Increases In GFAP Research Articles

Activation of astrocytic NMDA receptors counteracted Aβ-induced reduction of BDNF and elevation of GFAP and complement 3 in the hippocampal astrocytes

N-methyl-D-aspartate receptors (NMDARs) play a crucial role in mediating Amyloid-β (Aβ) synaptotoxicity. Our previous studies have demonstrated an opposite (neuroprotection and neurotoxicity) effect of activating astrocytic and neuronal NMDARs with higher dose (10 μM) of NMDA, an agonist of NMDARs. By contrast, activating neuronal or astrocyitc NMDARs with lower dose (1 μM) of NMDA both exerts neuroprotective effect in Aβ-induced neurotoxicity. However, the underlying mechanism of activating astrocytic NMDARs with lower dose of NMDA to protect against Aβ neurotoxicity remains unclear. Based on our previous related work, in this study, using a co-cultured cell model of primary hippocampal neurons and astrocytes, we further investigated the possible factors involved in 1 μM of NMDA activating astrocytic NMDARs to oppose Aβ-induced synaptotoxicity. Our results showed that activation of astrocytic NMDARs by 1 μM NMDA rescued Aβ-induced reduction of brain-derived neurotrophic factor (BDNF), and inhibited Aβ-induced increase of GFAP, complement 3 (C3) and activation of NF-κB. Furthermore, blockade of astrocytic GluN2A with TCN201 abrogated the ability of 1 μM NMDA to counteract the effects of Aβ decreasing BDNF, and increasing GFAP, C3 and activation of NF-κB. These findings suggest that activation of astrocytic NMDARs protect against Aβ-induced synaptotoxicity probably through elevating BDNF and suppressing GFAP and C3. Our present research provides valuable insights for elucidating the underlying mechanism of astrocytic NMDARs activation resisting the toxic effects of Aβ.

JR5558 mice are a reliable model to investigate subretinal fibrosis

Subretinal fibrosis is a major untreatable cause of poor outcomes in neovascular age-related macular degeneration. Mouse models of subretinal fibrosis all possess a degree of invasiveness and tissue damage not typical of fibrosis progression. This project characterises JR5558 mice as a model to study subretinal fibrosis. Fundus and optical coherence tomography (OCT) imaging was used to non-invasively track lesions. Lesion number and area were quantified with ImageJ. Retinal sections, wholemounts and Western blots were used to characterise alterations. Subretinal lesions expand between 4 and 8 weeks and become established in size and location around 12 weeks. Subretinal lesions were confirmed to be fibrotic, including various cell populations involved in fibrosis development. Müller cell processes extended from superficial retina into subretinal lesions at 8 weeks. Western blotting revealed increases in fibronectin (4 wk and 8 wk, p < 0.001), CTGF (20 wks, p < 0.001), MMP2 (12 wks and 20 wks p < 0.05), αSMA (12 wks and 20 wks p < 0.05) and GFAP (8 wk and 12 wk, p ≤ 0.01), consistent with our immunofluorescence results. Intravitreal injection of Aflibercept reduced subretinal lesion growth. Our study provides evidence JR5558 mice have subretinal fibrotic lesions that grow between 4 and 8 weeks and confirms this line to be a good model to study subretinal fibrosis development and assess treatment options.

B. glomerulata promotes neuroprotection against ischemic stroke by inhibiting apoptosis through the activation of PI3K/AKT/mTOR pathway

BackgroundBrassaiopsis glomerulata (Blum) Regel (B.glomerulata) is recognized as a traditional Chinese medicine (TCM) primarily used for promoting blood circulation and removing stasis. It is frequently utilized in the treatment of injuries resulting from falls and bumps. PurposeDespite its effective use in clinical treatment for ischemic stroke (IS), there are currently no reports on its composition and mechanism of action, which affects its promotion. The study investigated the chemical components and molecular mechanisms of B.glomerulata, with the following components: UPLC-Q-TOF-MS, network pharmacology Analysis and experimental verification in vivo and vitro. MethodsThe effect of B.glomerulata on interfering with ischemic stroke was assessed on MCAO/R rats and ORD cell model. Then the compositional analysis was conducted using UPLC-Q-TOF-MS. Furthermore, network pharmacology and molecular docking techniques were explored to identify potential targets and pathways. The predicted mechanisms of action were ultimately confirmed by immunohistochemistry and protein blotting. ResultsB. glomerulata exhibited neuroprotective effects in MCAO/R rats by reductions in hippocampal and cortical neuronal damage, brain infarction, and cerebral edema. Both in vivo and in vitro experiments demonstrated that it decreased ROS and MDA levels, increased SOD and GSH levels, thereby inhibiting oxidative stress. Moreover, the improvements in neuronal morphology and the modulation of Nissl bodies suggested a potential mechanism underlying its neuroprotective action. Additionally, B.glomerulata exhibited concentration-dependent reductions in Bax and Caspase-3 expressions, along with increases in GFAP, Bcl2/Bax ratio, p-PI3K, p-AKT, and p-mTOR levels. ConclusionB.glomerulata exhibited neuroprotective effects against cerebral ischemia-reperfusion injury both in vivo and in vitro. It prevented oxidative stress damage and inhibited apoptosis of ischemic stroke through the PI3K/AKT/mTOR pathway.

