Iba1 Expression Research Articles

Electroacupuncture alleviates the symptom of depression in mice by regulating the cGAS-STING-NLRP3 signaling.

To investigate the therapeutic effect of electroacupuncture (EA) on chronic and unpredictable mild stress (CUMS)-induced depression in mice and the underlying mechanism. Male C57BL/6 mice were randomly divided into 6 groups: Control, CUMS, CUMS+EA-placebo, CUMS+EA, CUMS+ ad-NC, CUMS+ ad-cGAS-shRNA. CUMS was utilized to establish the depression model in mice. The behavioral changes were determined by the forced swimming, open field, and sucrose preference experiments. The pathological changes in the hippocampus tissue were evaluated by HE staining. The release of TNF-α, IL-1β, IL-6, 5-HT, and NE in the hippocampus tissue was determined by ELISA. IBA-1 expression detected by the immunofluorescence was used to represent the activity of microglia. Western blot and RT-PCR were utilized to measure the expression of Bax, bcl-2, cGAS, STING, TBK1, IRF3, and NLRP3. The depression behavior in CUMS mice was significantly alleviated by the treatment of EA and cGAS-shRNA, accompanied by ameliorated hippocampus pathological changes, declined production of TNF-α, IL-1β, and IL-6, elevated secretion of 5-HT and NE, and inhibition on the activity of microglia. Furthermore, significantly elevated expression level of Bax, cGAS, STING, TBK1, IRF3, and NLRP3 and declined expression level of bcl-2 were observed in the CUMS+EA and CUMS+ ad-cGAS-shRNA groups. EA significantly mitigated the symptom of depression in mice, which was closely associated with the repressed neuroinflammation, increased monoamine concentration, inactivated microglia, and inhibited cGAS-STING-NLRP3 signaling.

Vincamine alleviates brain injury by attenuating neuroinflammation and oxidative damage in a mouse model of Parkinson's disease through the NF-κB and Nrf2/HO-1 signaling pathways.

Parkinson's disease (PD)is a neurodegenerative disease featured by progressive loss of nigrostriatal dopaminergic neurons, the etiology of which is associated with the existence of neuroinflammatory response and oxidative stress. Vincamine is an indole alkaloid that was reported to exhibit potent anti-inflammatory and antioxidant properties in many central and/or peripheral diseases. Nevertheless, the specific role of vincamine in PD development remains unknown. In our study, dopaminergic neuron loss was determined through immunohistochemistry staining and western blot analysis of tyrosine hydroxylase (TH)expression in the substantia nigra (SN)of PD mice. Reactive oxygen species (ROS)production and malondialdehyde (MDA),superoxide dismutase (SOD),and glutathione (GSH)levels were detected through DHE staining and commercially available kits to assess oxidative stress. Pro-inflammatorycytokine (TNF-α,IL-1β, and IL-6)levels in the SN were measured via RT-qPCRand western blot analysis. Microglial and astrocyte activation was examined through immunofluorescence staining of Iba-1 (microglia marker) and GFAP (astrocyte marker) in the SN. The regulation of vincamine on the NF-κBand Nrf2/HO-1pathway was estimated through western blot analysis. Our results showed that vincamine treatment decreased TNF-α,IL-1β,and IL-6 mRNA and protein levels, reduced GFAP and Iba-1 expression, decreased ROS production and MDA level, and increased SOD activity and GSH level in the SN of PD mice. Mechanically, vincamine repressed the phosphorylation levels of p65, IKKβ, and IκBα but enhanced the protein levels of Nrf2 and HO-1 in PD mice. Collectively, vincamine plays a neuroprotective role in PD mouse models by alleviating neuroinflammation and oxidative damage via suppressing the NF-κBpathway and activating the Nrf2/HO-1pathway.

Nerolidol rescues hippocampal injury of diabetic rats through inhibiting NLRP3 inflammasome and regulation of MAPK/AKT pathway.

