Microglial Inflammatory Response Research Articles

Long-term neurological implications of severe acute respiratory syndrome coronavirus 2 infections in neonates: Innate immune memory and chronic neuroinflammation

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection can cause long-term neurological complications in adults. However, the mechanisms by which early-life SARS-CoV-2 infection increases the risk of abnormal neurodevelopment remain unknown.Recent studies have shown an association with chronic proinflammatory cytokine/chemokine production in the central nervous system (CNS). Therefore, it was hypothesised that innate immune activation and induction of innate immune memory may play a potential role in the neonatal brain. Haematopoietic stem cells in the bone marrow are exposed to SARS-CoV-2, SARS-CoV-2 envelope protein (E protein), lipopolysaccharide (LPS)-bound spike proteins (S1 and S2 proteins), and damage-associated molecular patterns (DAMPs). Myeloid progenitors enter the stroma of the choroid plexus and are further directed to incessantly supply the brain parenchyma with resident innate immune cells. The S proteins-LPS complex can cross the blood–brain barrier and plays an important role in microglial and astrocytic inflammatory responses and innate immune memory. Persistently activated microglia with memory release pro-inflammatory cytokines/chemokines which contribute to abnormal synaptic development in the frontal lobe and cerebellum, potentially leading to long-term neurological complications, similar to those observed in autism spectrum disorder (ASD). In addition, this hypothesis suggests that bacterial and fungal products may act as adjuvants to S proteins and may also explain why S proteins alone are insufficient to induce neuroinflammation in neonates.

Regulation of microglia-mediated inflammation by host lncRNA Gm20559 upon flaviviral infection

BackgroundJapanese Encephalitis Virus (JEV) and West Nile Viruses (WNV) are neurotropic flaviviruses which cause neuronal death and exaggerated glial activation in the central nervous system. Role of host long non coding RNAs in shaping microglial inflammation upon flavivirus infections has been unexplored. This study attempted to decipher the role of lncRNA Gm20559 in regulating microglial inflammatory response in context of flaviviruses. MethodsAntisense oligonucleotide LNA Gapmers designed against lncRNA Gm20559 and non-specific site (negative control) were used for Gm20559 knockdown in JEV and WNV-infected N9 microglial cells. Upon establishing successful Gm20559 knockdown, expression of various proinflammatory cytokines, chemokines, interferon-stimulated genes (ISGs) and RIG-I were checked by qRT-PCR and cytometric bead array. Western Blotting was done to analyse the phosphorylation level of various inflammatory markers and viral non-structural protein expression. Plaque Assays were employed to quantify viral titres in microglial supernatant upon knocking down Gm20559. Effect of microglial supernatant on HT22 neuronal cells was assessed by checking expression of apoptotic protein and viral non-structural protein by Western Blotting. ResultsUpregulation in Gm20559 expression was observed in BALB/c pup brains, primary microglia as well as N9 microglia cell line upon both JEV and WNV infection. Knockdown of Gm20559 in JEV and WNV-infected N9 cell led to the reduction of major proinflammatory cytokines – IL-1β, IL-6, IP-10 and IFN-β. Inhibition of Gm20559 upon JEV infection in N9 microglia also led to downregulation of RIG-I and OAS-2, which was not the case in WNV-infected N9 microglia. Phosphorylation level of P38 MAPK was reduced in case of JEV-infected N9 microglia and not WNV-infected N9 microglia. Whereas phosphorylation of NF-κB pathway was unchanged upon Gm20559 knockdown in both JEV and WNV-infected N9 microglia. However, treating HT22 cells with JEV and WNV-infected microglial supernatant with and without Gm20559 could not trigger cell death or influence viral replication. ConclusionKnockdown studies on lncRNA Gm20559 suggests its pivotal role in maintaining the inflammatory milieu of microglia in flaviviral infection by modulating the expression of various pro-inflammatory cytokines. However, Gm20559-induced increased microglial proinflammatory response upon flavivirus infection fails to trigger neuronal death.

