Neuroinflammatory State Research Articles

P2X7 Receptor Antagonism as a Potential Therapy in Amyotrophic Lateral Sclerosis.

This review focuses on the purinergic ionotropic receptor P2X7 (P2X7R) as a potential target for developing drugs that delay the onset and/or disease progression in patients with amyotrophic lateral sclerosis (ALS). Description of clinical and genetic ALS features is followed by an analysis of advantages and drawbacks of transgenic mouse models of disease based on mutations in a bunch of proteins, particularly Cu/Zn superoxide dismutase (SOD1), TAR-DNA binding protein-43 (TDP-43), Fused in Sarcoma/Translocated in Sarcoma (FUS), and Chromosome 9 open reading frame 72 (C9orf72). Though of limited value, these models are however critical to study the proof of concept of new compounds, before reaching clinical trials. The authors also provide a description of ALS pathogenesis including protein aggregation, calcium-dependent excitotoxicity, dysfunction of calcium-binding proteins, ultrastructural mitochondrial alterations, disruption of mitochondrial calcium handling, and overproduction of reactive oxygen species (ROS). Understanding disease pathogenic pathways may ease the identification of new drug targets. Subsequently, neuroinflammation linked with P2X7Rs in ALS pathogenesis is described in order to understand the rationale of placing the use of P2X7R antagonists as a new therapeutic pharmacological approach to ALS. This is the basis for the hypothesis that a P2X7R blocker could mitigate the neuroinflammatory state, indirectly leading to neuroprotection and higher motoneuron survival in ALS patients.

Characterization of Immune Cell-derived Extracellular Vesicles and Studying Functional Impact on Cell Environment.

The neuroinflammatory state of the central nervous system (CNS) plays a key role in physiological and pathological conditions. Microglia, the resident immune cells in the brain, and sometimes the infiltrating bone marrow-derived macrophages (BMDMs), regulate the inflammatory profile of their microenvironment in the CNS. It is now accepted that the extracellular vesicle (EV) populations from immune cells act as immune mediators. Thus, their collection and isolation are important to identify their contents but also evaluate their biological effects on recipient cells. The present data highlight chronological requirements for EV isolation from microglia cells or blood macrophages including the ultracentrifugation and size-exclusion chromatography (SEC) steps. A non-targeted proteomic analysis permitted the validation of protein signatures as EV markers and characterized the biologically active EV contents. Microglia-derived EVs were also functionally used on primary culture of neurons to assess their importance as immune mediators in the neurite outgrowth. The results showed that microglia-derived EVs contribute to facilitate the neurite outgrowth in vitro. In parallel, blood macrophage-derived EVs were functionally used as immune mediators in spheroid cultures of C6 glioma cells, the results showing that these EVs control the glioma cell invasion in vitro. This report highlights the possibility to evaluate the EV-mediated immune cell functions but also understand the molecular bases of such a communication. This deciphering could promote the use of natural vesicles and/or the in vitro preparation of therapeutic vesicles in order to mimic immune properties in the microenvironment of CNS pathologies.

Positron Emission Tomography Imaging of Long-Term Expression of the 18 kDa Translocator Protein After Sudden Cardiac Arrest in Rats.

Knowledge about the neuroinflammatory state during months after sudden cardiac arrest is scarce. Neuroinflammation is mediated by cells that express the 18 kDa translocator protein (TSPO). We determined the time course of TSPO-expressing cells in a rat model of sudden cardiac arrest using longitudinal in vivo positron emission tomography (PET) imaging with the TSPO-specific tracer [18F]DAA1106 over a period of 6 months. Five male Sprague Dawley rats were resuscitated from 6 min sudden cardiac arrest due to ventricular fibrillation, three animals served as shams. PET measurements were performed on day 5, 8, 14, 90, and 180 after intervention. Magnetic resonance imaging was performed on day 140. Imaging was preceded by Barnes Maze spatial memory testing on day 3, 13, 90, and 180. Specificity of [18F]DAA1106 binding was confirmed by Iba-1 immunohistochemistry. [18F]DAA1106 accumulated bilaterally in the dorsal hippocampus of all sudden cardiac arrest animals on all measured time points. Immunohistochemistry confirmed Iba-1 expressing cells in the hippocampal CA1 region. The number of Iba-1-immunoreactive objects per mm2 was significantly correlated with [18F]DAA1106 uptake. Additionally, two of the five sudden cardiac arrest rats showed bilateral TSPO-expression in the striatum that persisted until day 180. In Barnes Maze, the relative time spent in the target quadrant negatively correlates with dorsal hippocampal [18F]DAA1106 uptake on day 14 and 180. After sudden cardiac arrest, TSPO remains expressed over the long-term. Sustainable treatment options for neuroinflammation may be considered to improve cognitive functions after sudden cardiac arrest.

