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The PICK1/TLR4 complex on microglia is involved in the regulation of LPS-induced sepsis-associated encephalopathy
Abstract
The treatment options for sepsis-associated encephalopathy caused by systemic inflammation are still not sufficient. Protein kinase C interaction protein 1 (PICK1) has attracted much attention because of its important physiological functions in many tissues. However, its role in sepsis-associated encephalopathy remains elusive. Our study results revealed that the expression levels of PICK1 protein in mice with lipopolysaccharide-induced sepsis-associated encephalopathy were not significantly changed, but PICK1 deficiency led to excessive activation of microglia and Toll-like receptor (TLR)4 pathways, which aggravated the sepsis- associated encephalopathy. We also observed that PICK1 and TLR4 form a complex in microglial cells, thereby providing brain protection. These findings contribute to our understanding of the important role of PICK1 in sepsis and may provide novel therapeutic targets to treat sepsis-associated encephalopathy.
... Recent research has consistently demonstrated the critical role of the NF-κB signaling pathway in microglial [105]. In addition, research has presented that Protein kinase C interaction Protein 1 (PICK1) can form a complex with TLR4, offering neuroprotection in SAE [106]. During sepsis, LPS from gram-negative bacteria binds to TLR4 on microglia, initiating both MyD88 and TRIF-dependent signaling pathways as well as inflammasome activation [106]. ...
... In addition, research has presented that Protein kinase C interaction Protein 1 (PICK1) can form a complex with TLR4, offering neuroprotection in SAE [106]. During sepsis, LPS from gram-negative bacteria binds to TLR4 on microglia, initiating both MyD88 and TRIF-dependent signaling pathways as well as inflammasome activation [106]. In addition to LPS, host-derived danger signals such as HMGB1 and ATP, which are released during sepsis, can also activate microglial TLR2/4 and P2 × 7 receptors. ...
... Recent studies focusing on microglia have been conducted to explore potential SAE treatments. Several analyses have demonstrated that inhibiting microglial activation can alleviate neurological symptoms and improve long-term cognitive function [106,114,128,188]. In light of these findings, we have compiled an overview of the latest therapeutic strategies targeting microglia for the treatment of cognitive impairment in SAE. ...
Sepsis-associated encephalopathy (SAE) is a neurological dysfunction induced by sepsis, with symptoms ranging from mild delirium to deep coma. About 70% of patients with severe systemic infection develop SAE and with more than half of surviving patients suffering from long-term cognitive deficits, which seriously damaged the quality of their daily life and brought a heavy burden to society. The pathogenesis of SAE is multifactorial, including activated inflammation, blood- brain barrier (BBB) disruption, cerebral blood flow impairment, and neurotransmitter disturbances. Microglia mediate multiple SAE pathologies. In this review, we summarized the most recent findings in the roles of microglia in every stage of SAE pathogenesis, focusing on the molecular pathways in microglia activation and downstream effects. We also demonstrated the novel therapeutic studies targeting microglia in SAE. Deep insight into the role of microglia in SAE is of great importance in exploring pathogenesis and developing effective remedies of SAE.
... 63 Toll-like receptors are key in pathogen recognition and immune activation. 64 In LPS-induced SAE mouse models, activation of the TLR4 signaling pathway in microglial cells exacerbates SAE, 65 while this activation is dependent on the transcriptional expression of IRF7. 66 The TLR signaling pathway has also been shown to stimulate CCL2 activation in renal tubular epithelial cells, aggravating sepsis-induced renal injury. ...
Purpose
Sepsis can induce sepsis-associated encephalopathy (SAE), with Ulinastatin (UTI) serving a critical anti-inflammatory role. This study aimed to identify the hub genes in an SAE mouse model following UTI intervention and investigate the underlying molecular mechanisms.
Materials and Methods
Through differential expression analysis to obtain differentially expressed genes (DEGs), ie, UTI vs CLP (DEGs1) and Con vs CLP (DEGs2). After taking the intersection of the genes with opposite differential trends in these two parts and immune-related genes (IRGs), DE-IRGs were obtained. Hub genes in the protein-protein interaction (PPI) network were then determined using six algorithms from the Cytohubba plugin in Cytoscape. Gene set enrichment analysis (GSEA) was employed to explore the functional relevance of these hub genes. Additionally, the immune microenvironment across the three groups was compared, and hub gene-related drugs were predicted using an online database. Finally, qRT-PCR was used to validate the expression of the hub genes in hippocampal tissue from CLP mice.
Results
RNA sequencing obtained 864 differentially expressed genes (DEGs) (CLP vs Con) and 279 DEGs (UTI vs CLP). Taking the intersection of DEGs with opposite expression trends yielded 165 DEGs. Six key genes (ICAM - 1, IRF7, IL - 1β, CCL2, IL - 6 and SOCS3) were screened by six algorithms. Immune infiltration analysis found that Treg cells were reversed after treatment with UTI in the diseased state. A total of 106 hub - gene - related drugs were predicted, among which BINDARIT - CCL2 and LIFITEGRAST - ICAM1 showed particularly high affinities. The qRT - PCR verification results were consistent with the sequencing results.
Conclusion
In conclusion, ICAM-1, IRF7, IL-1β, CCL2, IL-6, and SOCS3 were identified as potential therapeutic targets in SAE mice treated with UTI. This study offers theoretical support for UTI as a treatment option for SAE.
... Its deficiency impairs autophagic function, exacerbating sepsisinduced acute lung injury (Mo et al., 2018). In sepsis-associated encephalopathy, PICK1 forms a complex with TLR4, exerting a protective effect against inflammatory damage (Wang et al., 2021). Additionally, PICK1 mitigates lipopolysaccharide-induced apoptosis in renal tubular epithelial cells, protecting against sepsis-related acute kidney injury (Dou et al., 2021). ...
PICK1 plays a crucial role in mammalian spermatogenesis. Here, we integrated single-molecule long-read and short-read sequencing to comprehensively examine PICK1 expression patterns in adult Baoshan pig (BS) testes. We identified the most important transcript ENSSSCT00000000120 of PICK1, obtaining its full-length coding sequence (CDS) spanning 1254 bp. Gene structure analysis located PICK1 on pig chromosome 5 with 14 exons. Protein structure analysis reflected that PICK1 consisted of 417 amino acids containing two conserved domains, PDZ and BAR_PICK1. Phylogenetic analysis underscored the evolutionary conservation and homology of PICK1 across different mammalian species. Evaluation of protein interaction network, KEGG, and GO pathways implied that interacted with 50 proteins, predominantly involved in glutamatergic synapses, amphetamine addiction, neuroactive ligand-receptor interactions, dopaminergic synapses, and synaptic vesicle recycling, and PICK1 exhibited significant correlation with DLG4 and TBC1D20. Functional annotation identified that PICK1 was involved in 9 GOs, including seven cellular components and two molecular functions. ceRNA network analysis suggested BS PICK1 was regulated by seven miRNA targets. Moreover, qPCR expression analysis across 15 tissues highlighted that PICK1 was highly expressed in the bulbourethral gland and testis. Subcellular localization analysis in ST (Swine Tesits) cells demonstrated that PICK1 significantly localized within the cytoplasm. Overall, our findings shed new light on PICK1’s role in BS reproduction, providing a foundation for further functional studies of PICK1.
... With the aim to stimulate the neuroinflammation we used LPS. LPS induces the transformation of microglial cells from their resting form into an activated M1 phenotype via receptor complex CD14/TLR4/MD2 and stimulates the production of detrimental proinflammatory cytokines, chemokines and prostaglandins (Abdo Qaid et al. 2024;Wang et al. 2021). The result showed that 250 ng/ml LPS was sufficient to stimulate inflammatory response, characterized by morphology changes in microglia, the transformation from the "resting" ramified to activated "alerted" phenotype ( Figure 2A). ...
Tauopathies encompass a group of disorders characterized by abnormal accumulation of tau protein into neurofibbrilary pathology and neuroinflammation. Understanding how glial cells interact with neurons is important to unraveling the complex mechanisms driving the progression of tauopathies. In this study, we established a multi-component cellular model for tauopathies comprising neuronal cells inducible expressing pathological truncated tau, primary microglia, and astrocytes. We successfully identified optimal culture conditions, with 1% serum supplemented with B27 proving to be most effective in enhancing neuronal protein expression, tau levels, and cell viability. This condition supported improved neuronal differentiation and synaptic marker expression, reflecting a more robust neuronal phenotype compared to monocultures. The increase in pro-inflammatory cytokines production underscores the model's capability to replicate the inflammatory environment characteristic of neurodegenerative diseases. Importantly, we observed that truncated tau significantly modulates key signalling pathways, notably the CX3CL1-CX3CR1 and CD47-SIRP-α pathways. We found that pathological changes in tauopathies altered neuron-glia interactions, leading them towards a more quiescent glial state.The alterations in signaling were accompanied by changes in tau phosphorylation, with neuroinflammation exacerbating hyperphosphorylation of truncated tau, while minimally affecting endogenous tau. In conclusion, our study highlights that pathological truncated tau significantly affects the expression of membrane-anchored fractalkine. This differential modulation, coupled with changes in inflammatory conditions, suggests that early-stage tau pathology can influence neuroinflammatory responses and may still allow neuronal cells to engage in protective mechanisms.
