reduction in the load of Aβ42 spots/plaques in the brain of AD model mice (male). Another branched oligosaccharide, bianntennary core pentasaccharide (BCP), had a relatively high binding specificity for Aβ42 oligomers (Aβ42O) and acted as an antagonistic binding partner for Aβ42O. Free BCP effectively blocked/prevented further assembly of Aβ42O and their toxicity to neural and vascular endothelial cell lines. Since BCP is also a signaling component of membrane targets (glycolipids, glycoproteins or receptors), it seemed that BCP had two opposing effects on the binding of Aβ42O to target cells. This study's findings suggest that these branched oligosaccharides may be potential candidates for blocking or preventing Aβ42 aggregation and Aβ42O cytotoxicity/neurotoxicity, respectively, and that IMO-like or free BCP-like oligosaccharide deficiencies in the brain may be one of the underlying mechanisms for Aβ42 aggregation and Aβ42O cytotoxicity.

Anisomycin alleviates cognitive impairments and pathological features in 3xTg-AD mice

Neuroinflammation plays a pivotal role in the pathogenesis of Alzheimer’s disease (AD). Anisomycin is a pyrrolidine antibiotic isolated from Streptomyces griseolus, which is an efficient anti-inflammatory agent that functions both in vivo and in vitro. However, it is not clear whether anisomycin can exert neuroprotective effect in AD. In the present study, anisomycin was intragastrically administrated to female triple-transgenic AD (3xTg-AD) model mice, then Morris water maze test was used to observe the long-term spatial memory of mice, the in vivo hippocampal field potential recording was performed to evaluate the synaptic plasticity, the western blot and immunofluorescence were employed to detect pathological changes, and the bioinformatics analysis was used to predict the potential target of anisomycin exerting effects in AD. The results showed that anisomycin ameliorated the long-term spatial memory deficits, improved LTP depression and increased the expression of PSD-95, reduced the Aβ and tau pathologies, and alleviated the activation of microglia and astrocytes in the brains of 3xTg-AD mice. In addition, the results from bioinformatics analysis showed that the potential target of anisomycin focused on inflammatory pathway. These results indicated that anisomycin exerts neuroprotective effects in 3xTg-AD mice by alleviating neuroinflammation, but the potential mechanism of anisomycin exerting neuroprotective effects needs to be further investigated.

SERS detection of dopamine in artificial cerebrospinal fluid and in Parkinson’s disease-induced mouse cortex using a hybrid ZnO@Ag nanostructured platform

Rapid, direct, and sensitive detection of metabolites or clinically relevant biomarkers in complex biological samples is still challenging. We report a label-free surface-enhanced Raman scattering (SERS)-based approach for highly sensitive detection of dopamine (DA) in artificial cerebrospinal fluid (aCSF) and mouse brain tissue samples. The hybrid SERS-active sensing platform was designed to maximize the hot spot distribution. It encompasses a network of flexible and polymeric nanotrenches and nanogaps fabricated by nanoimprint lithography (NIL) covered by thin films of zinc oxide (ZnO) and silver (Ag) deposited using pulsed laser deposition (PLD) and direct current magnetron sputtering (DC-MS), respectively. The growth of the ZnO and Ag thin films was assessed by scanning electron microscopy (SEM) technique. The proposed SERS substrate benefits from the interlayered semiconductor–metal contribution in SERS enhancement, leading to a versatile sensing platform with improved detection sensitivity through the increased hot spots distribution. Using this strategy, the proposed assay can achieve an ultralow DA determination at the nM level, a short testing time (<30 min), and high signal reproducibility (RSD 11 %). We also assessed the sensing attributes of our in house developed SERS platform to detect DA in spiked aCSF samples, and a limit of detection (LOD) of 10 μM was achieved. Additionally, after in vivo inducing Parkinson’s disease (PD) in mice, relevant biological samples (striatum and cortical brain tissue) were investigated by SERS and ELISA. Given its good stability and accuracy in complex samples, the SERS-ELISA analysis has great potential to be a powerful tool for the reliable detection of DA in remote conditions.

Brain organoids: A new tool for modelling of neurodevelopmental disorders.