Aberrant Enhanced NLRP3 Inflammasomes and Cell Pyroptosis in the Brains of Prion-Infected Rodent Models Are Largely Associated with the Proliferative Astrocytes.

Neuroinflammation is a common pathological feature in a number of neurodegenerative diseases, which is mediated primarily by the activated glial cells. Nucleotide-binding oligomerization domain-like receptor pyrin domain-containing-3 (NLRP3) inflammasome-associated neuroinflammatory response is mostly considered. To investigate the situation of the NLRP3-related inflammation in prion disease, we assessed the levels of the main components of NLRP3 inflammasome and its downstream biomarkers in the scrapie-infected rodent brain tissues. The results showed that the transcriptional and expressional levels of NLRP3, caspase-1, and apoptosis-associated speck-like protein (ASC) in the brains of scrapie-infected rodents were significantly increased at terminal stage. The increased NLPR3 overlapped morphologically well with the proliferated GFAP-positive astrocytes, but little with microglia and neurons. Using the brain samples collected at the different time-points after infection, we found the NLRP3 signals increased in a time-dependent manner, which were coincidental with the increase of GFAP. Two main downstream cytokines, IL-1β and IL-18, were also upregulated in the brains of prion-infected mice. Moreover, the gasdermin D (GSDMD) levels, particularly the levels of GSDMD-NT, in the prion-infected brain tissues were remarkably increased, indicating activation of cell pyroptosis. The GSDMD not only co-localized well with the astrocytes but also with neurons at terminal stage, also showing a time-dependent increase after infection. Those data indicate that NLRP3 inflammasomes were remarkably activated in the infected brains, which is largely mediated by the proliferated astrocytes. Both astrocytes and neurons probably undergo a pyroptosis process, which may help the astrocytes to release inflammatory factors and contribute to neuron death during prion infection.

Gastrodia elata polysaccharide alleviates Parkinson's disease via inhibiting apoptotic and inflammatory signaling pathways and modulating the gut microbiota.

Parkinson's disease (PD) is a common, chronic, and progressive degenerative disease of the central nervous system for which there is no effective treatment. Gastrodia elata is a well-known food and medicine homologous resource with neuroprotective potential. Gastrodia elata polysaccharide (GEP), which is a highly active and safe component in Gastrodia elata, is an important ingredient in the development of functional products. In this study, GEP was administered to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced mice over 3 weeks to investigate its neuroprotective effects. The results showed that GEP significantly alleviated the motor dysfunction of PD mice, inhibited the accumulation of α-synuclein, and reduced the loss of dopaminergic neurons in the brain. Moreover, GEP increased the Bcl-2/Bax ratio and decreased the cleaved-caspase-3 level, suggesting that GEP may ameliorate PD by preventing MPTP-induced mitochondrial apoptosis. GEP also significantly inhibited the increase of GFAP and decreased the levels of TNF-α, IL-1β, and IL-6 in the brain of PD mice, which may be the result of the inhibition of neuroinflammation by the inactivation of the TLR4/NF-κB pathway. Furthermore, the neuroprotective effects of GEP involve the gut-brain axis, as it has been shown that GEP regulated the dysbiosis of PD-related gut microbiota such as Akkermansia, Lactobacillus, Bacteroides, Prevotella, and Faecalibacterium, increased the content of microbial metabolites SCFAs in the colon and increased the level of occludin that repairs the intestinal barrier of PD mice. In conclusion, this study is expected to provide a theoretical basis for the development and application of functional products with GEP from the perspective of neuroprotective effects.

Evaluating the effect of post-traumatic hypoxia on the development of axonal injury following traumatic brain injury in sheep

Damage to the axonal white matter tracts within the brain is a key cause of neurological impairment and long-term disability following traumatic brain injury (TBI). Understanding how axonal injury develops following TBI requires gyrencephalic models that undergo shear strain and tissue deformation similar to the clinical situation and investigation of the effects of post-injury insults like hypoxia. The aim of this study was to determine the effect of post-traumatic hypoxia on axonal injury and inflammation in a sheep model of TBI. Fourteen male Merino sheep were allocated to receive a single TBI via a modified humane captive bolt animal stunner, or sham surgery, followed by either a 15 min period of hypoxia or maintenance of normoxia. Head kinematics were measured in injured animals. Brains were assessed for axonal damage, microglia and astrocyte accumulation and inflammatory cytokine expression at 4 hrs following injury. Early axonal injury was characterised by calpain activation, with significantly increased SNTF immunoreactivity, a proteolytic fragment of alpha-II spectrin, but not with impaired axonal transport, as measured by amyloid precursor protein (APP) immunoreactivity. Early axonal injury was associated with an increase in GFAP levels within the CSF, but not with increases in IBA1 or GFAP+ve cells, nor in levels of TNFα, IL1β or IL6 within the cerebrospinal fluid or white matter. No additive effect of post-injury hypoxia was noted on axonal injury or inflammation. This study provides further support that axonal injury post-TBI is driven by different pathophysiological mechanisms, and detection requires specific markers targeting multiple injury mechanisms. Treatment may also need to be tailored for injury severity and timing post-injury to target the correct injury pathway.