... Immune reactions in the brain are mainly characterized by the reactivity of brain resident immune cells, namely microglia. Microglial reactivity results in production of various cytokines and chemokines as well as the production of complement component 1q (C1q), prostaglandin E2 (PGE2) and free radicals (NO and ROS) [8][9][10]. In addition to microglia, astrocytes, cells with immunological properties are involved in neuroinflammatory processes. ...
Despite advances in antimicrobial and anti-inflammatory treatment, inflammation and its consequences remain a major challenge in the field of medicine. Inflammatory reactions can lead to life-threatening conditions such as septic shock, while chronic inflammation has the potential to worsen the condition of body tissues and ultimately lead to significant impairment of their functionality. Although the central nervous system has long been considered immune privileged to peripheral immune responses, recent research has shown that strong immune responses in the periphery also affect the brain, leading to reactive microglia, which belong to the innate immune system and reside in the brain, and neuroinflammation. The inflammatory response is primarily a protective mechanism to defend against pathogens and tissue damage. However, excessive and chronic inflammation can have negative effects on neuronal structure and function. Neuroinflammation underlies the pathogenesis of many neurological and neurodegenerative diseases and can accelerate their progression. Consequently, targeting inflammatory signaling pathways offers potential therapeutic strategies for various neuropathological conditions, particularly Parkinson’s and Alzheimer’s disease, by curbing inflammation. Here the blood–brain barrier is a major hurdle for potential therapeutic strategies, therefore it would be highly advantageous to foster and utilize brain innate anti-inflammatory mechanisms. The tricarboxylic acid cycle-derived metabolite itaconate is highly upregulated in activated macrophages and has been shown to act as an immunomodulator with anti-inflammatory and antimicrobial functions. Mesaconate, an isomer of itaconate, similarly reduces the inflammatory response in macrophages. Nevertheless, most studies have focused on its esterified forms and its peripheral effects, while its influence on the CNS remained largely unexplored. Therefore, this study investigated the immunomodulatory and therapeutic potential of endogenously synthesized itaconate and its isomer mesaconate in lipopolysaccharide (LPS)-induced neuroinflammatory processes. Our results show that both itaconate and mesaconate reduce LPS-induced neuroinflammation, as evidenced by lower levels of inflammatory mediators, reduced microglial reactivity and a rescue of synaptic plasticity, the cellular correlate of learning and memory processes in the brain. Overall, this study emphasizes that both itaconate and mesaconate have therapeutic potential for neuroinflammatory processes in the brain and are of remarkable importance due to their endogenous origin and production, which usually leads to high tolerance.
Supplementary Information
The online version contains supplementary material available at 10.1186/s12974-024-03188-3.
... Focusing on the TLRs, activation of the microglial TLR-2 and TLR-4 has been related to neuroinflammation and neuronal cell death [61,62]. Therefore, governing the LPS-stimulated microglia activation through downregulation of the TLR-4 receptor and its mediated signaling pathway proteins in addition to the suppression of the production of neurotoxic proinflammatory cytokines would be an efficient therapeutic approach for neuroinflammatory ailments. ...
Backgrounds and objectives
Microglia play a regulatory role in central nervous system inflammatory diseases, such as Alzheimer’s, Parkinson’s, and multiple sclerosis. Natural remedies like black cumin seeds ( Nigella sativa ) are rich in bioactive compounds that potentially can modulate inflammatory processes in the brain. In the current work, we studied the protective and anti-inflammatory properties of black cumin seed oil (BCSO) and its nano-form on lipopolysaccharide (LPS)-induced neurotoxicity in mice.
Materials and methods
Forty-eight mice were divided randomly into eight groups ( n =6), three control groups (negative control, BCSO control, nano-BCSO control), LPS group, and four treatment groups [BCSO+LPS, nano-BCSO+LPS, indomethacin (5 mg/kg)+LPS, BCSO+indomethacin(2.5 mg/kg)+LPS]. At the end of the experiment, the brain tissues were removed for histopathological and biochemical assessments. Malondialdehyde and interleukin (IL)-10 were assessed using enzyme-linked immunosorbent assay while the gene expression of IL-6, toll-like receptor-4, brain-derived neurotrophic factor, nerve growth factor, cyclooxygenase-2, and B-cell lymphoma-2 were assessed by real-time PCR. IL-1β was quantified immunohistochemically along with the histopathological studies of the cerebral cortex of mice brains.
Results and conclusions
In our study, BCSO and its nano-form demonstrated a reduction in LPS-induced neurotoxicity, exhibiting comparable or better anti-inflammatory effects to indomethacin. These treatments significantly elevated the gene expression levels of neuroprotective factors brain-derived neurotrophic factor and nerve growth factor in LPS-treated mice. Pretreatment with BCSO and its nano-form reduced the malondialdehyde levels, in addition to gene expressions of cyclooxygenase-2, toll-like receptor-4, IL-6, and B-cell lymphoma-2. Immunohistochemical analysis indicated a decrease in IL-1β with BCSO and the lowering effect of the nano-form was superior. The histopathological studies corroborated with biochemical and molecular findings, suggesting that BCSO and its nano-form attenuated the inflammation and enhanced the microglial antioxidative and anti-inflammatory status. BCSO could enhance the anti-inflammatory activity of indomethacin, so lower doses of indomethacin with BCSO may be suggested for protecting against the adverse effects of high doses of NSAIDs as gastritis. Consequently, BCSO can serve a potential stimulatory supplement of the immunity for neurodegenerative conditions.
... Moreover, a previous study illustrated that the PICK-TLR4 complex in microglia was involved in electroacupuncture to prevent neuroinflammation in sepsis-related encephalopathy 33 . Wang et al. have previously demonstrated that TLR4 can form a complex with PICK1 and then be transported to the cell membrane 34 , and the PICK1/TLR4 complex has also been implicated in the regulation of sepsis 35 . Consistent with this, the PICK1 in macrophages was found to interact with TLR4 protein via co-immunoprecipitation in this study. ...
Acute liver injury (ALI) may manifest at any phase of sepsis, yet an explicit therapeutic approach remains elusive. In this study, LPS and cecum ligation and puncture (CLP) were utilized to establish an inflammatory cell model and a murine model of sepsis-induced liver injury, respectively, aiming to explore the potential protective effect of protein interacting with C α kinase 1 (PICK1) on sepsis-induced ALI and its underlying mechanisms. In both the cell supernatant and the murine whole blood, the concentrations of inflammatory factors were quantified by ELISA, while the protein and mRNA expressions of PICK1, cleaved-PARP-1, caspase1, TLR4, IκBα, and NF-κB were assessed via western blot and qRT-PCR. The outcomes revealed that the knockdown of PICK1 increased the levels of inflammatory factors and apoptosis, alongside activation of TLR4/NF-κB signaling pathway-related factors in both in vivo and in vitro models. Moreover, the murine liver samples were subjected to Hematoxylin–Eosin (HE) staining for assessment of histopathological morphology. The HE staining and liver injury scoring results manifested a markedly exacerbated hepatic damage in PICK1 knockout mice as compared to WT mice following CLP. Furthermore, the liver macrophages were isolated from murine livers, and the expression and activity of the factors associated with the TLR4/NF-κB signaling pathway were verified through RT-qPCR and western blot, and EMSA assay demonstrated an augmented NF-κB activity subsequent to PICK1 knockout. Finally, the expression and localization of PICK1 in macrophages were further scrutinized via immunofluorescence, and the interaction between PICK1 and TLR4 was identified through co-immunoprecipitation. In conclusion, the knockdown of PICK1 appeared to modulate inflammatory factors by activating the TLR4/NF-κB signaling pathway, thereby exacerbating hepatic damage induced by sepsis.
... LPS и липопротеины могут действовать синергически, продуцируя цитокины отдельными путями, что позволяет предположить, что липопротеины могут играть важную роль в механизме развития сепсиса [70]. Кроме того, введение мышам липопротеина индуцировала перекрёстную толерантность к последующему воздействию как липопротеина, так и LPS, и даже целых бактерий в моделях эндотоксемии и перевязки слепой кишки и пункции [71]. Перекрёстная толерантность может опосредоваться через TLR2 и черезTLR6 мышиных макрофагов [43]. ...