Stimulation of toll-like receptor 4 downregulates the expression of α7 nicotinic acetylcholine receptors via histone deacetylase in rodent microglia

Microglia have both protective and degenerative roles in the central nervous system. The α7 nicotinic acetylcholine receptor (nAChR) is crucial in the regulation of the neuroprotective role in microglia. Recent studies have demonstrated decreased expression of α7 nAChR in brain in response to neuroinflammation, but the mechanism mediating the downregulation of the α7 nAChR has yet to be elaborated. Treatment of microglial cell line BV2 cells or rat primary cultured microglia with the inflammogen lipopolysaccharide (LPS) significantly decreased the expression of α7 nAChR mRNA in a time and concentration-dependent manner. The effects of LPS were prevented by pretreatment with TAK-242, a toll-like receptor 4 (TLR4) blocker. The LPS-induced downregulation of α7 nAChR was also prevented with trichostatin A, a histone deacetylase (HDAC) inhibitor, but not 5-aza-2′-deoxycytidine, a DNA methyltransferase inhibitor. Further pharmacological probing revealed that HDAC2 and HDAC3 were involved in the effects of LPS. Treatment of BV2 cells with LPS significantly reduced acetylation of histone H3 at lysine 9 of the α7 nAChR promoter. The current findings demonstrate that inflammation-evoked activation of TLR4 leads to the reduction of the neuroprotective function of microglia through the downregulation of the α7 nAChR. Also, histone modification could be crucial in the regulation of the neuroprotective role of microglia during neuroinflammatory states.

Brain amyloid in virally suppressed HIV-associated neurocognitive disorder.

ObjectiveTo determine whether virally suppressed HIV neuropathogenesis, a chronic neuroinflammatory state, promotes abnormal brain amyloid deposition.MethodsA total of 10 men with virally suppressed HIV-associated neurocognitive disorder (HAND), aged 46–68 years, underwent 11C-labeled Pittsburgh compound B PET. Data from the Australian Imaging, Biomarkers and Lifestyle (AIBL), including 39 cognitively normal individuals (aged 60–74 years), 7 individuals with mild cognitive impairment (MCI) (aged 64–71 years), and 11 individuals with Alzheimer disease (AD) (aged 55–74 years), were used as reference. Apart from more women, the AIBL cohort was demographically comparable with the HIV sample. Also, the AIBL PET data did not differ by sex. Cerebellum standardized uptake value ratio amyloid values within 22 regions of interest were estimated. In the HIV sample, apolipoprotein E (APOE) was available in 80%, CSF biomarkers in 60%, and 8–10 years of long-term health outcomes in 100%.ResultsHAND and the AIBL group with no cognitive deficits had similar amyloid deposition, which was lower than that in both the MCI and AD groups. At the individual level, one HAND case showed high amyloid deposition consistent with AD. This case also had a CSF-AD–like profile and an E4/E4 for APOE. Clinically, this case declined over 18 years with mild HAND symptoms first, followed by progressive memory decline 8–9 years after the study PET, then progression to severe dementia within 2–3 years, and lived a further 6 years. Another HAND case showed increased amyloid deposition restricted to the hippocampi. Two other HAND cases showed abnormally decreased amyloid in subcortical areas.ConclusionsRelative to cognitively normal older controls, brain amyloid burden does not differ in virally suppressed HAND at the group level. However, individual analyses show that abnormally high and low amyloid burden occur.