Lomerizine inhibits LPS-mediated neuroinflammation and tau hyperphosphorylation by modulating NLRP3, DYRK1A, and GSK3α/β.

Lomerizine is a calcium channel blocker that crosses the blood-brain barrier and is used clinically in the treatment of migraines. However, whether lomerizine is beneficial in modulating neuroinflammatory responses has not been tested yet. To assess the potential of lomerizine for repurposing as a treatment for neuroinflammation, we investigated the effects of lomerizine on LPS-induced proinflammatory responses in BV2 microglial cells, Alzheimer's disease (AD) excitatory neurons differentiated from induced pluripotent stem cells (iPSCs), and in LPS-treated wild type mice. In BV2 microglial cells, lomerizine pretreatment significantly reduced LPS-evoked proinflammatory cytokine and NLRP3 mRNA levels. Similarly, lomerizine pretreatment significantly suppressed the increases in Iba-1, GFAP, proinflammatory cytokine and NLRP3 expression induced by LPS in wild-type mice. In addition, lomerizine posttreatment significantly decreased LPS-stimulated proinflammatory cytokine and SOD2 mRNA levels in BV2 microglial cells and/or wild-type mice. In LPS-treated wild-type mice and AD excitatory neurons differentiated from iPSCs, lomerizine pretreatment ameliorated tau hyperphosphorylation. Finally, lomerizine abolished the LPS-mediated activation of GSK3α/β and upregulation of DYRK1A, which is responsible for tau hyperphosphorylation, in wild-type mice. These data suggest that lomerizine attenuates LPS-mediated neuroinflammatory responses and tau hyperphosphorylation and is a potential drug for neuroinflammation- or tauopathy-associated diseases.

Revisiting the effects of exercise on cerebral neurovascular functions in rats using multimodal assessment techniques

Physical activity in the form of aerobic exercise has many beneficial effects on brain function. Here, we aim to revisit the effects of exercise on brain morphology and neurovascular organization using a rat running model. Electrocorticography (ECoG) was integrated with laser speckle contrast imaging (LSCI) and applied to simultaneously detect CSD propagation and the corresponding neurovascular function. In addition, blood oxygenation level-dependent (BOLD) signal in fMRI was used to observe cerebral utilization of oxygen. Results showed significant decrease in somatosensory evoked potentials (SSEPs) and deceleration of CSD propagation in the EXE group. Western blot results in the EXE group showed significant increases in BDNF, GFAP, and NeuN levels and significant decreases in neurodegenerative disease markers. Decreases in SSEP and CSD parameters may result from exercise-induced increases in cerebrovascular system function and increases in the stability and buffering of extracellular ion concentrations and cortical excitability.

Intermittent Lead Exposure Induces Behavioral and Cardiovascular Alterations Associated with Neuroinflammation

The nervous system is the primary target for lead exposure and the developing brain appears to be especially susceptible, namely the hippocampus. The mechanisms of lead neurotoxicity remain unclear, but microgliosis and astrogliosis are potential candidates, leading to an inflammatory cascade and interrupting the pathways involved in hippocampal functions. Moreover, these molecular changes can be impactful as they may contribute to the pathophysiology of behavioral deficits and cardiovascular complications observed in chronic lead exposure. Nevertheless, the health effects and the underlying influence mechanism of intermittent lead exposure in the nervous and cardiovascular systems are still vague. Thus, we used a rat model of intermittent lead exposure to determine the systemic effects of lead and on microglial and astroglial activation in the hippocampal dentate gyrus throughout time. In this study, the intermittent group was exposed to lead from the fetal period until 12 weeks of age, no exposure (tap water) until 20 weeks, and a second exposure from 20 to 28 weeks of age. A control group (without lead exposure) matched in age and sex was used. At 12, 20 and 28 weeks of age, both groups were submitted to a physiological and behavioral evaluation. Behavioral tests were performed for the assessment of anxiety-like behavior and locomotor activity (open-field test), and memory (novel object recognition test). In the physiological evaluation, in an acute experiment, blood pressure, electrocardiogram, and heart and respiratory rates were recorded, and autonomic reflexes were evaluated. The expression of GFAP, Iba-1, NeuN and Synaptophysin in the hippocampal dentate gyrus was assessed. Intermittent lead exposure induced microgliosis and astrogliosis in the hippocampus of rats and changes in behavioral and cardiovascular function. We identified increases in GFAP and Iba1 markers together with presynaptic dysfunction in the hippocampus, concomitant with behavioral changes. This type of exposure produced significant long-term memory dysfunction. Regarding physiological changes, hypertension, tachypnea, baroreceptor reflex impairment and increased chemoreceptor reflex sensitivity were observed. In conclusion, the present study demonstrated the potential of lead intermittent exposure inducing reactive astrogliosis and microgliosis, along with a presynaptic loss that was accompanied by alterations of homeostatic mechanisms. This suggests that chronic neuroinflammation promoted by intermittent lead exposure since fetal period may increase the susceptibility to adverse events in individuals with pre-existing cardiovascular disease and/or in the elderly.