Sepsis is a life-threatening organ dysfunction caused by a disturbed response to infection. The development of sepsis is preceded by Systemic Inflammatory Response Syndrome (SIRS), which is the overall inflammatory response of the body to severe lesion. The role of opportunistic pathogens in the development of CER and sepsis may be considered proven, but the value of intestinal microbiome remains underestimated. The survey systematized data on the role of cell wall or membrane components of Gram-positive and Gram-negative bacteria - representatives of intestinal microbiome in pathogenesis of SIRS and sepsis.
... TLR4 was the first recognized member of the TLR family and is widely expressed in primary sensory neurons and glial cells. It is a vital transmembrane pattern-recognition receptor of the innate immune system and is involved in different pathological processes, such as sepsis, cardiac diseases, and ischemia/reperfusion injury [16,17]. TLR4 is also implicated in acute symptomatic seizures and contributes to an increased risk of chronic epilepsy [19]. ...
The toll-like receptor 4 (TLR4) pathway is involved in seizures. We investigated whether ultrasound-targeted microbubble destruction (UTMD)-mediated delivery of short hairpin RNA (shRNA) targeting the TLR4 gene (shRNA-TLR4) can reduce ischemia-induced seizures in rats with hyperglycemia. A total of 100 male Wistar rats were randomly assigned to five groups: (1) Sham; (2) normal saline (NS); (3) shRNA-TLR4, where rats were injected with shRNA-TLR4; (4) shRNA-TLR4 + US, where rats were injected with shRNA-TLR4 followed by ultrasound (US) irradiation; and (5) shRNA-TLR4 + microbubbles (MBs) + US, where rats were injected with shRNA-TLR4 mixed with MBs followed by US irradiation. Western blot and immunohistochemical staining were used to measure TLR4-positive cells. Half of the rats in the NS group developed tonic-clonic seizures, and TLR4 expression in the CA3 region of the hippocampus was increased in these rats. In addition, the NS group showed an increased number of TLR4-positive cells compared with the Sham group, while there was a decreased number of TLR4-positive cells in the shRNA, shRNA + US, and shRNA + MBs + US groups. Our findings indicate that the TLR4 pathway is involved in the pathogenesis of ischemia-induced seizures in hyperglycemic rats and that UTMD technology may be a promising strategy to treat brain diseases.
... When activated, microglia can generate many cytokines and metalloproteinases and cause neuronal dysfunction and memory impairment [5,64]. Activated microglia are consistently observed in both experimental models and septic patients; in particular, there is increased microglial activation in hippocampal tissue in patients who die of SAE [65][66][67]. In addition to microglial activation, astrocyte activation is also one of the most relevant phenomena in SAE [63,68,69]. ...
Sepsis-Associated Encephalopathy (SAE) is common in sepsis patients, with high mortality rates. It is believed that neuroinflammation is an important mechanism involved in SAE. High mobility group box 1 protein (HMGB1), as a late pro-inflammatory factor, is significantly increased during sepsis in different brain regions, including the hippocampus. HMGB1 causes neuroinflammation and cognitive impairment through direct binding to advanced glycation end products (RAGE) and Toll-like receptor 4 (TLR4). Electroacupuncture (EA) at Baihui (GV20) and Zusanli (ST36) is beneficial for neurological diseases and experimental sepsis. Our study used EA to treat SAE induced by lipopolysaccharide (LPS) in male Sprague–Dawley rats. The Y maze test was performed to assess working memory. Immunofluorescence (IF) and Western blotting (WB) were used to determine neuroinflammation and the HMGB1 signaling pathway. Results showed that EA could improve working memory impairment in rats with SAE. EA alleviated neuroinflammation by downregulating the hippocampus’s HMGB1/TLR4 and HMGB1/RAGE signaling, reducing the levels of pro-inflammatory factors, and relieving microglial and astrocyte activation. However, EA did not affect the tight junctions’ expression of the blood–brain barrier (BBB) in the hippocampus.
... Microglia are the primary brain cells expressing pattern recognition receptors (PRRs) such as toll-like (TLRs) and nod-like (NLRs) receptors to recognize accumulating PAMPs and DAMPs [21]. When activated, microglial TLRs (mainly TLR2 and TLR4), acting through MyD88-dependent signaling cascade, trigger the NF-κB/MAPK pathway leading to secretion of proinflammatory molecules including TNF-α, IL-1, IL-6, IL-12, and IFNγ [21,49]. Both experimental and clinical sepsis displayed rapid microglial reactivity, consistently accompanied by the production of these cytokines [16][17][18]20]. ...
Sepsis and acute liver failure are associated with severe endogenous intoxication. Microglia, which are the resident immune brain cells, play diverse roles in central nervous system development, surveillance, and defense, as well as contributing to neuroinflammatory reactions. In particular, microglia are fundamental to the pathophysiology of reactive toxic encephalopathies. We analyzed microglial ultrastructure, morphotypes, and phagocytosis in the sensorimotor cortex of cecal ligation and puncture (CLP) and acetaminophen-induced liver failure (AILF) Wistar rats. A CLP model induced a gradual shift of ~50% of surveillant microglia to amoeboid hypertrophic-like and gitter cell-like reactive phenotypes with active phagocytosis and frequent contacts with damaged neurons. In contrast, AILF microglia exhibited amoeboid, rod-like, and hypertrophic-like reactive morphotypes with minimal indications for efficient phagocytosis, and were mostly in contact with edematous astrocytes. Close interactions of reactive microglia with neurons, astrocytes, and blood–brain barrier components reflect an active contribution of these cells to the tissue adaptation and cellular remodeling to toxic brain damage. Partial disability of reactive microglia may affect the integrity and metabolism in all tissue compartments, leading to failure of the compensatory mechanisms in acute endogenous toxic encephalopathies.
... These subtypes differ in ligand specifity, cellular localization, mode of multimerization upon activation and downstream signaling pathway. They can be localized on the surface of the cell (TLR1, 2, 4, 6, 10) or in intracellular compartments (TLR3,7,8,9,11,12,13) [36]. In brief, TLR-signaling induces recruitment of specific adaptor molecules, leading to activation of the transcription factors, which stimulate production of inflammatory mediators, including cytokines and chemokines, such as TNF-α, IL-1α, IL-1β, IL-6, IL-8, IL-12, IL-23, macrophage inflammatory protein (MIP)-1α, and MIP-1β [37][38][39]. ...
Despite advances in antimicrobial and anti-inflammatory therapies, inflammation and its consequences still remain a significant problem in medicine. Acute inflammatory responses are responsible for directly life-threating conditions such as septic shock; on the other hand, chronic inflammation can cause degeneration of body tissues leading to severe impairment of their function. Neuroinflammation is defined as an inflammatory response in the central nervous system involving microglia, astrocytes, and cytokines including chemokines. It is considered an important cause of neurodegerative diseases, such as Alzheimer’s disease, Parkinson’s disease and amyotrophic lateral sclerosis. Lipopolysaccharide (LPS) is a strong immunogenic particle present in the outer membrane of Gram-negative bacteria. It is a major triggering factor for the inflammatory cascade in response to a Gram-negative bacteria infection. The use of LPS as a strong pro-inflammatory agent is a well-known model of inflammation applied in both in vivo and in vitro studies. This review offers a summary of the pathogenesis associated with LPS exposure, especially in the field of neuroinflammation. Moreover, we analyzed different in vivo LPS models utilized in the area of neuroscience. This paper presents recent knowledge and is focused on new insights in the LPS experimental model.
Sepsis-associated encephalopathy (SAE) is a critical neurological complication of sepsis and represents a crucial factor contributing to high mortality and adverse prognosis in septic patients. This study explored the contribution of NAT10-mediated messenger RNA (mRNA) acetylation in cognitive dysfunction associated with SAE, utilizing a cecal ligation and puncture (CLP)-induced SAE mouse model. Our findings demonstrate that CLP significantly upregulates NAT10 expression and mRNA acetylation in the excitatory neurons of the hippocampal dentate gyrus (DG). Notably, neuronal-specific Nat10 knockdown improved cognitive function in septic mice, highlighting its critical role in SAE. Proteomic analysis, RNA immunoprecipitation, and real-time qPCR identified GABA B R1 as a key downstream target of NAT10. Nat10 deletion reduced GABA B R1 expression, and subsequently weakened inhibitory postsynaptic currents in hippocampal DG neurons. Further analysis revealed that microglia activation and the release of inflammatory mediators lead to the increased NAT10 expression in neurons. Microglia depletion with PLX3397 effectively reduced NAT10 and GABA B R1 expression in neurons, and ameliorated cognitive dysfunction induced by SAE. In summary, our findings revealed that after CLP, NAT10 in hippocampal DG neurons promotes GABA B R1 expression through mRNA acetylation, leading to cognitive dysfunction.