Mast cells as mediators of astrocyte morphology associated with neuroinflammatory disease states

Abstract Morphological deficits observed in astrocytes, the most abundant glial cell type in the central nervous system, are associated with various neuroinflammatory diseases. The morphological and molecular astrocyte changes associated with multiple sclerosis are well-characterized and concentrated in the white matter of the brain. While the exact mechanisms behind this autoimmune disease are not fully understood, mast cells play a key role in the progression of white matter demyelination. We hypothesize mast cells orchestrate morphological astrocyte changes that are associated with neuroinflammatory disease states. To test this hypothesis, we used GFAP as a marker to visualize astrocyte morphology in white matter tracts of wild type (WT) and mast cell deficient KitW–sh/W–sh (sash) mice. Using skeleton analysis, we quantified astrocyte characteristics such as number of endpoints, branches, and branch length. Compared with WT mice, sash mice exhibited fewer branches (77 vs. 54 branches/cell) and endpoints per cell (49.5 vs. 24.5 P<0.001). These findings suggest that mast cells may drive altered astrocyte morphology and function in white matter. In order to determine whether this effect is due to altered development, studies using sash mice at time points across the lifespan are in progress. In addition, we are using diphtheria toxin in combination with novel genetic lines to ablate mast cells acutely in adult mice. Along with new molecular techniques, this will allow us to better characterize the interaction between mast cells and astrocytes in the adult and developing brain. Unraveling this interplay will be critical to understanding the role of mast cells in neurodevelopment and neuroinflammatory diseases such as multiple sclerosis.

Cytotoxic T cells infiltrate the brain and interact with microglia to reduce Alzheimer’s Disease pathogenesis

Abstract The role of the adaptive immune system, specifically T cells, remains poorly understood in Alzheimer’s disease (AD) pathogenesis research. Our lab generated an immune-deficient mouse model of AD (RAG2ɣc−/−5xfAD) which showed a dramatic increase in amyloid plaque load, an activated microglial neuroinflammatory state, and a decrease in the ability of microglia to phagocytose beta-amyloid. Restoration of adaptive immunity via bone marrow transplantation led to a corresponding decrease in amyloid plaques, indicating a fundamental role for adaptive immunity in AD pathology. Further adoptive transfers into these aged immune-deficient mice showed a significant increase in memory CD8+ T cells and microglia in the RAG2ɣc−/−5xfAD brain parenchyma via flow cytometry. Interestingly, T cells were observed adjacent to plaque-associated microglia in these brains via Immunohistochemistry. To determine whether T cell infiltration also occurs in immune-intact AD mice, flow cytometry was conducted on 5xfAD mice that develop amyloid plaques, PS19 mice that develop neurofibrillary tangles, PS19-5xfAD bigenic mouse model, and wild-type mice. This analysis validated the immune-deficient observations, demonstrating a significant increase in memory CD8+ T cells in 5xfAD mice brains but also revealed a further significant increase in the bigenic PS19-5xfAD model. Therefore, I hypothesize that cytotoxic T cells respond to the inflammatory AD environment by infiltrating the brain and directly interacting with microglia to decrease amyloid plaques. My initial data clearly demonstrate that further examination of the crosstalk between adaptive and innate immunity is needed to determine the mechanisms by which T cells influence AD pathogenesis.

Corticosterone Induces HMGB1 Release in Primary Cultured Rat Cortical Astrocytes: Involvement of Pannexin-1 and P2X7 Receptor-Dependent Mechanisms.

A major risk factor for major depressive disorder (MDD) is stress. Stress leads to the release of high-mobility group box-1 (HMGB1), which in turn leads to neuroinflammation, a potential pathophysiological basis of MDD. The mechanism underlying stress-induced HMGB1 release is not known, but stress-associated glucocorticoids could be involved. To test this, rat primary cultured cortical astrocytes, the most abundant cell type in the central nervous system (CNS), were treated with corticosterone and HMGB1 release was assessed by Western blotting and ELISA. Significant HMGB1 was released with treatment with either corticosterone or dexamethasone, a synthetic glucocorticoid. HMGB1 translocated from the nucleus to the cytoplasm following corticosterone treatment. HMGB1 release was significantly attenuated with glucocorticoid receptor blocking. In addition, inhibition of pannexin-1, and P2X7 receptors led to a significant decrease in corticosterone-induced HMGB1 release. Taken together, corticosterone stimulates astrocytic glucocorticoid receptors and triggers cytoplasmic translocation and extracellular release of nuclear HMGB1 through a mechanism involving pannexin-1 and P2X7 receptors. Thus, under conditions of stress, glucocorticoids induce astrocytic HMGB1 release, leading to a neuroinflammatory state that could mediate neurological disorders such as MDD.