NLRP3 upregulation related to sleep deprivation-induced memory and emotional behavior changes in TRPV1-/- mice

Sleep deprivation, which is a common problem in modern society, impairs memory function and emotional behavior. TRPV1, a subfamily of transient receptor potential cation channels, is abundantly expressed in the central nervous system and is associated with animal behavior. In this article, we report that TRPV1 deficiency in mice alleviates sleep deprivation-induced abnormal behaviors. We found that in the sleep-deprived mice, TRPV1 knockout increased the duration and visits in the central area in the open field task and increased visits to the open arms in the elevated plus maze. The TRPV1-/- mice performed better during the test stage in the Morris water maze phase after sleep deprivation. In the mPFC and hippocampus regions, western blotting results showed that TRPV1-/- attenuated sleep deprivation-induced increases in GFAP, NLRP3, and ASC and increased the expression of the mitochondrial marker Tom20. Immunofluorescence results showed that the action of TRPV1 knockout on NLRP3 was negatively correlated with Tom20 after sleep deprivation. Our results confirm that TRPV1 knockout attenuates sleep deprivation-induced behavioral disorders. The effect of TRPV1 on the behavior of sleep-deprived mice may be related to the neuroinflammation associated with mitochondria in the mPFC and hippocampus.

Longitudinal change in plasma biomarkers by amyloid PET status

AbstractBackgroundIt is important to better understand the longitudinal trajectories of plasma measures of proteins implicated in Alzheimer’s disease among cognitively normal older adults.MethodWe investigated whether baseline levels and longitudinal rates of change of three Quanterix SIMOA plasma measures (amyloid‐beta 42 to 40 ratio [Aβ42/Aβ40], glial fibrillary acidic protein [GFAP], and neurofilament light chain [NfL]) differ by amyloid group as determined using Pittsburgh compound B (PiB) PET in the Baltimore Longitudinal Study of Aging (N=147 PiB– and 57 PiB+; 640 longitudinal measures). All participants were cognitively normal at baseline. We used linear mixed effects models with longitudinal plasma measures as the outcome. We conducted an unadjusted analysis that included binary PiB group, time from baseline, and their interaction. We also conducted an adjusted analysis that additionally included the following explanatory variables: baseline age and its interaction with time, sex, APOE ε4 carrier vs noncarrier, race, and estimated glomerular filtration rate (eGFR) from serum creatinine concurrent with plasma measurements. All variables except time were mean‐centered.ResultParticipant characteristics are presented in Table 1. Figures 1 and 2 illustrate plasma biomarkers versus age and longitudinal visit schedule per participant, respectively. PiB+ individuals had lower Aβ42/Aβ40, higher GFAP, and higher NfL at baseline (Table 2). Both PiB groups exhibited longitudinal increases in GFAP and NfL (Table 3). Aβ42/Aβ40 declined longitudinally in the PiB– group, but annual change in the PiB+ group was not statistically significant. PiB group difference in rates of change in plasma measures was statistically significant for Aβ42/Aβ40 but not for NfL. Difference in rate of change in GFAP was statistically significant in the unadjusted model only. Adjusted model fixed effect estimates are depicted in Figure 3.ConclusionSteeper decline in plasma Aβ42/Aβ40 in the PiB– group suggests that individuals may exhibit decline in this measure prior to the onset of amyloid PET positivity. Despite exhibiting baseline PiB group differences, plasma GFAP and NfL have similar rates of increase in both PiB groups. Reliable detection of such changes at the individual level, prior to the onset of cognitive impairment, may enable better personalized prediction of future outcomes.

Effect of Player Position on Serum Biomarkers during Participation in a Season of Collegiate Football.