Acute ischemic stroke (AIS) remains a major cause of mortality and long-term disability worldwide, driven by complex and multifaceted etiological factors. Metabolic dysregulation, gastrointestinal microbiome alterations, and systemic inflammation are emerging as significant contributors to AIS pathogenesis. This review addresses the critical need to understand how these factors interact to influence AIS risk and outcomes. We aim to elucidate the roles of dysregulated adipokines in obesity, the impact of gut microbiota disruptions, and the neuroinflammatory cascade initiated by lipopolysaccharides (LPS) in AIS. Dysregulated adipokines in obesity exacerbate inflammatory responses, increasing AIS risk and severity. Disruptions in the gut microbiota and subsequent LPS-induced neuroinflammation further link systemic inflammation to AIS. Advances in neuroimaging and biomarker development have improved diagnostic precision. Here, we highlight the need for a multifaceted approach to AIS management, integrating metabolic, microbiota, and inflammatory insights. Potential therapeutic strategies targeting these pathways could significantly improve AIS prevention and treatment. Future research should focus on further elucidating these pathways and developing targeted interventions to mitigate the impacts of metabolic dysregulation, microbiome imbalances, and inflammation on AIS.
Despite advances in antimicrobial and anti-inflammatory treatment, inflammation and its consequences remain a major challenge in the field of medicine. Inflammatory reactions can lead to life-threatening conditions such as septic shock, while chronic inflammation has the potential to worsen the condition of body tissues and ultimately lead to significant impairment of their functionality. Although the central nervous system has long been considered immune privileged to peripheral immune responses, recent research has shown that strong immune responses in the periphery also affect the brain, leading to reactive microglia, which belong to the innate immune system and reside in the brain, and neuroinflammation. The inflammatory response is primarily a protective mechanism to defend against pathogens and tissue damage. However, excessive and chronic inflammation can have negative effects on neuronal structure and function. Neuroinflammation underlies the pathogenesis of many neurological and neurodegenerative diseases and can accelerate their progression. Consequently, targeting inflammatory signaling pathways offers potential therapeutic strategies for various neuropathological conditions, particularly Parkinson’s and Alzheimer’s disease, by curbing inflammation. Here the blood-brain barrier is a major barrier for potential therapeutic strategies, therefore it would be highly advantageous to foster and utilize brain innate anti-inflammatory mechanisms. The tricarboxylic acid cycle-derived metabolite itaconate is highly upregulated in activated macrophages and has been shown to act as an immunomodulator with anti-inflammatory and antimicrobial functions. Mesaconate, an isomer of itaconate, similarly reduces the inflammatory response in macrophages. Nevertheless, most studies have focused on its esterified forms and its peripheral effects, while its influence on the CNS remained largely unexplored. Therefore, this study investigated the immunomodulatory and therapeutic potential of endogenously synthesized itaconate and its isomer mesaconate in lipopolysaccharide (LPS)-induced neuroinflammatory processes. Our results show that both itaconate and mesaconate reduce LPS-induced neuroinflammation, as evidenced by lower levels of inflammatory mediators, reduced microglial reactivity and a rescue of synaptic plasticity, the cellular correlate of learning and memory processes in the brain. Overall, this study emphasizes that both itaconate and mesaconate have therapeutic potential for neuroinflammatory processes in the brain and are of remarkable importance due to their endogenous origin and production, which usually leads to high tolerance.
Липополисахариды грамотрицательных бактерий (LPS) считаются наиболее иммуногенными бактериальными компонентами, ассоциированными с развитием системного воспаления и сепсиса. В обзоре описываются механизмы действия LPS на основные органы-мишени,- легкие, сердце и головной мозг тяжелое поражение которых ассоциировано с развитием дыхательной и сердечной недостаточности.
Gram-negative bacterial lipopolysaccharides (LPS) are considered to be the most immunogenic bacterial components associated with the development of systemic inflammation and sepsis. The review describes the mechanisms of action of LPS on the main target organs - the lungs, heart and brain, the severe damage of which is associated with the development of respiratory and heart failure.
Sepsis-associated encephalopathy (SAE) is a cognitive impairment associated with sepsis that occurs in the absence of direct infection in the central nervous system or structural brain damage. Microglia are thought to be macrophages of the central nervous system, devouring bits of neuronal cells and dead cells in the brain. They are activated in various ways, and microglia-mediated neuroinflammation is characteristic of central nervous system diseases, including SAE. Here, we systematically described the pathogenesis of SAE and demonstrated that microglia are closely related to the occurrence and development of SAE. Furthermore, we comprehensively discussed the function and phenotype of microglia and summarized their activation mechanism and role in SAE pathogenesis. Finally, this review summarizes recent studies on treating cognitive impairment in SAE by blocking microglial activation and toxic factors produced after activation. We suggest that targeting microglial activation may be a putative treatment for SAE.
Sepsis is associated with multiple organ dysfunction, and the brain is particularly vulnerable. Sepsis-associated encephalopathy (SAE) increases the mortality of patients with sepsis; however, the pathogenesis of SAE remains unclear. Methane, the simplest aliphatic hydrocarbon, has been reported to have anti-inflammatory and organ-protective effects. This study aimed to investigate the effects of methane on the cognitive deficits in mice with experimental sepsis. We randomly divided C57BL/6 male mice into sham, cecal ligation and puncture (CLP), and CLP + methane-rich saline (MS) groups. Twenty-four hours after surgery, behavioral tests were conducted on surviving mice and the hippocampus were collected for biochemical analysis. We found that CLP resulted in cognitive deficits in septic mice. A physiological mechanistic investigation revealed that microglia in the hippocampus are largely activated, coupled with the production of inflammatory cytokines and reactive oxygen species (ROS). Notably, methane inhibited the activation of microglia in the hippocampus, reduced the severity of inflammation, diminished the generation of ROS, and ultimately alleviated behavioral impairment in septic mice. Together, these show that treatment with methane ameliorated cognitive deficits in septic mice, which is partly related to the anti-inflammatory and antioxidative effects in the hippocampus.
Activation of TLRs mediated the NF-κB signaling pathway plays an important pathophysiological role in cardiac hypertrophy. Triad3A, a ubiquitin E3 ligase, has been reported to negatively regulate NF-κB activation pathway via promoting ubiquitination and degradation of TLR4 and TLR9 in innate immune cells. The role of Triad3A in cardiac hypertrophic development remains unknown. The present study investigated whether there is a link between Triad3A and TLR4 and TLR9 in pressure overload induced cardiac hypertrophy. We observed that Triad3A levels were markedly reduced following transverse aortic constriction (TAC) induced cardiac hypertrophy. Similarly, stimulation of neonatal rat cardiac myocytes (NRCMs) with angiotensin-II (Ang II) significantly decreased Triad3A expression. To determine the role of Triad3A in TAC-induced cardiac hypertrophy, we transduced the myocardium with adenovirus expressing Triad3A followed by induction of TAC. We observed that increased expression of Triad3A significantly attenuated cardiac hypertrophy and improved cardiac function. To investigate the mechanisms by which Triad3A attenuated cardiac hypertrophy, we examined the Triad3A E3 ubiquitination on TLR4 and TLR9. We found that Triad3A promoted TLR4 and TLR9 degradation through ubiquitination. Triad3A mediated TLR4 and TLR9 degradation resulted in suppression of NF-κB activation. Our data suggest that Triad3A plays a protective role in the development of cardiac hypertrophy, at least through catalyzing ubiquitination-mediated degradation of TLR4 and TLR9, thus negatively regulating NF-κB activation.
Glial activation and neuroinflammation play significant roles in apoptosis as well as in the development of cognitive and memory deficits. Neuroinflammation is also a critical feature in the pathogenesis of neurodegenerative disorders such as Alzheimer and Parkinson's diseases. Previously, hesperetin has been shown to be an effective antioxidant and anti-inflammatory agent. In the present study, in vivo and in vitro analyses were performed to evaluate the neuroprotective effects of hesperetin in lipopolysaccharide (LPS)-induced neuroinflammation, oxidative stress, neuronal apoptosis and memory impairments. Based on our findings, LPS treatment resulted in microglial activation and astrocytosis and elevated the expression of inflammatory mediators such as phosphorylated-Nuclear factor-κB (p-NF-κB), tumor necrosis factor-α (TNF-α), and interleukin-1β (IL-1β) in the cortical and hippocampal regions and in BV2 cells. However, hesperetin cotreatment markedly reduced the expression of inflammatory cytokines by ameliorating Toll-like receptor-4 (TLR4)-mediated ionized calcium-binding adapter molecule 1/glial fibrillary acidic protein (Iba-1/GFAP) expression. Similarly, hesperetin attenuated LPS-induced generation of reactive oxygen species/lipid per oxidation (ROS/LPO) and improved the antioxidant protein level such as nuclear factor erythroid 2-related factor 2 (Nrf2) and Haem-oxygenase (HO-1) in the mouse brain. Additionally, hesperetin ameliorated cytotoxicity and ROS/LPO induced by LPS in HT-22 cells. Moreover, hesperetin rescued LPS-induced neuronal apoptosis by reducing the expression of phosphorylated-c-Jun N-terminal kinases (p-JNK), B-cell lymphoma 2 (Bcl-2)-associated X protein (Bax), and Caspase-3 protein and promoting the Bcl-2 protein level. Furthermore, hesperetin enhanced synaptic integrity, cognition, and memory processes by enhancing the phosphorylated-cAMP response element binding protein (p-CREB), postsynaptic density protein-95 (PSD-95), and Syntaxin. Overall, our preclinical study suggests that hesperetin conferred neuroprotection by regulating the TLR4/NF-κB signaling pathway against the detrimental effects of LPS.