Sex‐dependent Effects of High‐fat Diet and Stress on CNS Inflammation and Ceramides Production

The intake of a high‐fat diet (HFD) and exposition to stress are associated with development of a neuroinflammatory state in the central nervous system (CNS). Previously we found high levels of long‐chain (C18:0 and C20:0) ceramides in blood of rats fed an HFD that correlated with increased immobility in the forced swim test, suggesting that ceramides could contribute to the development of depressive‐like behavior. In the current study, we aimed to determine whether adding short‐term unpredictable stress (STUS) would exacerbate the effects of HFD on neuroinflammation and ceramide metabolism in the ventral hippocampus (VH) and the development of depressive‐like behavior. We hypothesized that exposure to HFD and STUS would cause a neuroinflammatory state and increased levels of long‐chained ceramides in the VH leading to depressive‐like behavior. To test this, male and female Sprague Dawley rats were administered either a low‐fat or high‐fat diet for eight weeks, while other groups of female and male rats were administered a low‐fat or high‐fat diet for 10 weeks with the inclusion of 4 days of random unpredictable stressors. After sacrifice, we assessed the expression of microglial (Iba‐1), neuroinflammatory state (NLRP3, HMGB1 and TNF‐α) and ceramide production markers (CerS5, CerS6, SMPD1 and SMPD3) in the VH. Female HFD rats showed increased expression of TNF‐α and HMGB‐1 independent of STUS, while male rats increased expression of Iba‐1, TNF‐α and NLRP3 after exposition to STUS, suggesting a sex‐dependent activation of inflammatory mediators. The assessment of enzymes that produce long‐chained ceramides revealed increased CerS5, CerS6 and SMPD1 mRNA only in the VH of female HFD rats exposed to STUS, suggesting sex differences in the production of long‐chained ceramides due to combined HFD and STUS. VH tissue and blood were collected to quantify ceramides by gas chromatography‐mass spectrometry (GC‐MS). Based on the expression of the enzymes involved in the production of ceramides, we expect that the GC‐MS assay will show increased C16 ceramides in female rats exposed to HFD and STUS. After diet completion, rats were subjected to sucrose grooming test and forced swim test to assess anhedonia and behavioral despair. Female HFD with STUS showed trending increases in grooming activity, reduced immobility and increased swimming time while male rats did not present any behavioral changes, suggesting less depressive‐like behavior in female rats. In conclusion, an HFD in combination with STUS causes sex‐dependent changes inflammatory state and long‐chained ceramides production without increasing depressive‐like behavior.Support or Funding InformationNIH‐NIGMS R25GM082406 (PHSU RISE Graduate Training Program), U54 MD007579, NIGMS/INBRE P20 GM103475, and NIH‐NIGMS R25GM096955 (UPR‐Ponce PRISE Program).

Effects of a Single Head Exposure to GSM-1800 MHz Signals on the Transcriptome Profile in the Rat Cerebral Cortex: Enhanced Gene Responses Under Proinflammatory Conditions.

Mobile communications are propagated by electromagnetic fields (EMFs), and since the 1990s, they operate with pulse-modulated signals such as the GSM-1800 MHz. The biological effects of GSM-EMF in humans affected by neuropathological processes remain seldom investigated. In this study, a 2-h head-only exposure to GSM-1800 MHz was applied to (i) rats undergoing an acute neuroinflammation triggered by a lipopolysaccharide (LPS) treatment, (ii) age-matched healthy rats, or (iii) transgenic hSOD1G93A rats that modeled a presymptomatic phase of human amyotrophic lateral sclerosis (ALS). Gene responses were assessed 24 h after the GSM head-only exposure in a motor area of the cerebral cortex (mCx) where the mean specific absorption rate (SAR) was estimated to be 3.22 W/kg. In LPS-treated rats, a genome-wide mRNA profiling was performed by RNA-seq analysis and revealed significant (adjusted p value < 0.05) but moderate (fold changes < 2) upregulations or downregulations affecting 2.7% of the expressed genes, including genes expressed predominantly in neuronal or in glial cell types and groups of genes involved in protein ubiquitination or dephosphorylation. Reverse transcription-quantitative PCR analyses confirmed gene modulations uncovered by RNA-seq data and showed that in a set of 15 PCR-assessed genes, significant gene responses to GSM-1800 MHz depended upon the acute neuroinflammatory state triggered in LPS-treated rats, because they were not observed in healthy or in hSOD1G93A rats. Together, our data specify the extent of cortical gene modulations triggered by GSM-EMF in the course of an acute neuroinflammation and indicate that GSM-induced gene responses can differ according to pathologies affecting the CNS.