Oral administration of sophoricoside (SOP) inhibits neuronal damage and neuroinflammation to curb neurodegeneration in Parkinson's disease

Neuronal apoptosis and neuroinflammation are key factors involved in the pathological changes of Parkinson's disease (PD). Sophoricoside (SOP) has shown anti-inflammatory and anti-apoptosis effects in various diseases. However, the role of SOP in PD has not been reported. In this experiment, we found that oral administration of SOP alleviated weight loss and motor symptoms in 1-Methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP)-injected mice. Further studies revealed that SOP inhibited inflammatory responses and neuronal apoptosis in the midbrain region of MPTP-injected mice. In vitro mechanistic study, we found that SOP exerts neuroprotective effects through a two-sided action. On the one hand, SOP inhibits Lipopolysaccharide (LPS)-induced inflammatory responses in microglia by inhibiting the Nuclear factor kappa-B(NF-κB) pathway. On the other hand, SOP inhibits 1-methyl-4-phenylpyridinium (MPP+)-induced neuronal apoptosis by regulating the Adenosine 5′-monophosphate (AMP)-activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR) signaling pathway. Thus SOP is expected to be a potential therapeutic agent for PD by targeting neuroinflammation and neuronal apoptosis.

Rumex crispus Leaf Extract Inhibits Lipopolysaccharide-Induced Inflammatory Response in BV-2 Microglia Cells

Background: Microglial cells are immune cells that operate within the central nervous system. Abnormally activated microglia cause neuroinflammation, which is linked with neurodegenerative disease. Previous research has revealed that Rumex crispus root extract exerts anti-inflammatory effects. However, it is not known whether Rumex crispus leaf extract (RLE) has anti-inflammatory effects on murine microglial cells, such as BV-2 cells. This study proposed to investigate the impact of RLE on inducing inflammation by LPS in BV-2 cells. Methods: LPS was used to induce inflammation in BV-2 cells, and then cell survival, changes in the levels of inflammation-related factors and pro-inflammatory cytokines, and NF-κB and MAPKs signaling pathway activity were evaluated in the presence or absence of RLE. Results: RLE treatment resulted in a reduction in nitric oxide (NO) production triggered by LPS without causing cytotoxic effects. In addition, both protein and mRNA expression levels of iNOS and COX-2, which were upregulated by LPS, were significantly decreased by RLE. Also, RLE effectively reduced the transcriptional expression and further suppressed the increased production of inflammatory cytokines by LPS stimulation. Additionally, RLE effectively suppressed the inflammatory response of BV-2 cells stimulated by LPS via interference with NF-κB and MAPK signaling pathways. Conclusions: Taken together, our results confirm the effective suppression of the inflammatory response induced by LPS in BV-2 cells by RLE. Consequently, we suggest that RLE holds promise as a preventive agent against diseases triggered by microglial inflammatory responses.

Chronic psychosocial stress triggers microglial-/macrophage-induced inflammatory responses leading to neuronal dysfunction and depressive-related behavior.

Chronic environmental stress and traumatic social experiences induce maladaptive behavioral changes and is a risk factor for major depressive disorder (MDD) and various anxiety-related psychiatric disorders. Clinical studies and animal models of chronic stress have reported that symptom severity is correlated with innate immune responses and upregulation of neuroinflammatory cytokine signaling in brain areas implicated in mood regulation (mPFC; medial Prefrontal Cortex). Despite increasing evidence implicating impairments of neuroplasticity and synaptic signaling deficits into the pathophysiology of stress-related mental disorders, how microglia may modulate neuronal homeostasis in response to chronic stress has not been defined. Here, using the repeated social defeat stress (RSDS) mouse model we demonstrate that microglial-induced inflammatory responses are regulating neuronal plasticity associated with psychosocial stress. Specifically, we show that chronic stress induces a rapid activation and proliferation of microglia as well as macrophage infiltration in the mPFC, and these processes are spatially related to neuronal activation. Moreover, we report a significant association of microglial inflammatory responses with susceptibility or resilience to chronic stress. In addition, we find that exposure to chronic stress exacerbates phagocytosis of synaptic elements and deficits in neuronal plasticity. Importantly, by utilizing two different CSF1R inhibitors (the brain penetrant PLX5622 and the non-penetrant PLX73086) we highlight a crucial role for microglia (and secondarily macrophages) in catalyzing the pathological manifestations linked to psychosocial stress in the mPFC and the resulting behavioral deficits usually associated with depression.