Neurodevelopmental disorders are mostly studied using mice as models. However, the mouse brain lacks similar cell types and structures as those of the human brain. In recent years, emergence of three-dimensional brain organoids derived from human embryonic stem cells or induced pluripotent stem cells allows for controlled monitoring and evaluation of early neurodevelopmental processes and has opened a window for studying various aspects of human brain development. However, such organoids lack original anatomical structure of the brain during maturation, and neurodevelopmental maturation processes that rely on unique cellular interactions and neural network connections are limited. Consequently, organoids are difficult to be used extensively and effectively while modelling later stages of human brain development and disease progression. To address this problem, several methods and technologies have emerged that aim to enhance the sophisticated regulation of brain organoids developmental processes through bioengineering approaches, which may alleviate some of the current limitations. This review discusses recent advances and application areas of human brain organoid culture methods, aiming to generalize optimization strategies for organoid systems, improve the ability to mimic human brain development, and enhance the application value of organoids.

Dynamic changes in Beclin-1, LC3B, and p62 in aldose reductase-knockout mice at different time points after ischemic stroke

Ischemic stroke is a brain injury caused by cerebral blood circulation disorders and is closely related to oxidative stress. Aldose reductase (AR) is a critical enzyme involved in oxidative stress. Autophagy has previously been found to play a key role in cerebral ischemia‒reperfusion injury. However, it is still unclear how autophagy molecules change after cerebral ischemia‒reperfusion injury in AR knockout mice (AR-/-). A transient middle cerebral artery occlusion (tMCAO) model was generated in AR-/- mice, and the neurological deficit scores of the mice were observed and recorded on Days 1, 3 and 5 after tMCAO. Neuronal damage in the ischemic penumbra was observed by TTC, HE, and Nissl staining. The expression of the autophagy-related molecules Beclin-1, LC3II/I, and P62 as well as that of molecules related to inflammation, oxidative stress, and neurological damage was detected by RT‒qPCR, western blotting, and immunofluorescence. Autophagosomes were observed using a transmission electron microscope. Cerebral ischemia‒reperfusion injury caused neurological deficits and ischemic infarction in tMCAO mice (P<0.01). Beclin-1, Bcl2/Bax, SOD, GSH-px, P62, PSD95, and TOM20 levels decreased (P<0.05), while IL-6, LC3II/I, and GFAP levels increased (P<0.01) in the AR-/- tMCAO-1d group and the AR-/- tMCAO-3d group, compared to those in the sham group. Beclin-1, Bcl2/Bax, NOX4, GSH-px, P62, and PSD95 levels increased (P<0.01), while IL-6, LC3II/I, and GFAP levels decreased (P<0.01) in the AR-/- tMCAO-5d group compared to those in the AR-/- tMCAO-1d group. Autophagosome formation was observed in tMCAO mice. In summary, the changes in autophagy proteins in the brain tissue of the AR-/- mice after tMCAO were more obvious on Days 1 and 3 after tMCAO. The expression of Beclin-1 and P62 decreased, and the expression of LC3B increased after cerebral ischemia‒reperfusion injury in AR-/- mouse brain tissue.

ADAR3 modulates neuronal differentiation and regulates mRNA stability and translation.

ADAR3 is a catalytically inactive member of the family of adenosine deaminases acting on RNA (ADARs). Here we have investigated its function in the context of the developing mouse brain. The expression of ADAR3 gradually increases throughout embryogenesis and drops after birth. Using primary cortical neurons, we show that ADAR3 is only expressed in a subpopulation of in vitro differentiated neurons, which suggests specific functions rather than being a general regulator of ADAR editing in the brain. The analysis of the ADAR3 interactome suggested a role in mRNA stability and translation, and we show that expression of ADAR3 in a neuronal cell line that is otherwise ADAR3-negative changes the expression and stability of a large number of mRNAs. Notably, we show that ADAR3 associates with polysomes and inhibits translation. We propose that ADAR3 binds to target mRNAs and stabilizes them in non-productive polysome complexes. Interestingly, the expression of ADAR3 downregulates genes related to neuronal differentiation and inhibits neurofilament outgrowth in vitro. In summary, we propose that ADAR3 negatively regulates neuronal differentiation, and that it does so by regulating mRNA stability and translation in an editing-independent manner.