Depressive-like phenotype evoked by lifelong nutritional omega-3 deficiency in female rats: Crosstalk among kynurenine, Toll-like receptors and amyloid beta oligomers

Depression is one of the most common psychiatric diseases and the prevalence of depressive symptoms in women is almost twice compared to men, although the reasons of this gender difference are not fully understood yet. Recently, soluble amyloid beta (Aβ)1-42 peptide has been receiving great importance in the development of depression, also considering that depression is highly comorbid with Alzheimer’s disease and other neurodegenerative illnesses. The central role played by Aβ in the development of depressive-like symptoms in rodents has been evidenced in environmental rodent model of depression. Indeed, we have previously found that lifelong exposure to n-3 polyunsaturated fatty acids (PUFA) deficient diet in female rats at 8 weeks of life leads to depressive like- symptoms and higher susceptibility to stress associated with increased Aβ levels. In order to understand if such effects were maintained over time, rats were exposed to the same diet regimen until 6 or 21 weeks of life. We found that both timepoints of exposure to n-3 PUFA deficient diet lead to depressive-like phenotype. Furthermore, a significant alteration in brain neurochemistry was retrieved. In particular, in hippocampal area a significant reduction in serotonin (5-HT) and noradrenaline (NA) content was evidenced. Considering the prominent role of NA in counterbalancing neuroinflammatory state, we quantified in the same brain area kynurenine levels, a metabolite of tryptophan implicated in inflammatory state and brought to the fore for its implication in depression. Interestingly, kynurenine levels were significantly increased in hippocampus (HIPP) of female rats exposed to such diet. In addition, lifelong deficiency in n-3 PUFA dietary intake led to systemic increase of corticosterone, hence hypothalamic pituitary adrenal (HPA) axis hyperactivation, and higher proinflammatory cytokine production. Increased production of kynurenine, along with HPA axis hyperactivation, have been associated with immune system modulation, particularly through Toll-like receptor type 2 (TLR2) and Toll-like receptor type 4 (TLR4) involvement. In addition, it has been shown that soluble forms of Aβ1-42 can induced depressive like-phenotype in consequence to a crosstalk between TLR4 and 5-HTergic system. Thus, considering that in this model we have previously reported increased plasma Aβ1-42 level, we quantified TRL2 and 4 expression in HIPP of treated rats. We found that chronic exposure to a diet characterized by very low n-3 PUFA content led to higher expression of TLR2 and TLR4 in HIPP of female treated rats, indicating an activation of the immune system and was accompanied by increased expression of oligomeric Aβ.Taken together, our data indicate that the pro-depressive effects induced by a diet poor in n-3 PUFA can be attributable to a shift of hippocampal tryptophan metabolism toward inflammatory metabolite ultimately corresponding to altered immune response and increased Aβ oligomerization.

CXCL12-induced rescue of cortical dendritic spines and cognitive flexibility.

Synaptodendritic pruning is a common cause of cognitive decline in neurological disorders, including HIV-associated neurocognitive disorders (HAND). HAND persists in treated patients as a result of chronic inflammation and low-level expression of viral proteins, though the mechanisms involved in synaptic damage are unclear. Here, we report that the chemokine CXCL12 recoups both cognitive performance and synaptodendritic health in a rodent model of HAND, which recapitulates the neuroinflammatory state of virally controlled individuals and the associated structural/functional deficiencies. CXCL12 preferentially regulates plastic thin spines on layer II/III pyramidal neurons of the medial prefrontal cortex via CXCR4-dependent stimulation of the Rac1/PAK actin polymerization pathway, leading to increased spine density and improved flexible behavior. Our studies unveil a critical role of CXCL12/CXCR4 signaling in spine dynamics and cognitive flexibility, suggesting that HAND - or other diseases driven by spine loss - may be reversible and upturned by targeting Rac1-dependent processes in cortical neurons.