Astrocytic Extracellular Vesicles Regulated by Microglial Inflammatory Responses Improve Stroke Recovery.

There are no effective treatments for post-stroke glial scar formation, which inhibits axonal outgrowth and functional recovery after stroke. We investigated whether astrocytic extracellular vesicles (AEVs) regulated by microglia modulate glial scars and improve stroke recovery. We found that peri-infarct glial scars comprised reactive astrocytes with proliferating C3d and decreased S100A10 expression in chronic stroke. In cultured astrocytes, microglia-conditioned media and treatment with P2Y1 receptor antagonists increased and reduced the area of S100A10- and C3d-expressing reactive astrocytes, respectively, by suppressing mitogen-activated protein kinase/nuclear factor-κβ (NF-κB)/tumor necrosis factor-α (TNF-α)/interleukin-1β signaling after oxygen-glucose deprivation. Intracerebral administrations of AEVs enriched miR-146a-5p, downregulated NF-κB, and suppressed TNF-α expressions, by transforming reactive astrocytes to those with S100A10 preponderance, causing functional recovery in rats subjected to middle cerebral artery occlusion. Modulating neuroinflammation in post-stroke glial scars could permit axonal outgrowth, thus providing a basis for stroke recovery with neuroprotective AEVs.

Redox Regulation of Microglial Inflammatory Response: Fine Control of NLRP3 Inflammasome through Nrf2 and NOX4.

The role of inflammation and immunity in the pathomechanism of neurodegenerative diseases has become increasingly relevant within the past few years. In this context, the NOD-like receptor protein 3 (NLRP3) inflammasome plays a crucial role in the activation of inflammatory responses by promoting the maturation and secretion of pro-inflammatory cytokines such as interleukin-1β and interleukin-18. We hypothesized that the interplay between nuclear factor erythroid 2-related factor 2 (Nrf2) and NADPH oxidase 4 (NOX4) may play a critical role in the activation of the NLRP3 inflammasome and subsequent inflammatory responses. After priming mixed glial cultures with lipopolysaccharide (LPS), cells were stimulated with ATP, showing a significant reduction of IL1-β release in NOX4 and Nrf2 KO mice. Importantly, NOX4 inhibition using GKT136901 also reduced IL-1β release, as in NOX4 KO mixed glial cultures. Moreover, we measured NOX4 and NLRP3 expression in wild-type mixed glial cultures following LPS treatment, observing that both increased after TLR4 activation, while 24 h treatment with tert-butylhydroquinone, a potent Nrf2 inducer, significantly reduced NLRP3 expression. LPS administration resulted in significant cognitive impairment compared to the control group. Indeed, LPS also modified the expression of NLRP3 and NOX4 in mouse hippocampus. However, mice treated with GKT136901 after LPS impairment showed a significantly improved discrimination index and recovered the expression of inflammatory genes to normal levels compared with wild-type animals. Hence, we here validate NOX4 as a key player in NLRP3 inflammasome activation, suggesting NOX4 pharmacological inhibition as a potent therapeutic approach in neurodegenerative diseases.

TPL2 kinase activity regulates microglial inflammatory responses and promotes neurodegeneration in tauopathy mice.