Investigating the influence of estrous cycle-dependent hormonal changes on neurogenesis in adult mice

ObjectiveNeurogenesis is the process of generating new neurons from neural stem cells (NSCs) in the adult brain. Sex hormones play an essential role in the development of the brain. The aim of this study was to evaluate the neurogenic changes in the brain at different phases of the estrous cycle in adult mice. Materials and methodsFemale NMRI mice were divided into four groups: 1- Estrous, 2- Proestrous, 3- Metestrous, and 4- Diestrous. Different stages of the estrous cycle were determined by staining of vaginal smears. The level of estrogen, progesterone, prolactin, follicle-stimulating hormone (FSH) and luteinizing hormone (LH) hormones was evaluated by the enzyme-linked immunosorbent assay (ELISA) method. The expression of brain-derived neurotrophic factor) BDNF), nerve growth factor (NGF), ciliary neurotrophic factor(CNTF)) genes in hippocampal and the expression of glial fibrillary acidic protein (GFAP) in subventricular zone (SVZ) tissue were evaluated. ResultsThe serum estrogen and FSH increased significantly in Proestrous group (p < 0.001). Also, progesterone and prolactin hormones were significantly increased in the Diaestrus group (p < 0.001). The expression levels of BDNF, NGF, and CNTF significantly increased in the hippocampal tissue of Proestrous and Diaestrus groups (p < 0.001). The number of GFAP+ cells in SVZ of the Proestrous and Diestrous groups had a significant increase (p < 0.05, p < 0.01, p < 0.001). ConclusionOur data showed that Changes in sex hormones, especially estrogen in the estrous cycle, can cause the production of new neurons and astrocytes in the hippocampus and SVZ. Therefore, the increase in neurotrophic factors in the Proestrus and Diestrus lead to neurogenesis in adult mice brains.

Identification and analysis correlation between hub genes and immune cell infiltration related to LPS-induced cognitive impairment

BackgroundThe occurrence of immunity and inflammation outside the central nervous system frequently results in acute cognitive impairment among elderly patients. However, there is currently a lack of standardized methods for diagnosing acute cognitive impairment. The objective of our study was to identify potential mRNA biomarkers and investigate the pathogenesis of acute cognitive impairment in mice brains. MethodsTo analyze changes in hub genes associated with acute cognitive impairment, bioinformatics analysis was performed on the mouse brain injury data of sterile saline control group and lipopolysaccharide (LPS) induced experimental group in Gene Expression Omnibus (GEO). Functional analysis was conducted using Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), which facilitated to identify some potential mRNA biomarkers for hub gene expression in mice brains. Additionally, the "CIBERSORT X″ R kit was employed to examine immune cell infiltrations of mice brains in LPS group and saline group. ResultsIn the LPS and saline group, 102 significantly upregulated differentially expressed genes (DEGs) and 32 downregulated DEGs were identified. The pathway enrichment analysis using GO and KEGG revealed that these DEGs were mainly related to the regulation of cytokine, cytokine-cytokine receptor interaction, as well as protein interaction with cytokine and cytokine receptor. Immune cell infiltration analysis indicated potential involvement of M1 macrophages, NK cells resting, T cells CD4 memory, and T cells CD8 naive in the process of cognitive impairment. By constructing a protein-protein interaction (PPI) network, five hub genes (Cxcl10, Cxcl12, Cxcr3, Gbp2, and Ifih1) showed significant associations with immune cell types by using a threshold Spearman's rank correlation coefficient of R > 0.50 and p < 0.05. ConclusionThe mRNA expression profile of the mice brain tissues in the LPS group differed from that in the normal saline group. These significantly expressed mRNAs may act an importance in the pathogenesis of acute cognitive impairment through mechanisms involving immunity and neuroinflammation.

An activatable near-infrared fluorescent probe with large Stokes shift for visualizing peroxynitrite in Alzheimer’s disease models