Inflammation in central nervous system (CNS) plays a vital role in neurodegenerative diseases such as Alzheimer's disease (AD), Parkinson's disease (PD), Lewy body dementia (DLB), HIV-related dementia and traumatic brain injury. Icariside II (ICS II), an active flavonoid compound derived from a Chinese herbal medicine Epimedium brevicornum Maxim, has been shown to possess a neuroprotective effect on AD model. However, whether ICS II has a directly protective effect on acute neuroinflammation remains still unclear. Therefore, the current study was designed to investigate the possible protective effect of ICS II on acute neuroinflammation induced by intracerebroventricular (ICV) injection of lipopolysaccharide (LPS), and further to explore its possible mechanism. After ICS II was prophylactically administered for 7 days before LPS injection, the rats were randomly divided into five groups as follows: sham group (n = 9), sham + ICS II-H (10 mg/kg) (n = 9), LPS (n = 14), LPS + ICS II-L (3 mg/kg) (n = 14), LPS + ICS II-H (10 mg/kg) (n = 14) groups, respectively. As expected, LPS injection exhibited neuronal morphological damage, and ionized calcium binding adapter molecule 1 (IBA-1) of microglia and glial fibrillary acidic protein (GFAP) of astrocyte were activated. However, pre-treatment with ICS II not only inhibited the activation of microglia and astrocyte, but also significantly reversed the expressions of inflammatory factors such as interleukin-1β (IL-1β), tumor necrosis factor (TNF-α), cyclooxygenase-2 (COX-2), as well as the expressions of Toll-Like receptor 4 (TLR4), myeloid differentiation factor 88 (MyD88) and TNF receptor associated factor 6 (TRAF6). Furthermore, ICS II inhibited the degradation of IκB and the following activation of NF-κB. Hence it is concluded that ICS II attenuates LPS-induced neuroinflammation through inhibiting TLR4/MyD88/NF-κB pathway in rats, and it has potential value as a new therapeutic agent to treat neuroinflammation-related diseases, such as AD.
Background:
Pharmaceutical treatment with probable anti-inflammatory substances that attack cells in various ways including receptors, ion channels, or transporter systems may slow down the progression of inflammatory conditions. Astrocytes and microglia are the most prominent target cells for inflammation in the central nervous system. Their responses upon inflammatory stimuli work through the NO/cyclic GMP/protein kinase G systems that can downregulate the ATP-induced Ca2+ signaling, as well as G protein activities which alter Na+ transporters including Na+/K+-ATPase pump activity, Toll-like receptor 4 (TLR4), glutamate-induced Ca2+ signaling, and release of pro-inflammatory cytokines. The rationale for this project was to investigate a combination of pharmaceutical substances influencing the NO and the Gi/Gs activations of inflammatory reactive cells in order to make the cells return into a more physiological state. The ATP-evoked Ca2+ signaling is important maybe due to increased ATP release and subsequent activation of purinergic receptors. A balance between intercellular Ca2+ signaling through gap junctions and extracellular signaling mediated by extracellular ATP may be important for physiological function.
Methods:
Astrocytes in primary cultures were incubated with lipopolysaccharide in a physiological glucose concentration for 24 h to induce inflammatory reactivity. The probable anti-inflammatory substances sildenafil and 1α,25-Dihydroxyvitamin D3 together with endomorphin-1, naloxone, and levetiracetam, were used in the presence of high glucose concentration in the medium to restore the cells. Glutamate-, 5-HT-, and ATP-evoked intracellular Ca2+ release, Na+/K+-ATPase expression, expression of inflammatory receptors, and release of tumor necrosis factor alpha were measured.
Results:
Sildenafil in ultralow concentration together with 1α,25-Dihydroxyvitamin D3 showed most prominent effects on the ATP-evoked intracellular Ca2+ release. The μ-opioid agonist endomorphin-1, the μ-opioid antagonist naloxone in ultralow concentration, and the antiepileptic agent levetiracetam downregulated the glutamate-evoked intracellular Ca2+ release and TLR4. The combination of the pharmaceutical substances in high glucose concentration downregulated the glutamate- and ATP-evoked Ca2+ signaling and the TLR4 expression and upregulated the Na+/K+-ATPase pump.
Conclusion:
Pharmaceutical treatment with the combination of substances that have potential anti-inflammatory effects, which attack different biochemical mechanisms in the cells may exert decisive effects to downregulate neuroinflammation in the nervous system.
Background: This study aimed at assessing the effect of a low-fat diet (LFD) in obese mice lacking toll–like receptors (Tlr) and understanding the expression and regulation of microRNAs during weight reduction. Methods: C57BL/6, Tlr5−/−, Tlr2−/− and Tlr4−/− mice were used in this study. A group of mice were fed with a high-fat diet (HFD) (58% kcal) for 12 weeks to induce obesity (diet-induced obesity, DIO). Another group that had been fed with HFD for eight weeks (obese mice) were switched to a low-fat diet (LFD) (10.5% kcal) for the next four weeks to reduce their body weight. The control mice were fed with a standard AIN-76A diet for the entire 12 weeks. The body weight of the mice was measured weekly. At the end of the experiment, epididymal fat weight and adipocyte size were measured. The differentially expressed miRNAs in the fat tissue was determined by next-generation sequencing with real-time quantitative reverse transcription polymerase chain reaction (RT–qPCR). Target prediction and functional annotation of miRNAs were performed using miRSystem database. Results: Switching to LFD significantly reduced the body weight and epididymal fat mass in the HFD-fed C57BL/6 and Tlr5−/− mice but not in Tlr2−/− and Tlr4−/− mice. Weight reduction significantly decreased the size of adipocytes in C57BL/6 but not in the Tlr knockout mice. In Tlr2−/− and Tlr4−/− mice, feeding with HFD and the subsequent weight reduction resulted in an aberrant miRNA expression in the epididymal fat tissue unlike in C57BL/6 and Tlr5−/−. However, target prediction and functional annotation by miRSystem database revealed that all the top 10 Kyoto Encyclopedia of Genes and Genomes (KEGG) database pathways of the dysregulated miRNAs during weight reduction in the C57BL/6 mice were also found in the regulated pathways of Tlr5−/−, Tlr2−/−, and Tlr4−/− strains. However, among these pathways, gene sets involved in arginine and proline metabolism and glutathione metabolism were mainly involved in the Tlr knockout mice but not in the C57BL/6 mice. Conclusions: In this study, we demonstrated that feeding of LFD leads to significant body weight reduction in C57BL/6 and Tlr5−/− mice, but not in Tlr2−/− and Tlr4−/− mice. Significant reduction in the size of adipocytes of epididymal fat was only found in C57BL/6, but not in Tlr5−/−, Tlr2−/−, and Tlr4−/− mice. The dysregulated miRNAs in Tlr2−/− and Tlr4−/− mice were different from those in C57BL/6 and Tlr5−/− strains. Among those miRNA-regulated pathways, arginine and proline metabolism as well as glutathione metabolism may have important roles in the Tlr knockout mice rather than in C57BL/6 mice.
Background/aims:
the pathogenesis of sepsis-associated encephalopathy (SAE) is multifactorial, involving neurotransmitter alterations, inflammatory cytokines, oxidative damage, mitochondrial dysfunction, apoptosis, and other factors. Mitochondria are major producers of reactive oxygen species, resulting in cellular injury. Omi/HtrA2 is a proapoptotic mitochondrial serine protease involved in caspase-dependent cell death; it is translocated from mitochondria to the cytosol after an apoptotic insult. We previously found that UCF-101, a specific inhibitor of Omi/HtrA2, has neuroprotective effects on cerebral oxidative injury and cognitive impairment in septic rats. In this study, the mechanisms and molecular pathways underlying these effects were investigated.
Methods:
Male Sprague-Dawley rats were subjected to cecal ligation and puncture (CLP) or sham-operated laparotomy and were administered vehicle or UCF-101 (10 µmol/kg). The hippocampus was isolated for subsequent analysis. Omi/HtrA2 expression in the mitochondria or cytosol was evaluated by immunofluorescence or western blotting. Terminal deoxynucleotidyl transferase dUTP nick end labeling staining was utilized to evaluate levels of apoptosis, and western blotting was used to evaluate apoptosis-related proteins, such as cleaved caspase-3, caspase-9, and poly (ADP-ribose) polymerase (PARP). Tight junction expression was assessed by immunofluorescence and western blotting. Mitochondrial function, inflammatory cytokines, and oxidative stress were also assayed. In addition, a wet/dry method was used to evaluate brain edema and Evans blue extravasation was used to evaluate blood-brain barrier (BBB) integrity.