This prospective cohort study examined the relationship between a panel of four serum proteomic biomarkers (glial fibrillary acidic protein [GFAP], ubiquitin C-terminal hydrolase-L1 [UCH-L1], total Tau, and neurofilament light chain polypeptide [NF-L]) in 52 players from two different cohorts of male collegiate student football athletes from two different competitive seasons of Division I National Collegiate Athletic Association Football Bowl Subdivision. This study evaluated changes in biomarker concentrations (as indicators of brain injury) over the course of the playing season (pre- and post-season) and also assessed biomarker concentrations by player position using two different published classification systems. Player positions were divided into: 1) speed (quarterbacks, running backs, halfbacks, fullbacks, wide receivers, tight ends, defensive backs, safety, and linebackers) versus non-speed (offensive and defensive linemen), and 2) "Profile 1" (low frequency/high strain magnitudes positions including quarterbacks, wide receivers, and defensive backs), "Profile 2" (mid-range impact frequency and strain positions including linebackers, running backs, and tight ends), and "Profile 3" (high frequency/low strains positions including defensive and offensive linemen). There were significant increases in GFAP 39.3 to 45.6 pg/mL and NF-L 3.5 to 5.4 pg/mL over the course of the season (p < 0.001) despite only five players being diagnosed with concussion. UCH-L1 decreased significantly, and Tau was not significantly different. In both the pre- and post-season blood samples Tau and NF-L concentrations were significantly higher in speed versus non-speed positions. Concentrations of GFAP, Tau, and NF-L increased incrementally from "Profile 3," to "Profile 2" to "Profile 1" in the post-season. UCH-L1 did not. GFAP increased (by Profiles 3, 2, 1) from 42.4 to 49.6 to 78.2, respectively (p = 0.051). Tau increased from 0.37 to 0.61 to 0.67, respectively (p = 0.024). NF-L increased from 3.5 to 4.9 to 8.2, respectively (p < 0.001). Although GFAP and Tau showed similar patterns of elevations by profile in the pre-season samples they were not statistically significant. Only NF-L showed significant differences between profiles 2.7 to 3.1 to 4.2 in the pre-season (p = 0.042). GFAP, Tau, and NF-L concentrations were significantly associated with different playing positions with the highest concentrations in speed and "Profile 1" positions and the lowest concentrations were in non-speed and "Profile 3" positions. Blood-based biomarkers (GFAP, Tau, NF-L) provide an additional layer of injury quantification that could contribute to a better understanding of the risks of playing different positions.

Effect of sex and gonadectomy on brain MPTP toxicity and response to dutasteride treatment in mice

The main neuropathological feature of Parkinson's disease (PD) is degeneration of dopamine (DA) neurons in the substantia nigra (SN); PD prevalence is higher in men, suggesting a role of sex hormones in neuroprotection. This study sought the effects of sex hormones in the brain in a mouse model of PD and modulation of steroid metabolism/synthesis with the 5α-reductase inhibitor dutasteride shown to protect 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) male mice. Male and female mice were gonadectomized (GDX) or SHAM operated. They were treated with vehicle or dutasteride (5 mg/kg) for 10 days and administered a low dose of MPTP (5.5 mg/kg) or saline on the 5th day to model early PD; brains were collected thereafter. Striatal measures of the active metabolite 1-methyl-4-phenylpyridinium (MPP+) contents showed no difference supporting an effect of the experimental conditions investigated. In SHAM MPTP male mice loss of striatal DA and metabolites, DA transporter (DAT) and vesicular monoamine transporter 2 (VMAT2) specific binding in the striatum and SN was prevented by dutasteride treatment; these changes were inversely correlated with glial fibrillary acidic protein (GFAP, an astrogliosis marker) levels. In SHAM female mice MPTP treatment had little or no effect on striatal and SN DA markers and GFAP levels whereas GDX male and female mice showed a similar loss of striatal DA markers and increase of GFAP. No effect of dutasteride treatment was observed in GDX male and female mice. In conclusion, sex differences in mice MPTP toxicity and response to dutasteride were observed that were lost upon gonadectomy implicating neuroinflammation.

Date seeds alleviate behavioural and neuronal complications of metabolic syndrome in rats

Unhealthy dietary habits can play a crucial role in metabolic damages, promoting alteration of neural functions through the lifespan. Recently, dietary change has been perceived as the first line intervention in prevention and/or treatment of metabolic damages and related diseases. In this context, our study was designed to assess the eventual therapeutic effect of date seeds administration on memory and learning and on neuronal markers in a rat Metabolic Syndrome model. For this purpose, 32 adult male Wistar rats were fed with standard diet or high-fat high-sugar diet during ten weeks. After this, 16 rats were sacrified and the remaining rats received an oral administration of 300 mg of date seeds/kg of body weight during four supplementary weeks. Before sacrifice, we evaluate cognitive performances by the Barnes maze test. Afterwards, neuronal, astrocytic, microtubular and oxidative markers were investigated by immunoblotting methods. In Metabolic syndrome rats, results showed impairment of spatial memory and histological alterations. We identified neuronal damages in hippocampus, marked by a decrease of NeuN and an increase of GFAP and pTau396. Finally, we recorded an increase in protein oxidation and lipid peroxidation, respectively identified by an up-regulation of protein carbonyls and 4-HNe. Interestingly, date seeds administration improved these behavioural, histological, neuronal and oxidative damages highlighting the neuroprotective effect of this natural compound. Liquid Chromatography-Mass Spectrometry (LC-MS) identified, in date seeds, protocatechuic acid, caffeoylshikimic acid and vanillic acid, that could potentially prevent the progression of neurodegenerative diseases, acting through their antioxidant properties.