Tumor progression locus 2 (TPL2) (MAP3K8) is a central signaling node in the inflammatory response of peripheral immune cells. We find that TPL2 kinase activity modulates microglial cytokine release and is required for microglia-mediated neuron death in vitro. In acute in vivo neuroinflammation settings, TPL2 kinase activity regulates microglia activation states and brain cytokine levels. In a tauopathy model of chronic neurodegeneration, loss of TPL2 kinase activity reduces neuroinflammation and rescues synapse loss, brain volume loss, and behavioral deficits. Single-cell RNA sequencing analysis indicates that protection in the tauopathy model was associated with reductions in activated microglia subpopulations as well as infiltrating peripheral immune cells. Overall, using various models, we find that TPL2 kinase activity can promote multiple harmful consequences of microglial activation in the brain including cytokine release, iNOS (inducible nitric oxide synthase) induction, astrocyte activation, and immune cell infiltration. Consequently, inhibiting TPL2 kinase activity could represent a potential therapeutic strategy in neurodegenerative conditions.

Metformin restrains ZIKV replication and alleviates virus-induced inflammatory responses in microglia.

The re-emergence of Zika virus (ZIKV) remains a major public health threat that has raised worldwide attention. Accumulating evidence suggests that ZIKV can cause serious pathological changes to the human nervous system, including microcephaly in newborns. Recent studies suggest that metformin, an established treatment for diabetes may play a role in viral infection; however, little is known about the interactions between ZIKV infection and metformin administration. Using fluorescent ZIKV by flow cytometry and immunofluorescence imaging, we found that ZIKV can infect microglia in a dose-dependent manner. Metformin diminished ZIKV replication without the alteration of viral entry and phagocytosis. Our study demonstrated that metformin downregulated ZIKV-induced inflammatory response in microglia in a time- and dose-dependent manner. Our RNA-Seq and qRT-PCR analysis found that type I and III interferons (IFN), such as IFNα2, IFNβ1 and IFNλ3 were upregulated in ZIKV-infected cells by metformin treatment, accompanied with the downregulation of GBP4, OAS1, MX1 and ISG15. Together, our results suggest that metformin-mediated modulation in multiple pathways may attribute to restraining ZIKV infection in microglia, which may provide a potential tool to consider for use in unique clinical circumstances.

M-Terphenylamines, Acting as Selective COX-1 Inhibitors, Block Microglia Inflammatory Response and Exert Neuroprotective Activity.

Inhibition of cyclooxygenase-2 (COX-2) has been extensively studied as an approach to reduce proinflammatory markers in acute brain diseases, but the anti-neuroinflammatory role of cyclooxygenase-1 (COX-1) inhibition has been rather neglected. We report that m-terphenylamine derivatives are selective COX-1 inhibitors, able to block microglia inflammatory response and elicit a neuroprotective effect. These compounds were synthesized via a three-component reaction of chalcones, β-ketoesters, and primary amines, followed by hydrolysis/decarboxylation of the ester group. Together with their synthetic intermediates and some urea derivatives, they were studied as inhibitors of COX-1 and COX-2. The m-terphenylamine derivatives, which were selective COX-1 inhibitors, were also analyzed for their ability to block microglia inflammatory and oxidative response. Compound 3b presented an interesting anti-inflammatory and neuroprotective profile by reducing nitrite release, ROS overproduction, and cell death in organotypic hippocampal cultures subjected to LPS. We thus show that COX-1 inhibition is a promising approach to provide enhanced neuroprotection against acute inflammatory processes, which are crucial in the development of a plethora of acute neurodegenerative injuries.

Protection of a novel velvet antler polypeptide PNP1 against cerebral ischemia-reperfusion injury