Despite the observation of diabetes-induced brain tissue damage and impaired learning and memory, the underlying mechanism of damage remains elusive, and effective, targeted therapeutics are lacking. Notably, the NLRP3 inflammasome is highly expressed in the hippocampus of diabetic individuals. Nerolidol, a naturally occurring compound with anti-inflammatory and antioxidant properties, has been identified as a potential therapeutic option for metabolic disorders. However, the ameliorative capacity of nerolidol on diabetic hippocampal injury and its underlying mechanism remain unclear. Network pharmacology and molecular docking was used to predict the signaling pathways and therapeutic targets of nerolidol for the treatment of diabetes. Then established a diabetic rat model using streptozotocin (STZ) combined with a high-fat diet and nerolidol was administered. Morris water maze to assess spatial learning memory capacity. Hematoxylin and eosin and Nissl staining was used to detect neuronal damage in the diabetic hippocampus. Transmission electron microscopy was used to detect the extent of damage to mitochondria, endoplasmic reticulum (ER) and synapses. Immunofluorescence was used to detect GFAP, IBA1, and NLRP3 expression in the hippocampus. Western blot was used to detect apoptosis (Bcl-2, BAX, and Cleaved-Caspase-3); synapses (postsynaptic densifying protein 95, SYN1, and Synaptophysin); mitochondria (DRP1, OPA1, MFN1, and MFN2); ER (GRP78, ATF6, CHOP, and caspase-12); NLRP3 inflammasome (NLRP3, ASC, and caspase-1); inflammatory cytokines (IL-18, IL-1β, and TNF-α); AKT (P-AKT); and mitogen-activated protein kinase (MAPK) pathway (P-ERK, P-p38, and P-JNK) related protein expression. Network pharmacology showed that nerolidol's possible mechanisms for treating diabetes are the MAPK/AKT pathway and anti-inflammatory effects. Animal experiments demonstrated that nerolidol could improve blood glucose, blood lipids, and hippocampal neuronal damage in diabetic rats. Furthermore, nerolidol could improve synaptic, mitochondrial, and ER damage in the hippocampal ultrastructure of diabetic rats by potentially affecting synaptic, mitochondrial, and ER-related proteins. Further studies revealed that nerolidol decreased neuroinflammation, NLRP3 and inflammatory factor expression in hippocampal tissue while also decreasing MAPK pathway expression and enhancing AKT pathway expression. However, nerolidol improves hippocampal damage in diabetic rats cannot be shown to improve cognitive function. In conclusion, our study reveals for the first time that nerolidol can ameliorate hippocampal damage, neuroinflammation, synaptic, ER, and mitochondrial damage in diabetic rats. Furthermore, we suggest that nerolidol may inhibit NLRP3 inflammasome and affected the expression of MAPK and AKT. These findings provide a new experimental basis for the use of nerolidol to ameliorate diabetes-induced brain tissue damage and the associated disease.

Wogonoside alleviates microglia-mediated neuroinflammation via TLR4/MyD88/NF-κB signaling axis after spinal cord injury

Wogonoside (WG) is a natural flavonoid extracted from Scutellariae Radix, recognized for its established anti-inflammatory properties. However, the role of WG in the context of neuroinflammation after spinal cord injury (SCI) remains inadequately elucidated. This study employed in silico, in vitro, and in vivo methodologies to investigate the impact of WG on microglia-mediated neuroinflammation after SCI. In the in silico experiment, we identified 15 potential target genes of WG associated with SCI. These genes were linked to the regulation of inflammatory response and immune defense. Molecular docking maps revealed toll-like receptor 4 as a molecular target for WG, demonstrating binding through a hydrogen bond (Lys263, Ser120). In lipopolysaccharide-stimulated BV2 cells and SCI mice, WG significantly attenuated microglial activation and facilitated a phenotype shift from M1 to M2. This was evidenced by the reversal of the increased expressions of Iba1, GFAP, and iNOS, as well as the decreased expression of Arg1. WG also suppressed the production of pro-inflammatory mediators (NO, TNF-α, IL-6, IL-1α, IL-1β, C1q). WG exerted these effects by suppressing the TLR4/MyD88/NF-κB signaling axis in microglia. Furthermore, by reducing levels of TNF-α, IL-1α, and C1q in supernatant of LPS-induced microglia, WG indirectly induced astrocytes change to A2 phenotype, evidenced by transcriptome sequencing result of primary mouse astrocytes. All these events above collectively created a favorable microenvironment, contributing to a significant alleviation of weight loss and neuronal damage at the lesion site of SCI mice. Our findings substantiate the efficacy of WG in mitigating neuroinflammation after SCI, thereby warranting further exploration.

Syringaresinol attenuates Tau phosphorylation and ameliorates cognitive dysfunction induced by sevoflurane in aged rats.

Cognitive dysfunction following anesthesia with agents such as sevoflurane is a significant clinical problem, particularly in elderly patients. This study aimed to explore the protective effects of the phytochemical syringaresinol (SYR) against sevoflurane-induced cognitive deficits in aged Sprague-Dawley rats and to determine the underlying mechanisms involved. We assessed the impact of SYR on sevoflurane-induced cognitive impairment, glial activation, and neuronal apoptosis through behavioral tests (Morris water maze), immunofluorescence, Western blotting for key proteins involved in apoptosis and inflammation, and enzyme-linked immunosorbent assays for interleukin-1β, tumor necrosis factor-α, and interleukin-6. SYR treatment mitigated sevoflurane-induced cognitive decline, reduced microglial and astrocyte activation (decreased Iba-1 and GFAP expression), and countered neuronal apoptosis (reduced Bax, cleaved-caspase3, and cleaved-PARP expression). SYR also enhanced Sirtuin-1 (SIRT1) expression and reduced p-Tau phosphorylation; these effects were reversed by the SIRT1 inhibitor EX527. SYR exerts neuroprotective effects on sevoflurane-induced cognitive dysfunction by modulating glial activity, apoptotic signaling, and Tau phosphorylation through the SIRT1 pathway. These findings could inform clinical strategies to safeguard cognitive function in patients undergoing anesthesia.