Oligodendroglial alterations in FTD caused by C9orf72 expansion

AbstractBackgroundThe most frequent genetic cause of Frontotemporal Dementia (FTD) and Amyotrophic Lateral Sclerosis (ALS) is the hexanucleotide repeat expansion in the C9orf72 gene. Until now, three different mechanisms have been proposed to explain its pathogenic effects: i) loss of function of the C9orf72 protein, ii) toxic gain of function from sense and antisense C9orf72 repeat RNA, or iii) toxic gain of function from Dipeptide Repeat Proteins. Another important hallmark associated with C9orf72 mutations is the TDP43 accumulation in neurons and glia. As in other neurodegenerative diseases, neuroinflammation is a common feature in FTD; however, the specific role of the innate immune system in FTD associated to specific subtypes of the disease is unknown.MethodIn this study, we investigated changes in the pattern of different glial markers (GFAP, YKL‐40, Iba‐1 and MBP) using immunohistochemistry in the frontal cortex and hippocampus of a cohort of 23 cases diagnosed with C9‐FTD/ALS with TDP43 pathology (C9‐FTD), 8 cases diagnosed with sporadic‐FTD TDP43 (sFTD) and 9 healthy controls (HC). Different in‐house semi‐automated MATLAB algorithms were developed and were used to quantify the immunoreactivity of these different markers.ResultWe found a significant increase of GFAP, YKL‐40 and Iba1 immunoreactivity in C9‐FTD and in the sporadic group compared with a group of healthy controls both in frontal cortex and hippocampus. We also found a decreased of MBP protein immunoreactivity in the group of C9‐FTD samples, both in grey and white matter from the frontal cortex, compared to control and sporadic group of samples. In C9‐FTD group we found a negative correlation between MBP and TDP43.ConclusionThese results indicate that FTD caused by C9orf72 is associated with myelin loss likely caused by the mutation or its distinct pathology. Understanding these biological processes will help to identify specific pathways associated with this FTLD subtype and help in the development of new therapeutic strategies implicating oligodendroglial cells as a target to reduce the disease progression and improve quality of life of FTD with C9orf72 expansion patients.

Connexin43 Expression and Associated Chronic Inflammation Presages the Development of Cerebral Radiation Necrosis.

Cerebral radiation necrosis (CRN) is a delayed complication of radiosurgery that can result in severe neurological deficits. The biological changes leading to necrotic damage may identify therapeutic targets for this complication. Connexin43 expression associated with chronic inflammation may presage the development of CRN. A mouse model of delayed CRN was used. The left hemispheres of adult female mice were irradiated with single-fraction, high-dose radiation using a Leksell Gamma Knife. The brains were collected 1 and 4 days, and 1-3 weeks after the radiation. The expression of connexin43, interleukin-1β (IL-1β), GFAP, isolectin B-4, and fibrinogen was evaluated using immunohistochemical staining and image analysis. Compared with the baseline, the area of connexin43 and IL-1β staining was increased in ipsilateral hemispheres 4 days after radiation. Over the following 3 weeks, the density of connexin43 gradually increased in parallel with progressive increases in GFAP, isolectin B-4, and fibrinogen labeling. The overexpression of connexin43 in parallel with IL-1β spread into the affected brain regions first. Further intensified upregulation of connexin43 was associated with escalated astrocytosis, microgliosis, and blood-brain barrier breach. Connexin43-mediated inflammation may underlie radiation necrosis and further investigation of connexin43 hemichannel blockage is merited for the treatment of CRN.

Neuroprotective and anticonvulsant effects of sinomenine in kainate rat model of temporal lobe epilepsy: Involvement of oxidative stress, inflammation and pyroptosis