AimWe isolated a novel polypeptide PNP1 from velvet antler and investigated the role of PNP1 in ischemia reperfusion and its associated mechanism. MethodsWe built the ischemia reperfusion mouse model by the middle cerebral artery occlusion (MCAO) approach. Thereafter, PNP-1 was injected via the tail vein, and neurological function was scored. Meanwhile, the tissue injury level was detected through hematoxylin & eosin (HE) and immunohistochemical (IHC) staining, inflammatory factor levels were determined with enzyme-linked immunosorbent assay (ELISA), while protein levels through Western blotting. In addition, vascular endothelial cells were used to construct the oxygen-glucose deprivation (OGD) injury model in vitro, so as to detect the intervention effect of PNP1 on endothelial injury. Additionally, microglial cells were utilized to construct the inflammatory injury model to examine the impact of PNP1 on the polarization of microglial cells. ResultsPNP1 suppressed hypoxic cerebral injury in MCAO mice, decreased the tissue inflammatory factors, promoted tissue angiogenesis, and reduced the ischemic penumbra area. Experimental results in vitro demonstrated that, PNP1 suppressed vascular endothelial cell injury, and inhibited microglial M1 polarization as well as inflammatory response. ConclusionVelvet antler polypeptide PNP1 isolated in this study has the anti-ischemic cerebral injury effect, and its mechanism is associated with suppressing vascular endothelial cell injury and microglial cell inflammatory response.

DpdtpA, A Multi-metal Ion Chelator, Attenuates Tau Phosphorylation and Microglial Inflammatory Response via Regulating the PI3K/AKT/GSK-3β Signal Pathways

Tau protein hyperphosphorylation and formation of intracellular neurofibrillary tangles (NFTs) are one of the histopathological hallmarks of Alzheimer's disease (AD) and positively correlated with the severity of AD symptoms. NFTs contain a large number of metal ions that play an important role in regulating tau protein phosphorylation and AD progression. Extracellular tau induces primary phagocytosis of stressed neurons and neuronal loss by activating microglia. Here, we studied the effects of a multi-metal ion chelator, DpdtpA, on tau-induced microglial activation and inflammatory responses and the underlying mechanisms. Treatment with DpdtpA attenuated the increase in the expression of NF-κB and production of inflammatory cytokines, IL-1β, IL-6 and IL-10, in rat microglial cells induced by expression of human tau40 proteins. Treatment with DpdtpA also suppressed tau protein expression and phosphorylation. Moreover, treatment with DpdtpA prevented tau-induced activation of glycogen synthase kinase-3β (GSK-3β) and inhibition of phosphatidylinositol-3-hydroxy kinase (PI3K)/AKT. Collectively, these results show that DpdtpA can attenuate tau phosphorylation and inflammatory responses of microglia by regulating the PI3K/AKT/GSK-3β signal pathways, providing a new option to alleviate neuroinflammation for the treatment of AD.

Opioids Alleviate Oxidative Stress via the Nrf2/HO-1 Pathway in LPS-Stimulated Microglia

Opioids are known to have antioxidant effects and to modulate microglial function under certain conditions. It has been previously shown that opioid ligands can effectively inhibit the release of proinflammatory cytokines when stimulated with lipopolysaccharide (LPS) and convert microglia to an anti-inflammatory polarization state. Here, we used C8-B4 cells, the mouse microglial cell line activated by LPS as a model to investigate the anti-inflammatory/antioxidant potential of selected opioid receptor agonists (DAMGO, DADLE, and U-50488). We found that all of these ligands could exert cytoprotective effects through the mechanism affecting LPS-induced ROS production, NADPH synthesis, and glucose uptake. Interestingly, opioids elevated the level of reduced glutathione, increased ATP content, and enhanced mitochondrial respiration in microglial cells exposed to LPS. These beneficial effects were associated with the upregulation of the Nrf2/HO-1 pathway. The present results indicate that activation of opioid signaling supports the preservation of mitochondrial function with concomitant elimination of ROS in microglia and suggest that an Nrf2/HO-1 signaling pathway-dependent mechanism is involved in the antioxidant efficacy of opioids. Opioid receptor agonists may therefore be considered as agents to suppress oxidative stress and inflammatory responses of microglia.