Isolation of microglia-derived extracellular vesicles: towards miRNA signatures and neuroprotection

The functional preservation of the central nervous system (CNS) is based on the neuronal plasticity and survival. In this context, the neuroinflammatory state plays a key role and involves the microglial cells, the CNS-resident macrophages. In order to better understand the microglial contribution to the neuroprotection, microglia-derived extracellular vesicles (EVs) were isolated and molecularly characterized to be then studied in neurite outgrowth assays. The EVs, mainly composed of exosomes and microparticles, are an important cell-to-cell communication process as they exhibit different types of mediators (proteins, lipids, nucleic acids) to recipient cells. The medicinal leech CNS was initially used as an interesting model of microglia/neuron crosstalk due to their easy collection for primary cultures. After the microglia-derived EV isolation following successive methods, we developed their large-scale and non-targeted proteomic analysis to (i) detect as many EV protein markers as possible, (ii) better understand the biologically active proteins in EVs and (iii) evaluate the resulting protein signatures in EV-activated neurons. The EV functional properties were also evaluated in neurite outgrowth assays on rat primary neurons and the RNAseq analysis of the microglia-derived EVs was performed to propose the most representative miRNAs in microglia-derived EVs. This strategy allowed validating the EV isolation, identify major biological pathways in EVs and corroborate the regenerative process in EV-activated neurons. In parallel, six different miRNAs were originally identified in microglia-derived EVs including 3 which were only known in plants until now. The analysis of the neuronal proteins under the microglial EV activation suggested possible miRNA-dependent regulation mechanisms. Taken together, this combination of methodologies showed the leech microglial EVs as neuroprotective cargos across species and contributed to propose original EV-associated miRNAs whose functions will have to be evaluated in the EV-dependent dialog between microglia and neurons.

Xiaoyao Pills Prevent Lipopolysaccharide-Induced Depression by Inhibiting Inflammation and Protecting Nerves.

Lipopolysaccharides are pro-inflammation mediators that can induce inflammation in the serum, hippocampus, and cortex of animals. And lipopolysaccharide-induced neuroinflammatory state resulted in significant depression-like behaviors, including reduced locomotor activity in the open field test, reduced saccharin preference, added immobility time in tail suspension test and forced swimming test, decreased comb time in the splash test, and increased latency to food in the novelty suppressed feeding test time, and reduced the levels of neurotrophic factors and synaptic proteins, and decreased Nissl bodies. Treatment with Xiaoyao Pills ameliorated the depression-like behavior, decreased the levels of inflammatory indicators, increased those of neurotrophic factors and synaptic proteins, and restored Nissl bodies. Our study suggests that lipopolysaccharides induce inflammation and nerve injury, thereby leading to depression. Xiaoyao Pills could be considered a potential therapeutic candidate for inflammation-induced depression.

Sex-specific effects of prenatal valproic acid exposure on sociability and neuroinflammation: Relevance for susceptibility and resilience in autism

Autism spectrum disorder (ASD) is a group of neurodevelopmental disorders with an incidence four times higher in boys than in girls. By analyzing the effect of sex in a mouse model of ASD, we were able to identify immune alterations that could underlie this sex bias.Pregnant mice were injected subcutaneously with 600 mg/kg of valproic acid (VPA) or saline at gestational day 12.5. Their male and female offspring were evaluated in a social interaction test at adulthood, and only male VPA mice showed reduced sociability levels and a lack of preference for the social stimulus over a novel object. We then analyzed the corticosterone (CORT) response to an inflammatory stimulus, as a measure of the hypothalamus-pituitary-adrenal (HPA) function, and the neuroinflammatory state in adult and young animals. Adult VPA males exhibited increased basal CORT levels, while VPA females showed levels comparable to controls. As male mice showed a blunted CORT response at PD21 when compared to female mice, we propose that this early dimorphism could explain the different effects of VPA on HPA function. In addition, prenatal VPA exposure resulted in altered astroglial and microglial cell density levels in the cerebellum and dentate gyrus of adult mice. These neuroinflammatory effects were more pronounced in females than males, and appeared at early developmental stages. Hence, these postnatal glial density differences could underlie the behavioral alterations observed in adulthood, when only males show a social deficit.Our work contributes to the understanding of biological mechanisms affected by VPA on male and female rodents and shed light on the study of possible resilience mechanisms in the female population and/or susceptibility to ASD in boys.