Oxidative stress, inflammation and pyroptosis are three of the most important mechanisms in the pathophysiology of temporal lobe epilepsy (TLE). Most people with TLE are refractory to the existing drugs. Sinomenine has shown neuroprotective effects through counteracting oxidative stress, inflammation and pyroptosis. In this study, we evaluated the effect of sinomenine on seizure behavior, oxidative stress, inflammation and pyroptosis markers in addition to its neuroprotective potential in intrahippocampal kainate-induced rat model of TLE. For this purpose, male rats (n = 60) were randomly divided into five groups, i.e., sham, kainate (lesion) with an intrahippocampal injection of kainate, kainate groups receiving sinomenine at doses of 30 or 50 mg/kg, and kainate group receiving valproic acid at a dose of 200 mg/kg (as the positive control). Our obtained data showed that sinomenine administration at a dose of 50 mg/kg can significantly decreases severity of seizures and incidence of status epilepticus (SE), hippocampal aberrant MFS and DNA fragmentation and prevents reduction of neuronal density. It also significantly restored level of ROS, MDA, HO-1 and SOD but its effect on GSH level was not significant. Additionally, sinomenine at a dose of 50 mg/kg partially counteracted the increase of NF-κB, TLR 4, TNFα, GFAP and caspase 1. These results suggest that sinomenine has anticonvulsant and neuroprotective effects by reducing hippocampal oxidative stress, inflammation, pyroptosis and apoptosis in intrahippocampal kainate model of TLE.

IRAK4 Inhibition Attenuates Interleukin‐1‐Induced Astrogliosis: A Potential Therapeutic Target for Neurodegenerative Disorders?

The activation of inflammatory signaling in the central nervous system (CNS) is a common component of many neurodegenerative conditions including traumatic brain injury (TBI), a disease for which there are currently no FDA‐approved pharmacotherapies. Inflammation orchestrates alterations in the astrocytic transcriptome, driving the process of astrogliosis and formation of A1‐type neurodegenerative astrocytes; an effect stemming in part from interleukin‐1 (IL‐1)‐mediated signaling. We hypothesize that IL‐1‐mediated activation of functional, astrocytic type 1 interleukin‐1 receptors (IL‐1R1) promotes reactive astrogliosis and neurotoxic A1‐type astrocyte formation following TBI. Further, we hypothesize that pharmacologically targeting astrocyte specific IL‐1R1‐mediated signaling may be effective in mitigating reactive astrogliosis and neurodegeneration inherent to TBI. In vivo, blast‐induced TBI rapidly increased cortical mRNA expression of interleukin‐1α (il‐1α), interleukin‐1β (il‐1β), il‐1r1 and glial fibrillary acidic protein (gfap). In vitro studies using murine astrocytes (ATCC, C8‐D1A) were used to characterize astrocyte‐specific responsiveness to direct, recombinant IL‐1α and IL‐1β exposure. IL‐1α and IL‐1β treatment increased gfap expression, indicative of significant astrocyte activation. Increased expression of il‐1r1 was also detected following IL‐1α or IL‐1β treatment of murine astrocytes in vitro, revealing an astrocyte‐specific positive feedback mechanism. IL‐1α treatment increased the expression of A1‐type astrocyte marker serping1, whereas IL‐1β treatment was found to dose‐dependently reduce serping1 expression. Interleukin‐1 receptor associated kinase‐4 (IRAK4), a kinase downstream of IL‐1R1 activation, is amenable to targeting with small molecule inhibitors. We hypothesized that IRAK4 may be a viable target for blocking IL‐1R1‐mediated signaling in astrocytes. Treatment of astrocytes with the selective IRAK4 inhibitor, AS2444697, attenuated IL‐1α‐induced increases in gfap, il‐1r1 and serping1 expression, indicative of a blockade of both reactive astrogliosis and the formation of neurotoxic astrocytes. Studies contained here provide the first evidence that the inhibition of IRAK4 may represent a potential strategy to inhibit the formation of neurotoxic A1‐type astrocytes in various neurodegenerative disorders, including TBI. Future studies will utilize in vivo murine models affording the conditional elimination and restoration of IL‐1R1 in astrocytes to better understand the role of IL‐1R1 signaling in TBI. Systematic characterization of the role of IRAK4 in the formation of A1‐type astrocytes will lead to a better understanding of how IL‐1R1 signaling contributes to the acute and chronic neurologic sequelae of TBI. Combined, these studies identify a potentially viable pharmacotherapeutic target required for IL‐1R1‐initiated signaling within astrocyte populations.Support or Funding InformationStudies were supported by the Brain &amp; Behavior Research Foundation, PhRMA Foundation, and the University of Cincinnati.