A Human Microglial Cell Line Expresses γ-Aminobutyric Acid (GABA) Receptors and Responds to GABA and Muscimol by Increasing Production of IL-8

Gamma-aminobutyric acid (GABA) is an essential neurotransmitter and an important regulator of neuroinflammation and disease. Microglia are important immune cells in the brain that express GABA receptors (GABAR) and respond to both GABA and GABAR agonists, yet the effect of GABA on microglial inflammatory responses is unclear. We hypothesized that GABA and GABAR agonists might modify the activation of a human microglial cell line (HMC3). We further hypothesized that Amanita muscaria extract (AME-1), which contained GABAR agonists (GABA and muscimol), would similarly stimulate HMC3. Ligand-gated GABAR (GABAAR) and G protein-coupled GABAR (GABABR) subunit expression was analyzed by qRT-PCR, metabolic activity was determined by nicotinamide adenine dinucleotide (NADH)-dependent oxidoreductase assay (XTT), reactive oxygen species (ROS) generation was analyzed by 2′,7′-dichlorodihydrofluorescein diacetate (DCFDA), and interleukin-8 (IL-8) production was analyzed by an enzyme-linked immunosorbent assay (ELISA). HMC3 expressed several neuroreceptors such as subunits of the GABAA receptor (GABAAR). HMC3 constitutively produce IL-8 and ROS. Both muscimol and GABA stimulated HMC3 to produce more IL-8 but had no effect on constitutive ROS production. GABA and muscimol altered the morphology and Iba1 localization of HMC3. GABA, but not muscimol, increased HMC3 metabolic activity. Similarly, AME-1 induced HMC3 to produce more IL-8 but not ROS and altered cell morphology and Iba1 localization. GABA induction of IL-8 was blocked by bicuculline, an antagonist of GABAAR. AME-1-induced production of IL-8 was not blocked by bicuculline, suggesting that AME-1’s effect on HMC3 was independent of GABAAR. In conclusion, these data show that GABA and GABA agonists stimulate HMC3 to increase their production of IL-8. Mixtures that contain GABA and muscimol, such as AME-1, have similar effects on HMC3 that are independent of GABAAR.

DEAD-box helicase 54 regulates microglial inflammatory response in rats with chronic constriction injuries through NF-κB/NLRP3 signaling axis.

Neuropathic pain (NP) is caused by damage to or disease of the somatosensory nervous system, but its mechanism is still not fully understood. In this study, DEAD-box helicase 54 (DDX54) was targeted, and its regulatory role was explored in a chronic constriction injury (CCI) rat model. Microglia and HMC3 cells were stimulated with LPS. The interaction between DDX54 and myeloid differentiation factor-88 adapter protein (MYD88) was verified. A CCI of sciatic nerve model in rats was established. Behavioral testing was performed before and after the CCI. The expressions of IL-1β, TNF-α, and IL-6 were upregulated, and those of DDX54, MYD88, NF-κB, and NOD-like receptor 3 (NLRP3) were upregulated in microglia and HMC3 cells after LPS induction. DDX54 knockdown in microglia and HMC3 cells inhibited IL-1β, TNF-α, and IL-6 expressions and downregulated the protein levels of MYD88, p-NF-κB p65 (p-p65), and NLRP3. DDX54 overexpression promoted the stability of MYD88 mRNA. DDX54 binds to the MYD88-3'-untranslated region (UTR). DDX54 interference in rats could alleviate the decrease of paw withdrawal mechanical threshold (PWMT) and paw withdrawal thermal latency (PWTL) induced by CCI, inhibit Iba1 expression, and reduce inflammatory factors as well as MYD88 and NF-κB expressions. DDX54 promotes the activation of NF-κB/NLRP3 signaling by regulating MYD88 mRNA stability, thereby affecting inflammatory response and NP progression in CCI rats.NEW & NOTEWORTHY The role of DDX54 protein in LPS-induced microglia and a chronic constriction injury (CCI) rat model was investigated for the first time. DDX54 interference can inhibit microglial activation and reduce the secretion of inflammatory factors. The interaction between DDX54 protein and MYD88 mRNA was explored for the first time. DDX54 promotes NF-κB/NLRP3 signaling activation by regulating MYD88 transcription in a CCI rat model.