Alzheimer’s disease (AD), characterized by its incurable nature and prevalence among the elderly, has remained a focal point in medical research. Increasing evidence suggests that peroxynitrite (ONOO−) serves as a crucial biomarker for the diagnosis of AD. In this study, we present a novel, easily available, high-yield, and cost-effective near-infrared (NIR) fluorescent probe, CDCI-ONOO. This probe utilizes a coumarin-dicyanoisophorone conjugate as the fluorophore and diphenylphosphinic chloride as the recognition site, enabling the detection of ONOO− both in vitro and in vivo. Upon interaction with ONOO−, CDCI-ONOO exhibits a distinct maximum emission peak at 715 nm with a substantial Stokes shift of 184 nm. The probe demonstrates excellent selectivity and sensitivity (LOD = 144 nM), along with noticeable colorimetric and fluorescence changes after the reaction. Comprehensive analyses using high-performance liquid chromatography (HPLC), high-resolution mass spectrometry (HRMS), and density functional theory (DFT) calculations confirm that the reaction with ONOO− restores the initially quenched Intramolecular Charge Transfer (ICT), resulting in the formation of CDCI-OH, a product that emitting fluorescence in the near-infrared region. Furthermore, we demonstrated the successful application of CDCI-ONOO for ONOO− detection in neuronal cells and imaging of ONOO− in the brains of mice. These findings underscore the potential of CDCI-ONOO as a near-infrared fluorescent probe for in vivo ONOO− detection, offering a significant avenue for advancing our understanding of AD pathology and diagnosis.

ScCAD: Cluster decomposition-based anomaly detection for rare cell identification in single-cell expression data

Single-cell RNA sequencing (scRNA-seq) technologies have become essential tools for characterizing cellular landscapes within complex tissues. Large-scale single-cell transcriptomics holds great potential for identifying rare cell types critical to the pathogenesis of diseases and biological processes. Existing methods for identifying rare cell types often rely on one-time clustering using partial or global gene expression. However, these rare cell types may be overlooked during the clustering phase, posing challenges for their accurate identification. In this paper, we propose a Cluster decomposition-based Anomaly Detection method (scCAD), which iteratively decomposes clusters based on the most differential signals in each cluster to effectively separate rare cell types and achieve accurate identification. We benchmark scCAD on 25 real-world scRNA-seq datasets, demonstrating its superior performance compared to 10 state-of-the-art methods. In-depth case studies across diverse datasets, including mouse airway, brain, intestine, human pancreas, immunology data, and clear cell renal cell carcinoma, showcase scCAD’s efficiency in identifying rare cell types in complex biological scenarios. Furthermore, scCAD can correct the annotation of rare cell types and identify immune cell subtypes associated with disease, thereby offering valuable insights into disease progression.

Increased Expression of the Neuropeptides PACAP/VIP in the Brain of Mice with CNS Targeted Production of IL-6 Is Mediated in Part by Trans-Signalling.

Inflammation with expression of interleukin 6 (IL-6) in the central nervous system (CNS) occurs in several neurodegenerative/neuroinflammatory conditions and may cause neurochemical changes to endogenous neuroprotective systems. Pituitary adenylate cyclase-activating polypeptide (PACAP) and vasoactive intestinal polypeptide (VIP) are two neuropeptides with well-established protective and anti-inflammatory properties. Yet, whether PACAP and VIP levels are altered in mice with CNS-restricted, astrocyte-targeted production of IL-6 (GFAP-IL6) remains unknown. In this study, PACAP/VIP levels were assessed in the brain of GFAP-IL6 mice. In addition, we utilised bi-genic GFAP-IL6 mice carrying the human sgp130-Fc transgene (termed GFAP-IL6/sgp130Fc mice) to determine whether trans-signalling inhibition rescued PACAP/VIP changes in the CNS. Transcripts and protein levels of PACAP and VIP, as well as their receptors PAC1, VPAC1 and VPAC2, were significantly increased in the cerebrum and cerebellum of GFAP-IL6 mice vs. wild type (WT) littermates. These results were paralleled by a robust activation of the JAK/STAT3, NF-κB and ERK1/2MAPK pathways in GFAP-IL6 mice. In contrast, co-expression of sgp130Fc in GFAP-IL6/sgp130Fc mice reduced VIP expression and activation of STAT3 and NF-κB pathways, but it failed to rescue PACAP, PACAP/VIP receptors and Erk1/2MAPK phosphorylation. We conclude that forced expression of IL-6 in astrocytes induces the activation of the PACAP/VIP neuropeptide system in the brain, which is only partly modulated upon IL-6 trans-signalling inhibition. Increased expression of PACAP/VIP neuropeptides and receptors may represent a homeostatic response of the CNS to an uncontrolled IL-6 synthesis and its neuroinflammatory consequences.