Results:
After CLP treatment, the hippocampus exhibited a mild increase in Omi/HtrA2 expression; cytosolic Omi/HtrA2 expression increased significantly, whereas mitochondrial Omi/HtrA2 expression was reduced, indicating that CLP-induced oxidative stress resulted in the translocation of Omi/HtrA2 from mitochondria to the cytosol. Hippocampal cleaved caspase-3, caspase-9, and PARP levels were significantly higher in animals treated with CLP than in sham-operated animals, while XIAP expression was lower. Treatment with UCF-101 prevented the mobilization of Omi/HtrA2 from mitochondria to the cytosol, attenuated XIAP degradation, and decreased cleaved caspase-3, caspase-9, and PARP expression as well as apoptosis. UCF-101 also reversed the decreased mitochondrial complex I, II, and III respiration and the reduced ATP caused by CLP. In addition, UCF-101 treatment resulted in a significant improvement in BBB integrity, as demonstrated by increased occludin, claudin-5, and zonula occludens 1 levels and reduced Evans blue extravasation. No significant effects of UCF-101 on brain edema were found. Inflammatory cytokines and oxidative stress were significantly higher in the CLP-treated group than in the sham-operated group. However, the inhibition of Omi/HtrA2 by UCF-101 significantly alleviated these responses.
Conclusion:
Our data indicated that Omi/ HtrA2 regulates a mitochondria-dependent apoptotic pathway in a murine model of septic encephalopathy. Inhibition of Omi/HtrA2 by UCF-101 leads to neuroprotection by inhibiting the cytosolic translocation of Omi/HtrA2 and antagonizing the caspase-dependent apoptosis pathway. Therapeutic interventions that inhibit Omi/HtrA2 translocation or protease activity may provide a novel method to treat SAE.
Sepsis is a systemic inflammatory reaction caused by infection. Multiple organ failure ultimately leads to high morbidity and mortality. Unfortunately, therapies against these responses have been unsuccessful due to the insufficient underlying pathophysiological evidence. Protein interacting with C-kinase 1 (PICK1) has received considerable attention because of its important physiological functions in many tissues. However, its role in sepsis-induced acute lung injury (ALI) is unclear. In this study, we used cecal ligation and puncture (CLP) to establish a septic model and found that decreased microtubule-associated protein-1light chain 3 (LC3)-II/LC3-I in PICK1 −/− septic mice was caused by autophagy dysfunction. Consistently, the transmission electron microscopy (TEM) of bone marrow-derived macrophages (BMDMs) from PICK1 −/− mice showed the accumulation of autophagosomes as well. However, more serious damage was caused by PICK1 deficiency indicating that the disrupted autophagic flux was harmful to sepsis-induced ALI. We also observed that it was the impaired lysosomal function that mediated autophagic flux blockade, and the autophagy progress was relevant to PI3K-Akt-mTOR pathway. These findings will aid in the potential development of PICK1 with novel evidence of autophagy in sepsis treatment and prevention.
The anti-inflammatory effect of ginsenoside Rh2 (GRh2) has labeled it as one of the most important ginsenosides. The purpose of this study was to identify the anti-inflammatory and antioxidant effects of GRh2 using a lipopolysaccharide (LPS) challenge lung-injury animal model. GRh2 reduced LPS-induced proinflammatory mediator nitric oxide (NO), tumor necrosis factor-alpha, interleukin (IL)-1β, and anti-inflammatory cytokines (IL-4, IL-6, and IL-10) production in lung tissues. GRh2 treatment decreased the histological alterations in the lung tissues and bronchoalveolar lavage fluid (BALF) protein content; total cell number also reduced in LPS-induced lung injury in mice. Moreover, GRh2 blocked iNOS, COX-2, the phosphorylation of IκB-α, ERK, JNK, p38, Raf-1, and MEK protein expression, which corresponds with the growth of HO-1, Nrf-2, catalase, SOD, and GPx expression in LPS-induced lung injury. An in vivo experimental study suggested that GRh2 has anti-inflammatory effects, and has potential therapeutic efficacy in major anterior segment lung diseases.
Significance
Parkinson’s disease (PD) is the second most common neurodegenerative disorder. It is characterized by progressive deterioration of motor function caused by loss of dopamine-producing neurons. Currently, there is no treatment that could stop the progress of the disease. Loss-of-function mutations in Parkin account for ∼50% of early onset PD. Parkin functions as an E3 ubiquitin ligase that exhibits multiple protective roles especially in dopaminergic neurons. Here, we demonstrate that PICK1 directly binds to Parkin. PICK1 is a potent endogenous inhibitor of Parkin’s E3 ubiquitin ligase activity and blocks Parkin’s protective functions. Conversely, knockout of PICK1 enhances the neuroprotective effect of Parkin. Thus, the reduction of PICK1 may provide a therapeutic target for the treatment of PD.
Sepsis is not only a significant cause of mortality worldwide but has particularly devastating effects on the central nervous system of survivors. It is therefore crucial to understand the molecular structure, physiology, and events involved in the pathogenesis of sepsis-associated encephalopathy, so that potential therapeutic advances can be achieved. A key determinant to the development of this type of encephalopathy is morphological and functional modification of the blood–brain barrier (BBB), whose function is to protect the CNS from pathogens and toxic threats. Key mediators of pathologic sequelae of sepsis in the brain include cytokines, including TNF-α, and sphingolipids, which are biologically active components of cellular membranes that possess diverse functions. Emerging data demonstrated an essential role for sphingolipids in the pulmonary vascular endothelium. This raises the question of whether endothelial stability in other organs systems such as the CNS may also be mediated by sphingolipids and their receptors. In this review, we will model the structure and vulnerability of the BBB and hypothesize mechanisms for therapeutic stabilization and repair following a confrontation with sepsis-induced inflammation.
Toll-like receptor 4 (TLR4) signaling in the brain mediates autoimmune responses and induces neuroinflammation that results in neurodegenerative diseases, such as Alzheimer's disease (AD). The plant hormone osmotin inhibited lipopolysaccharide (LPS)-induced TLR4 downstream signaling, including activation of TLR4, CD14, IKKα/β, and NFκB, and the release of inflammatory mediators, such as COX-2, TNF-α, iNOS, and IL-1β. Immunoprecipitation demonstrated colocalization of TLR4 and AdipoR1 receptors in BV2 microglial cells, which suggests that osmotin binds to AdipoR1 and inhibits downstream TLR4 signaling. Furthermore, osmotin treatment reversed LPS-induced behavioral and memory disturbances and attenuated LPS-induced increases in the expression of AD markers, such as Aβ, APP, BACE-1, and p-Tau. Osmotin improved synaptic functionality via enhancing the activity of pre- and post-synaptic markers, like PSD-95, SNAP-25, and syntaxin-1. Osmotin also prevented LPS-induced apoptotic neurodegeneration via inhibition of PARP-1 and caspase-3. Overall, our studies demonstrated that osmotin prevented neuroinflammation-associated memory impairment and neurodegeneration and suggest AdipoR1 as a therapeutic target for the treatment of neuroinflammation and neurological disorders, such as AD.
Protein kinase C (PKC) plays a central role in the control of proliferation and differentiation of a wide range of cell types by mediating the signal transduction response to hormones and growth factors. Upon activation by diacylglycerol, PKC translocates to different subcellular sites where it phosphorylates numerous proteins, most of which are unidentified. We used the yeast two-hybrid system to identify proteins that interact with activated PKC alpha. Using the catalytic region of PKC fused to the DNA binding domain of yeast GAL4 as "bait" to screen a mouse T cell cDNA library in which cDNA was fused to the GAL4 activation domain, we cloned several novel proteins that interact with C-kinase (PICKs). One of these proteins, designated PICK1, interacts specifically with the catalytic domain of PKC and is an efficient substrate for phosphorylation by PKC in vitro and in vivo. PICK1 is localized to the perinuclear region and is phosphorylated in response to PKC activation. PICK1 and other PICKs may play important roles in mediating the actions of PKC.