Congenital localized cutaneous Langerhans cell histiocytosis in a Holstein calf.

Distinct solitary dermal nodules, either covered by an alopecic, or sometimes ulcerated, epidermis, were noticed on the head of a stillborn Holstein calf. The head was submitted for autopsy, and the nodules were found to consist of homogeneous, diffuse pale-yellow, soft-tissue masses with distinct margins that elevated the epidermis above the adjacent skin. Histologically, the dermal nodules were well-delineated on the deep margin approaching the cutaneous muscle and consisted of perivascular neoplastic infiltrates of round cells that in some places coalesced into sheets that extended into the dermis and subcutis. Neoplastic cells separated adnexa and collagen. Immunohistochemistry revealed intense tumor cell expression of vimentin, Iba1, E-cadherin, and CD204; expression of CD18 was faint. The masses were diagnosed as Langerhans cell histiocytosis. Congenital cutaneous Langerhans cell histiocytosis has not been reported previously in cattle, to our knowledge, and should be included in the differential diagnosis of congenital nodular skin lesions.

Central effects of peripherally administrated immune cells modulated by a psychoactive substance in aggression

IntroductionIt is known that the formation of aggressive behavior is accompanied by neurodegenerative and neuroinflammatory changes. Immune cells have a regulatory effect on the central nervous system functions, including regulation of behavior.ObjectivesWe first demonstrated that ex vivo chlorpromazine - modulated immune cells have a positive aggressive behavior editing effect. The aim of the study was to investigate the influence of the indicated cells on some central mechanisms underlying the development of aggressive reactions.Methods(CBAxC57Bl/6) F1 aggressive male mice, developed in conditions of chronic social stress, were undergoing the transplantation of syngeneic spleen lymphocytes with ex vivo chlorpromazine-modulated functional activity. In recipients the immunohistochemical analysis was performed assessing the expression of the microglial marker Iba1. The levels of brain-derived neurotrophic factor (Bdnf ) and cytokines was assessed by ELISA. For histological examination Nissl staining was applied.ResultsAggressive behavior editing after the chlorpromazine-modulated immune cells transplantation registered against the background of some structural and functional changes in the brain. It was found an increase in the density of pyramidal neurons in CA1 and CA3 hippocampal regions and augmented level of Bdnf. The decreased expression of microglial activation marker Iba1, accompanied with decreased levels of pro-inflammatory cytokines (IL-1β, IL-2, IL-6, INF-γ) and increased anti-inflammatory (IL-4) cytokine was found. Visualization of functionally active lymphocytes pre-treated with chlorpromazine in the brain parenchyma of aggressive recipients suggests a direct effect of injected lymphocytes on CNS.ConclusionsThe effect of chlorpromazine - modulated immune cells that edits aggressive behavior is realized by stimulating neurogenesis, neuroplasticity and reducing neuroinflammation.Disclosure of InterestNone Declared

352 Microglial Behavior and Iba-1 Expression: Evaluating the Cognitive Impact of Vascular Dementia and Long COVID

OBJECTIVES/GOALS: The study aims to explore the role of microglial behavior in cognitive impairment associated with vascular dementia (VaD) and long COVID. Using immunohistochemistry (IHC) and quantitative PCR (qPCR), we will assess the expression of Iba-1, a microglial activation marker, in subjects with VaD and SARS-CoV-2 infection. METHODS/STUDY POPULATION: Out of 48 female C57BL/6 mice, 24 had surgical intervention in the form of bilateral carotid artery stenosis (BCAS) for experimental induction of vascular dementia. After 2 weeks, 12 BCAS and 12 non-BCAS were infected with 1E4 PFU of mouse-adapted 10 (MA10) strain of SARS-CoV-2. 2 weeks post-infection, 4 weeks post-operatively, all animals were euthanized and tissues were processed for cDNA and histology. Immunofluorescence and RT-qPCR used to quantify microglia via Iba-1, BBB integrity via claudin-5 as well as occludin, GFAP, and integrin a5. RESULTS/ANTICIPATED RESULTS: We anticipate observing distinct patterns of microglial behavior in subjects with vascular dementia (VaD) and those with long COVID. Through immunohistochemistry (IHC), we expect to see increased Iba-1 expression, indicative of microglial activation. Quantitative PCR (qPCR) will likely corroborate these findings, showing elevated levels of Iba-1 mRNA. Lastly, we anticipate that the data will reveal interactions between microglia and the blood-brain barrier (BBB). These interactions could provide insights into how microglial behavior influences BBB integrity and, consequently, cognitive function in VaD and long COVID. DISCUSSION/SIGNIFICANCE: This study aims to clarify the role of microglia in cognitive decline linked to vascular dementia and long COVID. By categorizing patients based on microglial activation, we can better tailor treatments. The findings could lead to targeted therapies that address cognitive impairment in these conditions.