Nicotinamide mononucleotide mitigates neuroinflammation by enhancing GPX4-mediated ferroptosis defense in microglia

BackgroundNumerous neurological diseases involving neuroinflammation, particularly microglia, contribute to neuronal death. Ferroptosis is implicated in various diseases characterized by neuronal injury. Studies showed that nicotinamide mononucleotide (NMN) inhibits both neuroinflammation and ferroptosis. However, the mechanisms of NMN in both ferroptosis and neuroinflammation remain unclear. We aimed to explore the effects of NMN on neuroinflammation and the susceptibility of microglia to ferroptosis. MethodsFerroptosis markers in macroglia exposed to lipopolysaccharides (LPS) were analyzed using CCK8, flow cytometry, ELISA, and quantitative RT-PCR. The effects of NMN on LPS-induced ferroptosis in microglia were evaluated through flow cytometry, western blot, and immunofluorescence staining. RT-PCR analysis assessed the inflammatory cytokine production of microglia subjected to Ferrostatin-1-regulated ferroptosis. RNA sequencing elucidated the underlying mechanism of NMN-involved microglia ferroptosis under LPS induction. In BV2 microglia, an inhibitor of GPX4, RSL3, was employed to suppress GPX4 expression. Intracerebroventricular injection of LPS was performed to evaluate neuroinflammation and microglia activation in vivo. ResultsNMN effectively rescued LPS-induced ferroptosis and improved cell viability in microglia. Co-administration of NMN and ferrostatin-1 significantly reduced proinflammatory cytokine production in microglia following the introduction of LPS stimuli. Mechanistically, NMN facilitated glutathione (GSH) production, and enhanced resistance to lipid peroxidation occurred in a manner dependent on GPX4, repressing cytokine transcription and protecting cells from ferroptosis. RNA sequencing elucidated the underlying mechanism of NMN-associated microglia ferroptosis under LPS induction. Furthermore, simultaneous injection of NMN ameliorated LPS-induced ferroptosis and neuroinflammation in mouse brains. The data from the present study indicated that NMN enhances GPX4-mediated ferroptosis defense against LPS-induced ferroptosis in microglia by recruiting GSH, thereby inhibiting neuroinflammation. ConclusionTherapeutic approaches to effectively target ferroptosis in diseases using NMN, consideration should be given to both its anti-ferroptosis and anti-inflammatory effects to attain optimal outcomes, presenting promising strategies for treating neuroinflammation-related diseases or disorders.

1581: IGRT workflow for pre-clinical proton research and MRI-based response assessment in mouse brain

1581: IGRT workflow for pre-clinical proton research and MRI-based response assessment in mouse brain

Integrated multi-omics approaches reveal the neurotoxicity of triclocarban in mouse brain

Triclocarban (TCC) is an antimicrobial ingredient that commonly incorporated in many household and personal care products, raising public concerns about its potential health risks. Previous research has showed that TCC could cross the blood–brain barrier, but to date our understanding of its potential neurotoxicity at human-relevant concentrations remains lacking. In this study, we observed anxiety-like behaviors in mice with continuous percutaneous exposure to TCC. Subsequently, we combined lipidomic, proteomic, and metabolic landscapes to investigate the underlying mechanisms of TCC-related neurotoxicity. The results showed that TCC exposure dysregulated the proteins involved in endocytosis and neurodegenerative disorders in mouse cerebrum. Brain energy homeostasis was also altered, as evidenced by the perturbation of pyruvate metabolism, TCA cycle, and oxidative phosphorylation, which in turn caused mitochondrial dysfunction. Meanwhile, the changing trends of sphingolipid signaling pathway and overproduction of mitochondrial reactive oxygen species (mROS) could enhance the neural apoptosis. The in vitro approach further demonstrated that TCC exposure promoted apoptosis, accompanied by the overproduction of mROS and alteration in the mitochondrial membrane potential in N2A cells. Together, dysregulated endocytosis, mROS-related mitochondrial dysfunction and neural cell apoptosis are considered to be crucial factors for TCC-induced neurotoxicity, which may contribute to the occurrence and development of neurodegenerative disorders. Our findings provide novel perspectives for the mechanisms of TCC-triggered neurotoxicity.

Comprehensive Analysis of Age- and Sex-Related Expression of the Chaperone Protein Sigma-1R in the Mouse Brain

Comprehensive Analysis of Age- and Sex-Related Expression of the Chaperone Protein Sigma-1R in the Mouse Brain