Therapeutic strategies designed to inhibit the activation of microglia may lead to significant advancement in the treatment of most neurodegenerative diseases. Pyrroloquinoline quinone (PQQ) is a naturally occurring redox cofactor that acts as an essential nutrient, antioxidant, and has been reported to exert potent immunosuppressive effects. In the present study, the anti-inflammatory effects of PQQ was investigated in LPS treated primary microglia cells. Our observations showed that pretreatment with PQQ significantly inhibited the production of NO and PGE2 and suppressed the expression of pro-inflammatory mediators such as iNOS, COX-2, TNF-a, IL-1b, IL-6, MCP-1 and MIP-1a in LPS treated primary microglia cells. The nuclear translocation of NF-κB and the phosphorylation level of p65, p38 and JNK MAP kinase pathways were also inhibited by PQQ in LPS stimulated primary microglia cells. Further a systemic LPS treatment acute inflammation murine brain model was used to study the suppressive effects of PQQ against neuroinflammation in vivo. Mice treated with PQQ demonstrated marked attenuation of neuroinflammation based on Western blotting and immunohistochemistry analysis of Iba1-against antibody in the brain tissue. Indicated that PQQ protected primary cortical neurons against microglia-mediated neurotoxicity. These results collectively suggested that PQQ might be a promising therapeutic agent for alleviating the progress of neurodegenerative diseases associated with microglia activation.
Neuroinflammation is largely described in the acute phase after brain injury, as well as in chronic brain diseases. Cells that are directly or indirectly involved in immune responses compose the Central Nervous System. Microglia are resident cells of the central nervous system and, as peripheral macrophages, are activated in the presence of some cellular insult, producing a large number of cytokines and chemokines in order to remove toxins from the extracellular space. This activation can lead to a breakdown of the blood-brain barrier, production of reactive oxygen species that is involved in the progression of brain damage as occurs in septic encephalopathy. Given the growing importance of microglia in the field of neurotoxicology, we describe the role of microglia and the cellular mechanisms that activate these cells during sepsis. Thus, in this review we focused in the relationship between microglia and neuroinflammation associated with sepsis.
Abnormal aggregation of α-synuclein and sustained microglial activation are important contributors to the pathogenic processes of Parkinson's disease. However, the relationship between disease-associated protein aggregation and microglia-mediated neuroinflammation remains unknown. Here, using a combination of in silico, in vitro and in vivo approaches, we show that extracellular α-synuclein released from neuronal cells is an endogenous agonist for Toll-like receptor 2 (TLR2), which activates inflammatory responses in microglia. The TLR2 ligand activity of α-synuclein is conformation-sensitive; only specific types of oligomer can interact with and activate TLR2. This paracrine interaction between neuron-released oligomeric α-synuclein and TLR2 in microglia suggests that both of these proteins are novel therapeutic targets for modification of neuroinflammation in Parkinson's disease and related neurological diseases.
B cells are regarded for their capacity to produce antibody. However, recent advances in B cell biology have capitalized on old findings and demonstrated that B cells also release a broad variety of cytokines. As with T helper cells, B cells can be classified into subsets according to the cytokine milieu that they produce. One functional B cell subset, regulatory B cells (Bregs), has recently been shown to contribute to the maintenance of the fine equilibrium required for tolerance. Bregs restrain the excessive inflammatory responses that occur during autoimmune diseases or that can be caused by unresolved infections. Pivotal to Breg function is interleukin-10 (IL-10), which inhibits proinflammatory cytokines and supports regulatory T cell differentiation. This review reports and discusses the factors that are important for Breg differentiation and for their effector function in both mouse and human.
Microglia are the brain's tissue macrophages and are found in an activated state surrounding beta-amyloid plaques in the Alzheimer's disease brain. Microglia interact with fibrillar beta-amyloid (fAbeta) through an ensemble of surface receptors composed of the alpha(6)beta(1) integrin, CD36, CD47, and the class A scavenger receptor. These receptors act in concert to initiate intracellular signaling cascades and phenotypic activation of these cells. However, it is unclear how engagement of this receptor complex is linked to the induction of an activated microglial phenotype. We report that the response of microglial cells to fibrillar forms of Abeta requires the participation of Toll-like receptors (TLRs) and the coreceptor CD14. The response of microglia to fAbeta is reliant upon CD14, which act together with TLR4 and TLR2 to bind fAbeta and to activate intracellular signaling. We find that cells lacking these receptors could not initiate a Src-Vav-Rac signaling cascade leading to reactive oxygen species production and phagocytosis. The fAbeta-mediated activation of p38 MAPK also required CD14, TLR4, and TLR2. Inhibition of p38 abrogated fAbeta-induced reactive oxygen species production and attenuated the induction of phagocytosis. Microglia lacking CD14, TLR4, and TLR2 showed no induction of phosphorylated IkappaBalpha following fAbeta. These data indicate these innate immune receptors function as members of the microglial fAbeta receptor complex and identify the signaling mechanisms whereby they contribute to microglial activation.
Synaptic clustering of neurotransmitter receptors is crucial for efficient signal transduction and integration in neurons. PDZ domain-containing proteins such as PSD-95/SAP90 interact with the intracellular C termini of a variety of receptors and are thought to be important in the targeting and anchoring of receptors to specific synapses. Here, we show that PICK1 (protein interacting with C kinase), a PDZ domain-containing protein, interacts with the C termini of alpha-amino-3-hydroxy-5-methyl-isoxazole-4-propionic acid (AMPA) receptors in vitro and in vivo. In neurons, PICK1 specifically colocalizes with AMPA receptors at excitatory synapses. Furthermore, PICK1 induces clustering of AMPA receptors in heterologous expression systems. These results suggest that PICK1 may play an important role in the modulation of synaptic transmission by regulating the synaptic targeting of AMPA receptors.
Sepsis-associated encephalopathy (SAE) manifested clinically in acute and long-term cognitive impairments and associated with increased morbidity and mortality worldwide. The potential pathological changes of SAE are complex and remain to be elucidated. Pyroptosis, a novel programmed cell death, is executed by caspase-1-cleaved GSDMD N-terminal (GSDMD-NT) and we investigated it in peripheral blood immunocytes of septic patients previously. Here, a caspase-1 inhibitor VX765 was treated with CLP-induced septic mice. Novel object recognition test indicated that VX765 treatment reversed cognitive dysfunction in septic mice. Elevated plus maze, tail suspension test and open field test revealed that depressive-like behaviors of septic mice were relieved. Inhibited caspase-1 suppressed the expressions of GSDMD and its cleavage form GSDMD-NT, and reduced pyroptosis in brain at day 1 and day 7 after sepsis. Meantime, inhibited caspase-1 mitigated the expressions of IL-1β, MCP-1 and TNF-α in serum and brain, diminished microglia activation in septic mice, and reduced sepsis-induced brain-blood barrier disruption and ultrastructure damages in brain as well. Inhibited caspase-1 protected the synapse plasticity and preserved long-term potential, which may be the possible mechanism of cognitive functions protective effects of septic mice. In conclusion, caspase-1 inhibition exerts brain-protective effects against SAE and cognitive impairments in a mouse model of sepsis.
Delavatine A, an unusual isoquinoline alkaloid isolated from I. delavayi, was first studied for anti-inflammatory effect using lipopolysaccharide (LPS)-induced BV-2 microglia. In the present study, we found that delavatine A substantially suppressed the LPS-induced pro-inflammatory mediators, nitric oxide (NO), and tumor necrosis factor-a (TNF-a), interleukin-6 (IL-6), interleukin-1β (IL-1β) in BV-2 microglial cells. These effects resulted from the inhibition of their regulatory genes inducible NO synthase (iNOS), cycloxygenase-2 (COX-2) and TNF-a, IL-6, IL-1β. In addition, we examined several pathways related to inflammation. The results revealed that delavatine A significantly decreased LPS-induced the activation of nuclear factor-κB (NF-κB) by suppressing the p65 subunits, and the phosphorylation of IκBα, while not related to PI3K/Akt or MAPK pathways.
The breakdown of the blood-brain barrier (BBB) is a key event in the development of sepsis-induced brain damage. BBB opening allows blood-born immune cells to enter the CNS to provoke a neuroinflammatory response. Abnormal expression and activation of matrix metalloproteinases (MMP) was shown to contribute to BBB opening. Using different mouse genotypes in a model of LPS-induced systemic inflammation, our present report reveals phosphoinositide 3-kinase γ (PI3Kγ) as a mediator of BBB deterioration and concomitant generation of MMP by microglia. Unexpectedly, microglia expressing lipid kinase-deficient mutant PI3Kγ exhibited similar MMP regulation as wild-type cells. Our data suggest kinase-independent control of cAMP phosphodiesterase activity by PI3Kγ as a crucial mediator of microglial cell activation, MMP expression and subsequent BBB deterioration. The results identify the suppressive effect of PI3Kγ on cAMP as a critical mediator of immune cell functions.