CHOLESTATIC LIVER DISEASE AND BRAIN DYSFUNCTION: ROLE OF THE ARYL HYDROCARBON RECEPTOR

BackgroundCholestatic liver disease is associated with immune‐mediated inflammatory liver injury and tissue damage. This disorder is also associated with brain dysfunction and behavioural changes, notably fatigue, depression and social withdrawal. The mechanisms leading to these central nervous system abnormalities are unknown, however, they are associated with neuroinflammation in the brain. Microglia and astrocytes are two glial cell populations that play key roles in neuroinflammation. Activated glia display morphological changes, secrete cytokines, and mediate electrophysiological changes, altering the normal functioning of the brain. The aryl hydrocarbon receptor (AhR) is a transcription factor involved in the immune response. AhR is present on glia and its’ activation has been shown to reduce CNS inflammation. The role of the AhR in cholestatic liver disease has yet to examined.AimsTo study the function of the AhR and its ligands in an extensively characterized model of cholestic liver disease. We will test the hypothesis that activation of AhR in the brain will reduce neuroinflammation and associated behavioral deficits observed in cholestatic mice.MethodsMale C57Bl/6J mice had cholestasis induced by bile duct ligation (BDL); comparisons were made to sham‐operated controls. Mice were tested for social interaction with a 4‐week old juvenile male in their home cage and the number of social interaction attempts quantified. After this behavioral assay, mice were euthanized, brains were removed and processed for immunofluorescence analysis. Brain sections were stained for markers of microglia (IBA‐1) and astrocytes (GFAP). Microglia were counted and astrocyte activation was qualitatively assessed as higher GFAP immunofluorescence. PCR was used to quantify relative gene expression of AhR and its downstream gene targets (eg. CYP1A1) in mice that recived treatment with beta‐napthoflavone (BNF), a potent AhR agonist, or in vehicle treated controls.ResultsBDL mice made significantly fewer attempts to interact with the juvenile as compared to controls (P&lt;0.05). We also observed a significant increase in IBA‐1 immunoreactive cell numbers in both the CA1 region of the hippocampus and the hypothalamic paraventricular nucleus (PVN, P&lt;0.05). BDL mice also displayed marked increases in GFAP+ staining in the PVN, but not the CA1 region of hippocampus, in contrast to sham controls. Lastly, we found that BNF significantly upregulated CYP1A1 (P&lt;0.05) in the liver and brain prefrontal cortex of mice. We are currently examining whether BNF can downregulate neuroinflammation and improve decreased social interaction behaviors in cholestatic mice.ConclusionCholestatic liver damage was associated with impaired social behavior. Further, glial activation, an indicator of neuroinflammation was significantly increased in components of the brain limbic system associated with the response to stress, learning, and memory. Future experiments will address whether activation of the AhR will ameliorate neuroinflammation and behavioral changes observed in mice with cholestatic liver injury.Support or Funding InformationCanadian Institutes of Health Research

A187 CHOLESTATIC LIVER DISEASE AND BRAIN DYSFUNCTION: ROLE OF THE ARYL HYDROCARBON RECEPTOR

Abstract Background Cholestatic liver disease is associated with immune-mediated inflammatory liver injury. This disorder is also associated with brain dysfunction and behavioural changes, notably fatigue, depression and social withdrawal. The mechanisms leading to these central nervous system abnormalities are unknown, however, they are associated with neuroinflammation. Microglia and astrocytes are two glial populations that play key roles in neuroinflammation. Activated glia display morphological changes, secrete cytokines, and mediate electrophysiological changes, altering the normal functioning of the brain. The aryl hydrocarbon receptor (AhR) is a transcription factor involved in the immune response. AhR is present on glia and its’ activation has been shown to reduce neuroinflammation. The role of the AhR in cholestatic liver disease has yet to be examined. Aims To study the function of the AhR in a model of cholestic liver disease. We will test the hypothesis that activation of AhR in the brain will reduce neuroinflammation and behavioral deficits observed in cholestatic mice. Methods Male C57Bl/6J mice had cholestasis induced by bile duct ligation (BDL); comparisons were made to sham-operated controls. Mice were tested for social interaction with a 4-week old juvenile in their home cage and the number of social interaction attempts quantified. Next, mice were euthanized, brains were removed and processed for immunohistochemistry. Brain sections were stained for markers of microglia (IBA-1) and astrocytes (GFAP). Microglia were counted and astrocyte activation was qualitatively assessed. PCR was used to quantify gene expression of AhR and its downstream gene targets (eg. CYP1A1) in mice that recived treatment with beta-napthoflavone (BNF), an AhR agonist, or in vehicle treated controls. Results BDL mice made significantly fewer attempts to interact with the juvenile as compared to controls (P&amp;lt;0.05). We also observed a significant increase in IBA-1 immunoreactive cell numbers in both the CA1 region of the hippocampus and the hypothalamic paraventricular nucleus (PVN, P&amp;lt;0.05). BDL mice also displayed marked increases in GFAP+ staining in the PVN, but not the CA1, in contrast to sham controls. Lastly, we found that BNF significantly upregulated CYP1A1 (P&amp;lt;0.05) in the liver and prefrontal cortex of mice. We are currently examining whether BNF can reduce neuroinflammation and improve decreased social interaction in cholestatic mice. Conclusions Cholestatic liver damage was associated with impaired social behavior. Further, glial activation, an indicator of neuroinflammation was increased in components of the limbic system associated with the response to stress, learning, and memory. Future experiments will address whether activation of the AhR will ameliorate neuroinflammation and behavioral changes observed in mice with cholestatic liver injury. Funding Agencies CCC, CIHR