IL-6 mediates olfactory dysfunction in a mouse model of allergic rhinitis

BackgroundImmune-inflammatory response is a key element in the occurrence and development of olfactory dysfunction (OD) in patients with allergic rhinitis (AR). As one of the core factors in immune-inflammatory responses, interleukin (IL)-6 is closely related to the pathogenesis of allergic diseases. It may also play an important role in OD induced by diseases, such as Sjögren’s syndrome and coronavirus disease 2019. However, there is no study has reported its role in OD in AR. Thus, this study aimed to investigate the role of IL-6 in AR-related OD, in an attempt to discover a new target for the prevention and treatment of OD in patients with AR. MethodsDifferential expression analysis was performed using the public datasets GSE52804 and GSE140454 for AR, and differentially expressed genes (DEGs) were obtained by obtaining the intersection points between these two datasets. IL-6, a common differential factor, was obtained by intersecting the DEGs with the General Olfactory Sensitivity Database (GOSdb) again. A model of AR mice with OD was developed by sensitizing with ovalbumin (OVA) to verify the reliability of IL-6 as a key factor of OD in AR and explore the potential mechanisms. Furthermore, a supernatant and microglia co-culture model of nasal mucosa epithelial cells stimulated by the allergen house dust mite extract Derp1 was established to identify the cellular and molecular mechanisms of IL-6-mediated OD in AR. ResultsThe level of IL-6 in the nasal mucosa and olfactory bulb of AR mice with OD significantly increased and showed a positive correlation with the expression of olfactory bulb microglia marker Iba-1 and the severity of OD. In-vitro experiments showed that the level of IL-6 significantly increased in the supernatant after the nasal mucosa epithelial cells were stimulated by Derp1, along with significantly decreased barrier function of the nasal mucosa. The expression levels of neuroinflammatory markers IL-1β and INOS increased after a conditioned culture of microglia with the supernatant including IL-6. Then knockdown (KD) of IL-6R by small interfering RNA (siRNA), the expression of IL-1β and INOS significantly diminished. ConclusionIL-6 plays a key role in the occurrence and development of OD in AR, which may be related to its effect on olfactory bulb microglia-mediated neuroinflammation.

Methamphetamine Enhancement of HIV-1 gp120-Mediated NLRP3 Inflammasome Activation and Resultant Proinflammatory Responses in Rat Microglial Cultures.

Human Immunodeficiency Virus type 1 (HIV-1)-associated neurocognitive disorders (HANDs) remain prevalent in HIV-1-infected individuals despite the evident success of combined antiretroviral therapy (cART). The mechanisms underlying HAND prevalence in the cART era remain perplexing. Ample evidence indicates that HIV-1 envelope glycoprotein protein 120 (gp120), a potent neurotoxin, plays a pivotal role in HAND pathogenesis. Methamphetamine (Meth) abuse exacerbates HANDs, but how this occurs is not fully understood. We hypothesize that Meth exacerbates HANDs by enhancing gp120-mediated neuroinflammation. To test this hypothesis, we studied the effect of Meth on gp120-induced microglial activation and the resultant production of proinflammatory cytokines in primary rat microglial cultures. Our results show that Meth enhanced gp120-induced microglial activation, as revealed by immunostaining and Iba-1 expression, and potentiated gp120-mediated NLRP3 expression and IL-1β processing and release, as assayed by immunoblotting and ELISA. Meth also augmented the co-localization of NLRP3 and caspase-1, increased the numbers of NLRP3 puncta and ROS production, increased the levels of iNOS expression and NO production, and increased the levels of cleaved gasderminD (GSDMD-N; an executor of pyroptosis) in gp120-primed microglia. The Meth-associated effects were attenuated or blocked by MCC950, an NLRP3 inhibitor, or Mito-TEMPO, a mitochondrial superoxide scavenger. These results suggest that Meth enhances gp120-associated microglial NLRP3 activation and the resultant proinflammatory responses via mitochondria-dependent signaling.