Oxidative stress and inflammation is likely to be a major step in the development of sepsis-associated encephalopathy (SAE) and long-term cognitive impairment. To date, it is not known whether brain inflammation and oxidative damage are a direct consequence of systemic inflammation or whether these events are driven by brain resident cells, such as microglia. Therefore, the aim of this study is to evaluate the effect of minocycline on behavioral and neuroinflammatory parameters in rats submitted to sepsis. Male Wistar rats were subjected to sepsis by cecal ligation and puncture (CLP). The animals were divided into sham-operated (Sham+control), sham-operated plus minocycline (sham+MIN), CLP (CLP+control) and CLP plus minocycline (CLP+MIN) (100 μg/kg, administered as a single intracerebroventricular (ICV) injection). Some animals were killed 24 h after surgery to assess the breakdown of the blood brain barrier, cytokine levels, oxidative damage to lipids (TBARS) and proteins in the hippocampus. Some animals were allowed to recover for 10 days when step-down inhibitory avoidance and open-field tasks were performed. Treatment with minocycline prevented an increase in markers of oxidative damage and inflammation in the hippocampus after sepsis. This was associated with an improvement in long-term cognitive performance. In conclusion, we demonstrated that the inhibition of the microglia by an ICV injection of minocycline was able to decrease acute brain oxidative damage and inflammation as well as long-term cognitive impairment in sepsis survivors.
The transport of Toll-like Receptors (TLRs) to various organelles has emerged as an essential means by which innate immunity is regulated. While most of our knowledge is restricted to regulators that promote the transport of newly synthesized receptors, the regulators that control TLR transport after microbial detection remain unknown. Here, we report that the plasma membrane localized Pattern Recognition Receptor (PRR) CD14 is required for the microbe-induced endocytosis of TLR4. In dendritic cells, this CD14-dependent endocytosis pathway is upregulated upon exposure to inflammatory mediators. We identify the tyrosine kinase Syk and its downstream effector PLCγ2 as important regulators of TLR4 endocytosis and signaling. These data establish that upon microbial detection, an upstream PRR (CD14) controls the trafficking and signaling functions of a downstream PRR (TLR4). This innate immune trafficking cascade illustrates how pathogen detection systems operate to induce both membrane transport and signal transduction.
The mechanisms or pathophysiology that leads to preterm brain damage including white matter damage during development are complex and not fully understood. Intrauterine infection/inflammation can significantly affect perinatal brain development and result in significant alterations in brain structure and function. Glial cells and Toll-like receptors (TLRs) are vital players in central nervous system immune response; dysregulation of this response plays an important role in brain damage. Intrauterine infection/inflammation has immunomodulatory effects and induces specific alterations in the TLRs response in many tissues. Recent findings indicate that intrauterine infection/inflammation could promote inflammatory processes in brain and in glial cells by up-regulating cytokines and inflammatory mediators, and by activating signaling pathways and transcriptional factors (nuclear factor-kappaB) implicated in inflammatory injury. TLRs may be involved in intrauterine infection-mediated inflammatory signaling, and intrauterine infection/inflammation could interfere with the TLR4 recruitment into the lipid rafts, leading to an effect on the TLR signaling transduction. In summary, current results suggest that TLRs are key mediators of intrauterine infection/inflammation induced preterm brain damage.
Septic encephalopathy is a frequent complication in severe sepsis, the pathogenesis and mechanisms of which are not fully understood. Here, we review recent advances in our understanding of septic encephalopathy, from molecular mechanisms to behavioral alterations, from diagnostic tools to potential therapeutic agents.
Recent insights into septic encephalopathy include: microcirculatory failure precedes changes in evoked potential responses; blood-brain barrier alteration is prevented by reducing intercellular adhesion molecule expression and pericyte detachment; reducing infiltration of CD68 macrophages and inhibiting complement activation alleviates neuroinflammation in septic encephalopathy; and reducing mitochondrial dysfunction and inducible nitric oxide synthase expression can restore altered brain function. In addition, other factors such as the circulating levels of growth hormone are independent predictors for mortality and correlate with the severity of sepsis. Similar to humans, septic rats present recognition memory impairment and depressive-like symptoms but not anxiety-like behavior and will serve as efficient models to study the underlying mechanisms of septic encephalopathy.
Septic encephalopathy is a dynamic disease caused by a complex network of systems and pathways going awry. More insights into the pathogenesis of septic encephalopathy are expected to lead to new cellular and molecular targets, which in turn will permit design of specific septic encephalopathy-alleviating drugs and prevent its negative influence on survival.
Patients with sepsis often manifest disorientation, somnolence, asterixis and coma, symptoms also seen in portasystemic encephalopathy. Altered plasma concentrations of the neutral amino acids and in creased blood-brain transport of these acids may play a role in portasystemic encephalopathy. Plasma amino acids and blood-brain barrier transport of neutral amino acids were investigated in a rat model of abdominal sepsis, cecal ligation and puncture. The blood-brain transport was studied by the technique of Oldendorf with carbon-14-amino acids 12 and 24 hours after the induction of sepsis. In similar groups of animals, isolation of brain capillaries was carried out by the technique of Hjelle and the capillaries were incubated with carbon-14-amino acids to study transport activity. Plasma and brain amino acids were deranged in a fashion similar to the derangements seen in portasystemic encephalopathy, with a decrease in plasma branched chain amino acids and an increase in most neutral amino acids in brain. The changes were most pronounced after 24 hours. The brain uptake of several neutral amino acids was increased in the septic rats, while the uptake of lysine, a basic amino acid, was normal. In the brain capillaries isolated from septic rats, tyrosine and leucine transport was also greater than in sham-operated animals. Elevated neutral amino acids may play a role in the encephalopathy encountered in septic patients similar to its role in patients with portasystemic encephalopathy, as similar mechanisms appear to be operating.
Septic encephalopathy is a complication of sepsis, and it is closely associated with the increased mortality of the sufferers. Pathophysiology of septic encephalopathy is not still completely understood. In an attempt to provide insight into the pathogenesis of septic encephalopathy, a light and electron microscopic investigation has been carried out in a rat model of intraperitoneal sepsis. Experimental fecal peritonitis was induced in Wistar rats which have been monitored for 6 h and sacrificed to harvest the samples of frontal cortex. Vital parameters and morphometric data obtained from investigation of the microvessels were then compared with the sham-operated and unoperated controls. In addition to the discernible drop in the blood pressure and in rectal temperature following initial increases, unstable but usually increased heart rate and marked respiratory failure were recorded. Estimation of the percentage of the microvessel area occupied by edema revealed the presence of significantly more perimicrovascular edema in the experimental fecal peritonitis group compared to both sham-operated and unoperated controls, while no significant difference was present between the latter two groups. Electron microscopic investigation confirmed the presence of distinctive perimicrovascular edema in the fecal peritonitis group although the endothelial cells were linked by tight junctions which appeared morphologically intact. Although it might be premature to draw any strict parallels between the septic encephalopathy in humans and the findings observed in the present model, the results may suggest that the edema observed around the microvessels would bare a role in the pathogenesis of the septic encephalopathy probably by affecting the exchange of oxygen and nutrients with carbon dioxide and waste products between the blood and brain parenchyma.
Toll-like receptors (TLRs) play a critical role in the immune system as sensors of microbial infection. Signaling downstream from TLRs is initiated by the recruitment of adaptor proteins, including MyD88 and TIRAP. These adaptors play essential roles in TLR signaling, but the mechanism of their function is currently unknown. Here we demonstrate that TIRAP and MyD88 have distinct functions and describe a mechanism of recruitment of TIRAP and MyD88 to TLR4. We find that TIRAP contains a phosphatidylinositol 4,5-bisphosphate (PIP2) binding domain, which mediates TIRAP recruitment to the plasma membrane. TIRAP then functions to facilitate MyD88 delivery to activated TLR4 to initiate signal transduction. These results establish that phosphoinositide-mediated adaptor recruitment initiates a specific signal-transduction pathway.
PICK1 is a peripheral membrane protein conserved from Caenorhabditis elegans to the human. It is expressed in many tissues with high levels in brain and testis. Inside cells, PICK1 is localized at the perinuclear region as well as specialized structures such as synapses of neurons. PICK1 contains a PDZ domain and a BAR domain. The PDZ domain of PICK1 binds to a large number of membrane proteins, especially proteins with C-terminal type II PDZ-binding motifs. The BAR domain of PICK1 binds to lipid molecules, mainly phosphoinositides. While the PDZ domain and the linker region of PICK1 enhance BAR domain's lipid binding, the C-terminal region of PICK1 inhibits its lipid binding. PICK1 regulates the subcellular localization and surface expression of its PDZ-binding partners. Lipid binding of PICK1's BAR domain is important for this regulation. With its PDZ domain interacting with membrane proteins and its BAR domain binding to lipids, the unique structure of PICK1 enables it to couple membrane proteins to protein-trafficking machinery.
TRAM couples endocytosis of Toll-like receptor 4 to the induction of interferon-beta
- Kagan
Islet-cell autoantigen 69 mediates the antihyperalgesic effects of electroacupuncture on inflammatory pain by regulating spinal glutamate receptor subunit 2 phosphorylation through protein interacting with C-kinase 1 in mice
- Han