Protective effect of TNIP2 on the inflammatory response of microglia after spinal cord injury in rats

BackgroundSpinal cord injury (SCI) is a devastating disease that can lead to tissue loss and neurological dysfunction. TNIP2 is a negative regulator of NF-κB signaling due to its capacity to bind A20 and suppress inflammatory cytokines-induced NF-κB activation. However, the anti-inflammatory role of TNIP2 in SCI remains unclear. Our study's intention was to evaluate the effect of TNIP2 on the inflammatory response of microglia after spinal cord injury in rats. MethodsHE staining and Nissl staining were performed on day 3 following SCI to analyze the histological changes. To further investigate the functional changes of TNIP2 after SCI, we performed immunofluorescence staining experiments. The effect of LPS on TNIP2 expression in BV2 cells was examined by western blot. The levels of TNF-α, IL-1β, and IL-6 in spinal cord tissues of rats with SCI and in BV2 cells with LPS were measured by using qPCR. ResultsTNIP2 expression was closely associated with the pathophysiology of SCI in rats, and TNIP2 was involved in regulating functional changes in microglia. TNIP2 expression was increased during SCI in rats and that overexpression of TNIP2 inhibited M1 polarization and pro-inflammatory cytokine production in microglia, which might ultimately protect against inflammatory responses through the MAPK and NF-κB signaling pathways. ConclusionsThe present study provides evidence for a role of TNIP2 in the regulation of inflammation in SCI and suggests that induction of TNIP2 expression alleviated the inflammatory response of microglia.

Effectiveness of Albumin-Fused Thioredoxin against 6-Hydroxydopamine-Induced Neurotoxicity In Vitro.

Parkinson's disease (PD) is a neurodegenerative disorder caused by oxidative stress-dependent loss of dopaminergic neurons in the substantia nigra and elevated microglial inflammatory responses. Recent studies show that cell loss also occurs in the hypothalamus in PD. However, effective treatments for the disorder are lacking. Thioredoxin is the major protein disulfide reductase in vivo. We previously synthesized an albumin-thioredoxin fusion protein (Alb-Trx), which has a longer plasma half-life than thioredoxin, and reported its effectiveness in the treatment of respiratory and renal diseases. Moreover, we reported that the fusion protein inhibits trace metal-dependent cell death in cerebrovascular dementia. Here, we investigated the effectiveness of Alb-Trx against 6-hydroxydopamine (6-OHDA)-induced neurotoxicity in vitro. Alb-Trx significantly inhibited 6-OHDA-induced neuronal cell death and the integrated stress response. Alb-Trx also markedly inhibited 6-OHDA-induced reactive oxygen species (ROS) production, at a concentration similar to that inhibiting cell death. Exposure to 6-OHDA perturbed the mitogen-activated protein kinase pathway, with increased phosphorylated Jun N-terminal kinase and decreased phosphorylated extracellular signal-regulated kinase levels. Alb-Trx pretreatment ameliorated these changes. Furthermore, Alb-Trx suppressed 6-OHDA-induced neuroinflammatory responses by inhibiting NF-κB activation. These findings suggest that Alb-Trx reduces neuronal cell death and neuroinflammatory responses by ameliorating ROS-mediated disruptions in intracellular signaling pathways. Thus, Alb-Trx may have potential as a novel therapeutic agent for PD.

Thymopentin (TP-5) prevents lipopolysaccharide-induced neuroinflammation and dopaminergic neuron injury by inhibiting the NF-κB/NLRP3 signaling pathway.