Abstract 5522: Elucidating the role of P-selectin/P-selectin ligand-1 axis in lung cancer brain metastasis

Abstract Lung cancer (LC) stands as a prominent cause of cancer-related mortality. Brain metastasis (BM) is a common occurrence in LC, constituting nearly 50% of all BM cases with the brain emerging as the preferred site for LC metastasis. Unfortunately, current LCBM treatment options yield limited success, as reflected in the median survival of BM patients at 7.8 months. Consequently, there is an imperative need for novel and more efficacious therapeutic approaches targeting LCBM. Evidence suggests that brain microenvironment cells, particularly microglia and tumor-associated macrophages (TAMs), play a pivotal role in facilitating the colonization and growth of LCBM. Previous investigations from our lab have identified the involvement of the P-selectin/P-selectin glycoprotein ligand-1 (SELP/PSGL-1) axis in microglia/TAMs immunophenotypes, promoting pro-tumorigenic activity. This led us to explore the role of the SELP/PSGL-1 axis in the interactions between LCBM cells and the brain microenvironment1. Thus, we conducted Immunostaining on human tissue samples, revealing increased expression of SELP, PSGL-1, PD-1, PD-L1, IBA-1, and CD68 in LCBM tissues. Simultaneously, we employed advanced 3D models to investigate the intricate interactions between cancer cells and microglia/TAMs. Actively inhibiting the SELP/PSGL-1 axis with pharmacological agents led to a significant decrease in migration and invasion when LC cells were co-cultured with microglia and TAMs. Moreover, SELP inhibition prompted a shift in microglia/TAMs state towards an M1-like phenotype, enhancing the ability of CD4/CD8 cells to target cancer cells in our unique 3D LCBM models. This effect was further enhanced when co-treated with a PD-1 inhibitor, demonstrating a potential synergistic impact in targeting both pathways for therapeutic intervention. Our results suggest that combining SELP inhibition with other immunomodulators, including a small molecule PD-L1 inhibitor developed in our lab2, could synergistically enhance both innate and adaptive anti-tumor immune responses. Taken together, this study may enhance our understanding of LCBM immunogenicity and contribute to the development of novel immunotherapeutic approaches that can improve patient outcomes.1.Yeini, E., et al. P-selectin axis plays a key role in microglia immunophenotype and glioblastoma progression. Nat Commun 12, 1912 (2021). 2. Acúrcio, R.C., et al. Therapeutic targeting of PD-1/PD-L1 blockade by novel small-molecule inhibitors recruits cytotoxic T cells into solid tumor microenvironment. J Immunother Cancer 10(2022). Citation Format: Koren Salomon, Eilam Yeini, Neta Frommer, Irisi Kamer, Jair Bar, Iris Barshack, Ronit Satchi-Fainaro. Elucidating the role of P-selectin/P-selectin ligand-1 axis in lung cancer brain metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5522.

Dietary linoleic acid supplementation protects against obesity-induced microglial reactivity in mice

We investigated whether linoleic acid (LA) supplementation could modulate emotional behavior and microglia-related neuroinflammation. For that, male mice of C57BL/6J genetic background fed either a high-fat diet (HFD) or a standard diet (STD) for 12 weeks, were treated with a vehicle or LA solution for 5 weeks before being evaluated for emotional behavior using a battery of behavioral tests. The animals were subsequently sacrificed and their brains collected and processed for immunofluorescence staining, targeting microglia-specific calcium-binding proteins (IBA-1). Neuroinflammation severity was assessed in multiple hypothalamic, cortical and subcortical brain regions. We show an anxio-depressive-like effect of sustained HFD feeding that was neither alleviated nor worsened with LA supplementation. However, increased IBA-1 expression and microgliosis in the HFD group were largely attenuated by LA supplementation. These observations demonstrate that the anti-neuroinflammatory properties of LA are not restricted to hypothalamic areas but are also evident at the cortical and subcortical levels. This study discloses that neuroinflammation plays a role in the genesis of neuropsychiatric disorders in the context of obesity, and that LA supplementation is a useful dietary strategy to alleviate the impact of obesity-related neuroinflammation.

Sex differences in luteinizing hormone aggravates Aβ deposition in APP/PS1 and Aβ1-42-induced mouse models of Alzheimer's disease

Alzheimer's disease (AD) exhibits a higher incidence rate among older women, and dysregulation of the hypothalamic-pituitary-gonadal (HPG) axis during aging is associated with cognitive impairments and the development of dementia. luteinizing hormone (LH) has an important role in CNS function, such as mediating neuronal pregnenolone production, and modulating neuronal plasticity and cognition. The sex differences in LH and its impact on Aβ deposition in AD individuals remain unclear, with no reported specific mechanisms. Here, we show through data mining that LH-related pathways are significantly enriched in female AD patients. Additionally, LH levels are elevated in female AD patients and exhibit a negative correlation with cognitive levels but a positive correlation with AD pathology levels, and females exhibit a greater extent of AD pathology, such as Aβ deposition. In vivo, we observed that the exogenous injection of LH exacerbated behavioral impairments induced by Aβ1-42 in mice. LH injection resulted in worsened neuronal damage and increased Aβ deposition. In SH-SY5Y cells, co-administration of LH with Aβ further exacerbated Aβ-induced neuronal damage. Furthermore, LH can dose-dependently decrease the levels of NEP and LHR proteins while increasing the expression of GFAP and IBA1 in vivo and in vitro. Taken together, these results indicate that LH can exacerbate cognitive impairment and neuronal damage in mice by increasing Aβ deposition. The potential mechanism may involve the reduction of NEP and LHR expression, along with the exacerbation of Aβ-induced inflammation.