Differential fracture response to traumatic brain injury suggests dominance of neuroinflammatory response in polytrauma

Polytraumatic injuries, specifically long bone fracture and traumatic brain injury (TBI), frequently occur together. Clinical observation has long held that TBI can accelerate fracture healing, yet the complexity and heterogeneity of these injuries has produced conflicting data with limited information on underlying mechanisms. We developed a murine polytrauma model with TBI and fracture to evaluate healing in a controlled system. Fractures were created both contralateral and ipsilateral to the TBI to test whether differential responses of humoral and/or neuronal systems drove altered healing patterns. Our results show increased bone formation after TBI when injuries occur contralateral to each other, rather than ipsilateral, suggesting a role of the nervous system based on the crossed neuroanatomy of motor and sensory systems. Analysis of the humoral system shows that blood cell counts and inflammatory markers are differentially modulated by polytrauma. A data-driven multivariate analysis integrating all outcome measures showed a distinct pathological state of polytrauma and co-variations between fracture, TBI and systemic markers. Taken together, our results suggest that a contralateral bone fracture and TBI alter the local neuroinflammatory state to accelerate early fracture healing. We believe applying a similar data-driven approach to clinical polytrauma may help to better understand the complicated pathophysiological mechanisms of healing.

P3‐055: SYSTEMS BIOLOGY PLATFORM TO ENABLE NEUROINFLAMMATORY TARGET ID, VALIDATION, AND PATIENT SELECTION FOR DISEASE‐MODIFYING ALZHEIMER'S DISEASE THERAPEUTICS

Chronic neuroinflammation is a hallmark of Alzheimer's disease and other neurodegenerative disorders.Microglial dysfunction is likely a significant contributor to the onset and progression of Alzheimer's disease. The elucidation of non-innate immune functions of microglia in CNS development have revealed new therapeutic strategies for how microglial function might be modulated to protect the aging brain. In vitroassays based on isolated microglia do not fully model the neuroinflammatory environment that arises during the progression of Alzheimer's disease. Therefore, a predictive methodology for identification and validation of microglia targets in the neurodegenerative brain has remained elusive. Tiaki Therapeutics systems biology platform models chronic neuroinflammation associated with Alzheimer's disease and other neurodegenerative disorders. One integral part of the system is an brain slice assay, which permits the longitudinal analysis of all CNS cell types within their authentic matrix and inter-cellular environment. RNA expression profiles were obtained for bulk tissue and isolated microglia to establish the transcriptomic signature of neuroinflammation in the platform. Bioinformatics was used to compare the neuroinflammatory signature of the platform to transcriptomic data from patients. The initial implementation of the Tiaki platform indicated that after an initial period of dynamic changes in gene expression, the platform exhibits a stable neuroinflammatory state that can be detected and quantified by transcriptomic analyses. The gene expression signature observed in the platform was significantly correlated with gene expression data from Alzheimer's patients. Tiaki has developed a novel approach to authentically model chronic neuroinflammation that aligns with Alzheimer's disease gene expression in order to develop disease-modifying therapeutics against microglia targets that demonstrate beneficial effects on neurons. The assay is uniquely positioned to provide longitudinal genomic, as well as both target engagement and efficacy, biomarkers with high translational value.

Single Neuron and Microglia Gene Expression Networks Demonstrate Cellular Subphenotype Shifts and Altered Glial‐Neuronal Signaling in Solitary Nucleus during Alcohol Withdrawal: A Time Series

Alcohol use disorder remains a major societal and healthcare challenge motivating investigation into novel mechanisms of alcohol addiction. Negative‐reinforcement models of addiction postulate that avoidance of negative physical and emotional symptoms during withdrawal drive drug‐seeking. Alcohol withdrawal syndrome is characterized by autonomic dysregulation and negative emotion implicating the strongly connected amygdala and solitary nucleus (NTS) involved in autonomic inputs, outputs, and emotional states in the pathophysiology of alcohol withdrawal. Our previous work demonstrates a neuroinflammatory state in the amygdala during alcohol withdrawal, which we speculate contributes to the negative physical and emotional withdrawal symptoms and consequently drug‐seeking. Here, we investigated the effect of alcohol withdrawal on gene expression in single glucagon‐like peptide 1 (GLP‐1) neurons, adrenergic neurons, and microglia in the NTS. GLP‐1 neurons in this region receive autonomic inputs via vagal afferents and signal to amygdala neurons co‐expressing GLP‐1 receptors and corticotropin‐releasing factor. NTS adrenergic neurons respond to higher‐order inputs to maintain cardiovascular homeostasis. Control, ethanol‐dependent, and rats undergoing ethanol withdrawal at 8, 32, and 176 hour time points were sampled. Single cells were harvested using using laser‐capture microdissection, and microfluidic RT‐qPCR was employed to generate a high‐dimensional dataset. Cellular subphenotypes were characterized and local glial‐neuronal signaling networks were described based on gene networks that defined these subphenotypes. We found an increase in inflammatory subphentoypes in the 8 and 32 hour withdrawal condition with the 176 hour withdrawal condition demonstrating decreased GABA receptor expression. Preproglucagon (Gcg), the GLP‐1 precursor transcript, was found to be upregulated only in alcohol withdrawal conditions. The levels of this transcript opposed levels of the norepinephrine precursor tyrosine hydroxylase transcript suggesting these two signaling ligands have oppositional regulation in alcohol withdrawal.Support or Funding InformationU01HL133360‐01This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Soursop fruit extract mitigates scopolamine-induced amnesia and oxidative stress via activating cholinergic and Nrf2/HO-1 pathways.