Neuroinflammation plays a pivotal role in neurodegenerative diseases, including Parkinson's disease, Alzheimer's disease, amyotrophic lateral sclerosis and stroke, and is accompanied by excessive release of inflammatory cytokines and mediators by activated microglia. Microglial inflammatory response inhibition may be an effective strategy for preventing inflammatory disorders. However, the reciprocal connections between the central nervous system (CNS) and immune system have not been elucidated. Thus far, these links have been proven to mainly involve immuno- and neuropeptides. The pentapeptide thymopentin (TP-5) exerts a significant immunomodulatory effect; however, its antineuroinflammatory effects and underlying mechanism are still unclear. In this study, lipopolysaccharide (LPS) was used to establish an inflammation model, and the therapeutic effect of TP-5 was evaluated. Behavioral tests showed that TP-5 treatment could improve the performance of LPS-treated mice in the open field and pole test, but not hanging wire test. TP-5 also attenuated neuronal lesions in the brains of LPS-treated mice. TP-5 reduced cytotoxicity and morphological changes in activated microglia. Label-free quantitative analysis indicated that the expression of multiple proteins and the activation of associated signaling pathways were altered by TP-5. Moreover, TP-5 could inhibit LPS-induced neuroinflammation in the brain and BV2 microglia and the expression of major genes in the NF-κB/NLRP3 signaling pathway. Additionally, tyrosine hydroxylase (TH) expression downregulation was rescued in the LPS+TP-5 group compared with the LPS group. We conclude that TP-5 exerts neuroprotection by alleviating LPS-induced inflammatory damage and dopaminergic neurodegeneration. The protective effect of TP-5 may involve the NF-κB/NLRP3 signaling pathway.

Microglial Responses to Stress-Induced Depression: Causes and Consequences.

Chronic stress is a major risk factor for various psychiatric diseases, including depression; it triggers various cellular and structural changes, resulting in the alteration of neurocircuitry and subsequent development of depression. Accumulating evidence suggests that microglial cells orchestrate stress-induced depression. Preclinical studies of stress-induced depression revealed microglial inflammatory activation in regions of the brain that regulate mood. Although studies have identified several molecules that trigger inflammatory responses in microglia, the pathways that regulate stress-induced microglial activation remain unclear. Understanding the exact triggers that induce microglial inflammatory activation can help find therapeutic targets in order to treat depression. In the current review, we summarize the recent literature on possible sources of microglial inflammatory activation in animal models of chronic stress-induced depression. In addition, we describe how microglial inflammatory signaling affects neuronal health and causes depressive-like behavior in animal models. Finally, we propose ways to target the microglial inflammatory cascade to treat depressive disorders.

Formononetin Inhibits Microglial Inflammatory Response and Contributes to Spinal Cord Injury Repair by Targeting the EGFR/MAPK Pathway

ABSTRACT Zhenbao Pill contains many Chinese herbal medicinal ingredients and has been proven to have therapeutic effects on the repair of spinal cord injury (SCI). This study attempts to investigate the role of formononetin (FMN), an ingredient of Zhenbao Pill, in regulating neuroinflammation after SCI and the underlying mechanism. Primary microglia isolated from the spinal cord of newborn rats and human microglial clone 3 (HMC3) cells were stimulated with IL-1β followed by FMN incubation. The cell viability and inflammatory cytokine levels were detected. The target of FMN was predicted and screened using databases. By silencing or overexpression of epidermal growth factor receptor (EGFR), the anti-neuroinflammatory effect of FMN was assessed in vitro. In vivo, FMN was intraperitoneally injected into rats after SCI followed by the neurological function and histopathology examination. The isolated microglia were in high purity, and the different concentrations of FMN incubation had no toxic effects on primary microglia and HMC3 cells. FMN reduced the inflammatory cytokine levels (TNF-α and IL-6) in a concentration-dependent manner. EGFR silencing or FMN incubation decreased p-EGFR and p-p38 levels and down-regulated inflammatory cytokine levels in IL-1β-stimulated cells or supernatants. Nevertheless, the effects of FMN on microglial inflammation were reversed by EGFR overexpression. In vivo, FMN treatment improved the neuromotor function, repaired tissue injury, and inhibited EGFR/p38MAPK phosphorylation. Formononetin inhibits microglial inflammatory response and contributes to SCI repair via the EGFR/p38MAPK signaling pathway.