MyD88-mediated signaling is critical for the generation of seizure responses and cognitive impairment in a model of anti-N-methyl-D-aspartate receptor encephalitis.

We previously demonstrated that interleukin-1 receptor-mediated immune activation contributes to seizure severity and memory loss in anti-N-methyl-D-aspartate receptor (NMDAR) encephalitis. In the present study, we assessed the role of the myeloid differentiation primary response gene 88 (MyD88), an adaptor protein in Toll-like receptor signaling, in the key phenotypic characteristics of anti-NMDAR encephalitis. Monoclonal anti-NMDAR antibodies or control antibodies were infused into the lateral ventricle of MyD88 knockout mice (MyD88-/-) and control C56BL/6J mice (wild type [WT]) via osmotic minipumps for 2 weeks. Seizure responses were measured by electroencephalography. Upon completion of the infusion, the motor, anxiety, and memory functions of the mice were assessed. Astrocytic (glial fibrillary acidic protein [GFAP]) and microglial (ionized calcium-binding adaptor molecule 1 [Iba-1]) activation and transcriptional activation for the principal inflammatory mediators involved in seizures were determined using immunohistochemistry and quantitative real-time polymerase chain reaction, respectively. As shown before, 80% of WT mice infused with anti-NMDAR antibodies (n = 10) developed seizures (median = 11, interquartile range [IQR] = 3-25 in 2 weeks). In contrast, only three of 14 MyD88-/- mice (21.4%) had seizures (0, IQR = 0-.25, p = .01). The WT mice treated with antibodies also developed memory loss in the novel object recognition test, whereas such memory deficits were not apparent in MyD88-/- mice treated with anti-NMDAR antibodies (p = .03) or control antibodies (p = .04). Furthermore, in contrast to the WT mice exposed to anti-NMDAR antibodies, the MyD88-/- mice had a significantly lower induction of chemokine (C-C motif) ligand 2 (CCL2) in the hippocampus (p = .0001, Sidak tests). There were no significant changes in the expression of GFAP and Iba-1 in the MyD88-/- mice treated with anti-NMDAR or control antibodies. These findings suggest that MyD88-mediated signaling contributes to the seizure and memory phenotype in anti-NMDAR encephalitis and that CCL2 activation may participate in the expression of these features. The removal of MyD88 inflammation may be protective and therapeutically relevant.

ATP-P2X7 signaling mediates brain pathology while contributing to viral control in perinatal Zika virus infection

Zika virus (ZIKV), the causative agent of Zika fever, is a flavivirus transmitted by mosquitoes of the Aedes genus. Zika virus infection has become an international concern due to its association with severe neurological complications such as fetal microcephaly. Viral infection can induce the release of ATP in the extracellular environment, activating receptors sensitized by extracellular nucleotides, such as the P2X7 receptor. This receptor is the primary purinergic receptor involved in neuroinflammation, neurodegeneration, and immunity. In this work, we investigated the role of ATP-P2X7 receptor signaling in Zika-related brain abnormalities. Wild-type mice (WT) and P2X7 receptor-deficient (P2X7-/-) C57BL/6 newborn mice were subcutaneously inoculated with 5 × 106plaque-forming units of ZIKV or mock solution. P2X7 receptor expression increased in the brain of Zika virus-infected mice compared to the mock group. Comparative analyses of the hippocampi from WT and P2X7-/-mice revealed that the P2X7 receptor increased hippocampal damage in CA1/CA2 and CA3 regions. Doublecortin expression decreased significantly in the brains of ZIKV-infected mice. WT ZIKV-infected mice showed impaired motor performance compared to P2X7-/- infected mice. WT ZIKV-infected animals showed increased expression of glial markers GFAP (astrocytes) and IBA-1 (microglia) compared to P2X7-/- infected mice. Although the P2X7 receptor contributes to neuronal loss and neuroinflammation, WT mice were more efficient in controlling the viral load in the brain than P2X7 receptor-deficient mice. This result was associated with higher induction of TNF-α, IFN-β, and increased interferon-stimulated gene expression in WT mice than P2X7-/-ZIKV-infected. Finally, we found that the P2X7 receptor contributes to inhibiting the neuroprotective signaling pathway AKT/mTOR while stimulating the caspase-3 activation, possibly two distinct pathways contributing to neurodegeneration. These findings suggest that ATP-P2X7 receptor signaling contributes to the antiviral response in the brain of ZIKV-infected mice while increasing neuronal loss, neuroinflammation, and related brain abnormalities.