Current therapeutic interventions for memory loss are inadequate and are associated with numerous adverse effects. There is an urgent need for new alternative agents for the treatment of memory loss and related disorders. Here, we investigated the potential neuroprotective role of soursop fruit extract (SSFE) in scopolamine (SCO)-induced amnesia and oxidative damage in the hippocampus of rats. Thirty-five rats were randomly allocated into 5 groups: control, SCO, SSFE, SCO, SSFE+SCO and N-acetylcysteine (NAC) + SCO. SCO-treatment increased acetylcholine esterase activity and decreased hippocampal levels of acetylcholine, serotonin, dopamine, norepinephrine, and histamine. The level of ATP increased. SCO-treated rats showed a disturbance in oxidative status, which was evident through the increase in malondialdehyde, and nitrites/nitrates and a decrease in cellular antioxidant molecules including glutathione, superoxide dismutase, catalase, glutathione reductase, and glutathione peroxidase. A disturbance was also observed via downregulation of the nuclear factor erythroid 2-related factor 2 and heme oxygenase-1 defense pathways. SCO-treatment enhances a neuroinflammatory state, as indicated by the release of tumor necrosis factor- α and interleukin-1β and increased inducible nitric oxide synthase and mRNA expression. SCO-treatment decreased the expression of the anti-apoptotic protein, B cell lymphoma 2 and increased the expression of the pro-apoptotic protein, Bcl-2 associated X protein, caspase-3 and cytochrome c in hippocampal neurons. SSFE pretreatment markedly ameliorated hippocampal changes. Our findings revealed that SSFE exerts its potential anti-amnestic effect mainly through the activation of the cholinergic system and Nrf2/HO-1 pathway.

Indirubin-3′-monoxime prevents aberrant activation of GSK-3β/NF-κB and alleviates high fat-high fructose induced Aβ-aggregation, gliosis and apoptosis in mice brain

Deciphering the molecular mechanisms of amyloid pathology and glial cell-mediated neuroinflammation, offers a novel avenue for therapeutic intervention against neurodegeneration. Recent findings demonstrate a crucial link between activation of glycogen synthase kinase-3β (GSK-3β), amyloid deposition and a neuroinflammatory state. However, studies demonstrating the pharmacological effects of GSK-3β inhibition and the interlinked molecular mechanisms still remain elusive. The present study explores whether high fat-high fructose diet (HFFD)-induced neuropathological changes could be alleviated by indirubin-3′-monoxime (IMX), a GSK-3β inhibitor. Male Swiss albino mice (8 weeks old) were fed with normal pellet or HFFD for 60 days. HFFD mice were treated with IMX once daily for last 7 days of the experimental period. HFFD fed-mice had significant amyloid deposits in cerebral cortex and hippocampus, and protein expression analyses showed activation of GSK-3β, nuclear translocation of NF-κB p65 and upregulation of inflammatory (TNF-α, IL-6, COX-2), astrocytic (GFAP), glial surface (CD-68) and pro-apoptotic markers (Bax and caspase-3). IMX treatment promotes the inhibitory phosphorylation of GSK-3β at Ser9 and moreover, a marked reduction in the phosphorylation of IKK-β, which prevents translocation and activation of NF-κB. Protein expression studies in IMX-treated brain tissues positively correlate with the anti-neuroinflammatory effects of GSK-3β inhibition. Taken together, our results provide substantial evidence that IMX could potentially attenuate neuroinflammation in coordination with the master transcription factor-NF-κB.