Effects of Exercise Training and L-Arginine Loaded Chitosan Nanoparticles on Hippocampus Histopathology, β-Secretase Enzyme Function, APP, Tau, Iba1and APOE-4 mRNA in Aging Rats.

The objective of this study was to evaluate the combined and independent effects of exercise training and L-Arginine loaded chitosan nanoparticles (LA CNPs) supplementation on hippocampal Tau, App, Iba1, and ApoE gene expression, oxidative stress, β-secretase enzyme activity, and hippocampus histopathology in aging rats. Thirty-five male Wistar rats were randomly assigned to five groups (n = 7 in each): Young (8 weeks old), Old (20 months old), old + L-arginine supplementation (Old Sup), old + exercise (Old Exe) and old + L-arginine supplementation + exercise (Old Sup + Exe). LA CNPs were administered to the supplement groups through gavage at a dosage of 500mg/kg/day for 6-weeks. Exercise groups were subjected to a swimming exercise program five days/week for the same duration. Upon the completion of their interventions, the animals underwent behavioral and open-field task tests and were subsequently sacrificed for hippocampus genetic and histopathological evaluation. For histopathological analysis of brain, Cresyl violet staining was used. Congo Red staining was employed to confirm amyloid plaques in the hippocampus. Expressions of Tau, App, Iba1, and ApoE genes were determined by real-time PCR. In contrast to the Old group, Old Exe and Old Sup + Exe groups spent more time in the central space in the open field task (p < 0.05) and have more live cells in the hippocampus. Old rats (Old, Old Sup and Old Exe groups) exhibited a significant Aβ peptide accumulation and increases in APP, Tau, Iba1, APOE-4 mRNA and MDA, along with decreases in SOD compared to the young group (p < 0.05). However, LA CNPs supplementation, exercise, and their combination (Old Sup, Old Exe and Old Sup + Exe) significantly reduced MDA, Aβ plaque as well as APP, Tau, Iba1, and APOE-4 mRNA compared to the Old group (p < 0.05). Consequently, the administration of LA CNPs supplements and exercise might regulate the risk factors of hippocampus cell and tissue.

Sodium butyrate activates the KATP channels to regulate the mechanism of Parkinson's disease microglia model inflammation.

Parkinson's disease (PD) is a common neurodegenerative disorder. Microglia-mediated neuroinflammation has emerged as an involving mechanism at the initiation and development of PD. Activation of adenosine triphosphate (ATP)-sensitive potassium (KATP ) channels can protect dopaminergic neurons from damage. Sodium butyrate (NaB) shows anti-inflammatory and neuroprotective effects in some animal models of brain injury and regulates the KATP channels in islet β cells. In this study, we aimed to verify the anti-inflammatory effect of NaB on PD and further explored potential molecular mechanisms. We established an in vitro PDmodel in BV2 cells using 1-methyl-4-phenylpyridinium (MPP+ ). The effects of MPP+ and NaB on BV2 cell viability were detected by cell counting kit-8 assays. The morphology of BV2 cells with or without MPP+ treatment was imaged via an optical microscope. The expression of Iba-1 was examined by the immunofluorescence staining. The intracellular ATP content was estimated through the colorimetric method, and Griess assay was conducted to measure the nitric oxide production. The expression levels of pro-inflammatory cytokines and KATP channel subunits were evaluated by reverse transcription-quantitativepolymerase chain reaction and western blot analysis. NaB (5 mM) activated the KATP channels through elevating Kir6.1 and Kir6.1 expression in MPP+ -challenged BV2 cells. Both NaB and pinacidil (a KATP opener) suppressed the MPP+ -induced activation of BV2 cells and reduced the production of nitrite and pro-inflammatory cytokines in MPP+ -challenged BV2 cells. NaB treatment alleviates the MPP+ -induced inflammatory responses in microglia via activation of KATP channels.

Curcumin abrogates cobalt-induced neuroinflammation by suppressing proinflammatory cytokines release, inhibiting microgliosis and modulation of ERK/MAPK signaling pathway

Curcumin, a bioactive polyphenol derived from turmeric, has been reported to have anti-inflammatory properties. The current study investigated the anti-inflammatory effect of curcumin in the hippocampal subfields (CA1 and CA3) after exposure to cobalt (Co) and the impact of ERK protein. Twenty-eight albino Wistar rats were divided into four groups, each with seven randomly selected rats as follows: Control (distilled water), Cobalt (Co) only (40 mg/kg), 120 mg/kg or 240 mg/kg curcumin + Co (40 mg/kg). Treatment was via oral gavage for 28 days. We performed a biochemical investigation to determine the levels of proinflammatory cytokines (TNFα and IL-1β). Furthermore, we conducted an immunohistochemical evaluation to assess the expression of IBA1 by microglial cells and the immunoexpression of ERK protein in the hippocampus. Results revealed a significant (p<0.05) elevation in the tissue level of TNFα and IL-1β, an increase in the number of IBA1-positive microglia, and upregulation of ERK protein in the hippocampal subfields of the rats after exposure to cobalt-only. Nevertheless, pretreatment with curcumin restored these parameters to levels comparable to control. In conclusion, our results showed that curcumin abrogated the Co-induced neuroinflammation by suppressing the release of proinflammatory biomarkers, reducing microgliosis, and modulating the ERK/MAPK pathway.

Depression-like Behavior Induced by Repeated Administration of Dexamethasone to Lipopolysaccharide-inflamed Mice.

Over the years, animal models of depression have been developed by loading chronic stress, inducing neuroinflammation, or administering drugs that induce depression; however, these results have poor reproducibility. Therefore, it is necessary to develop animal models that exhibit definitive symptoms of depression for studies on potential therapeutics. This study was aimed at investigating depression-like symptoms and their pathogenesis in lipopolysaccharide (LPS)-inflamed mice treated with dexamethasone (DEX). Male ICR mice were injected with LPS, followed by injection with DEX a day later and each day for 6 consecutive days. Depression-like behavior, expression of the glial markers glial fibrillary acidic protein (GFAP) and ionized calcium-binding adapter molecule 1 (Iba1), and the number of the immature neuronal marker doublecortin (DCX)-positive cells were assessed using tail-suspension test (TST), forced swim test (FST), western blot analysis, and immunohistochemical analysis. Mice in the LPS+DEX group had significantly longer immobility time in the TST and FST than did those in the LPS- or DEX-only and control groups on day 7 post-LPS administration. GFAP and Iba1 expression was significantly elevated in the hippocampus of mice in the LPS group than in those of mice in the control group. Moreover, a significantly lower number of DCX-positive cells was observed in the hippocampal dentate gyrus of mice in the LPS+DEX group compared with that in mice in the LPS- or DEX-only and control groups on day 7 after LPS administration. Repeated DEX administration to LPS-inflamed mice may induce definitive depression-like symptoms by decreasing the number of immature neurons in the hippocampal dentate gyrus. This novel mouse model of depression was produced by repeated administration of steroids to inflamed mice.

Resveratrol protects dopaminergic neurons in a mouse model of Parkinson's disease by regulating the gut-brain axis via inhibiting the TLR4 signaling pathway

To investigate the protective effect of resveratrol on intestinal barrier in 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP)-induced Parkinson's disease (PD) mouse models and its mechanism for regulating TLR4/MyD88/NF-κB signaling to protect dopaminergic neurons. Fifty-two C57BL/6J mice were randomized into control group (n= 12), MPTP group (n=14), MPTP + resveratrol (30 mg/kg) group (n=13), and MPTP + resveratrol (90 mg/kg) group (n=13), and mouse models were established by intraperitoneal MPTP (30 mg/kg) injection for 7 days in the latter 3 groups. Behavioral tests were conducted to evaluate the effect of resveratrol on motor symptoms of the mice. Western blotting was used to detect the expression of TH, α-syn, ZO-1, Claudin-1, TLR4, MyD88, and NF-κB in the brain tissues of the mice. Immunohistochemistry, immunofluorescence, ELISA and transmission electron microscopy were used to verify the effect of resveratrol for suppressing inflammation and protecting the intestinal barrier. Compared with those in the normal control group, the mice in MPTP group showed significant changes in motor function, number of dopaminergic neurons, neuroinflammation, levels of LPS and LBP, and expressions of tight junction proteins in the intestinal barrier. Resveratrol treatment significantly improved motor function of the PD mice (P < 0.01), increased the number of neurons and TH protein expression (P < 0.05), down-regulated the expressions of GFAP, Iba-1, and TLR4, lowered fecal and plasma levels of LPS and LBP (P < 0.05), restored the expression levels of ZO-1 and Claudin-1 (P < 0.01), and down-regulated the expressions of TLR4, MyD88, and NF-κB in the colon tissue (P < 0.05). The mice with resveratrol treatment at 30 mg/kg showed normal morphology of the tight junction complex with neatly and tightly arranged intestinal villi. Resveratrol repairs the intestinal barrier by inhibiting TLR4/MyD88/NF-κB signaling pathway-mediated inflammatory response, thereby improving motor function and neuropathy in mouse models of MPTP-induced PD.