Aberrant activation of hippocampal astrocytes causes neuroinflammation and cognitive decline in mice.

Reactive astrocytes are associated with neuroinflammation and cognitive decline in diverse neuropathologies; however, the underlying mechanisms are unclear. We used optogenetic and chemogenetic tools to identify the crucial roles of the hippocampal CA1 astrocytes in cognitive decline. Our results showed that repeated optogenetic stimulation of the hippocampal CA1 astrocytes induced cognitive impairment in mice and decreased synaptic long-term potentiation (LTP), which was accompanied by the appearance of inflammatory astrocytes. Mechanistic studies conducted using knockout animal models and hippocampal neuronal cultures showed that lipocalin-2 (LCN2), derived from reactive astrocytes, mediated neuroinflammation and induced cognitive impairment by decreasing the LTP through the reduction of neuronal NMDA receptors. Sustained chemogenetic stimulation of hippocampal astrocytes provided similar results. Conversely, these phenomena were attenuated by a metabolic inhibitor of astrocytes. Fiber photometry using GCaMP revealed a high level of hippocampal astrocyte activation in the neuroinflammation model. Our findings suggest that reactive astrocytes in the hippocampus are sufficient and required to induce cognitive decline through LCN2 release and synaptic modulation. This abnormal glial-neuron interaction may contribute to the pathogenesis of cognitive disturbances in neuroinflammation-associated brain conditions.

Regulation of inflammatory response by LINC00346 via miR-25-3p-mediated modulation of the PTEN/PI3K/AKT/NF-κB pathway.

Long intergenic non-coding RNA 346 (LINC00346) has been reported to be involved in the development of atherosclerosis and specific cancers by affecting signaling pathways. However, its function in inflammation has not been thoroughly studied. Therefore, its expression pattern and function were determined in the human macrophage-like cell line THP-1. Lipopolysaccharide (LPS) treatment induced the expression of LINC00346. LPS-induced NF-κB activation and proinflammatory cytokine expression were suppressed or enhanced by the overexpression or knockdown of LINC00346, respectively. Analyses using dual luciferase assay and decoy RNAs that could block RNA-RNA interactions indicated that LINC00346 improves phosphatase and tensin homolog (PTEN) expression by sponging miR-25-3p. Subsequently, PTEN suppresses phosphoinositide-3 kinase (PI3K)-mediated conversion of phosphatidylinositol-4,5-bisphosphate (PIP2) into phosphatidylinositol-3,4,5-trisphosphate (PIP3) as well as consequent activation of protein kinase B (AKT) and NF-κB. Interestingly, database analysis revealed that the expression levels of LINC00346 and PTEN were simultaneously decreased in breast cancer tissues. Further analyses conducted using a breast cancer cell line, MDA-MB-231, confirmed the functional relationship among LINC00346, miR-25-3p, and PTEN in LPS-induced activation of NF-κB. These results indicate that miR-25-3p-sponging activity of LINC00346 affects the balance between PTEN and PI3K as well as the downstream activation of AKT/NF-κB pathway in inflammatory conditions.

Subsequent correlated changes in complement component 3 and amyloid beta oligomers in the blood of patients with Alzheimer's disease.

INTRODUCTION: Alzheimer's disease (AD) involves the complement cascade, with complement component 3 (C3) playing a key role. However, the relationship between C3 and amyloid beta (Aß) in blood is limited. METHODS: Plasma C3 and Aß oligomerization tendency (AßOt) were measured in 35 AD patients and 62 healthy controls. Correlations with cerebrospinal fluid (CSF) biomarkers, cognitive impairment, and amyloid positron emission tomography (PET) were analyzed. Differences between biomarkers were compared in groups classified by concordances of biomarkers. RESULTS: Plasma C3 and AßOt were elevated in AD patients and in CSF or amyloid PET-positive groups. Weak positive correlation was found between C3 and AßOt, while both had strong negative correlations with CSF Aß42 and cognitive performance. Abnormalities were observed for AßOt and CSF Aß42 followed by C3 changes. DISCUSSION: Increased plasma C3 in AD are associated with amyloid pathology, possibly reflecting a defense response for Aß clearance. Further studies on Aß-binding proteins will enhance understanding of Aß mechanisms in blood.

Roles of lncRNAs in NF-κB-Mediated Macrophage Inflammation and Their Implications in the Pathogenesis of Human Diseases.

Over the past century, molecular biology's focus has transitioned from proteins to DNA, and now to RNA. Once considered merely a genetic information carrier, RNA is now recognized as both a vital element in early cellular life and a regulator in complex organisms. Long noncoding RNAs (lncRNAs), which are over 200 bases long but do not code for proteins, play roles in gene expression regulation and signal transduction by inducing epigenetic changes or interacting with various proteins and RNAs. These interactions exhibit a range of functions in various cell types, including macrophages. Notably, some macrophage lncRNAs influence the activation of NF-κB, a crucial transcription factor governing immune and inflammatory responses. Macrophage NF-κB is instrumental in the progression of various pathological conditions including sepsis, atherosclerosis, cancer, autoimmune disorders, and hypersensitivity. It orchestrates gene expression related to immune responses, inflammation, cell survival, and proliferation. Consequently, its malfunction is a key contributor to the onset and development of these diseases. This review aims to summarize the function of lncRNAs in regulating NF-κB activity in macrophage activation and inflammation, with a particular emphasis on their relevance to human diseases and their potential as therapeutic targets. The insights gained from studies on macrophage lncRNAs, as discussed in this review, could provide valuable knowledge for the development of treatments for various pathological conditions involving macrophages.

Protocol for delivering proteins, peptides, and shRNAs via the intrathecal route in the experimental allergic encephalomyelitis mouse model.

The blood-brain barrier hinders drug delivery to the central nervous system (CNS), particularly for large molecules. Here, we present a protocol for delivering proteins, peptides, and short hairpin RNAs (shRNAs) via the intrathecal (IT) route in the experimental allergic encephalomyelitis (EAE) mouse model. We describe steps for developing EAE in mice and administering treatments intrathecally. The insights into treatment efficacy that can be provided by this protocol make it an important tool for CNS research. For complete details on the use and execution of this protocol, please refer to Bhusal et al.1.

LINC01686 affects LPS-induced cytokine expression via the miR-18a-5p/A20/STAT1 axis in THP-1 cells.

BACKGROUND AND OBJECTIVE: Long noncoding RNAs (lncRNAs) are crucial in regulating various physiological and pathological processes, including immune responses. LINC01686 is a lncRNA with previously uncharacterized functions in immune regulation. This study aims to investigate the function of LINC01686 in lipopolysaccharide (LPS)-induced inflammatory responses in the human monocytic leukemia cell line THP-1 and its potential regulatory mechanisms involving miR-18a-5p and the anti-inflammatory protein A20. METHOD: THP-1 cells were stimulated with LPS to induce inflammatory responses, followed by analysis of LINC01686 expression levels. The role of LINC01686 in regulating the expression of interleukin (IL)-6, IL-8, A20, and signal transducer and activator of transcription 1 (STAT1) was examined using small interfering RNA-mediated knockdown. Additionally, the involvement of miR-18a-5p in LINC01686-mediated regulatory pathways was assessed by transfection with decoy RNAs mimicking the miR-18a-5p binding sites of LINC01686 or A20 messenger RNA. RESULTS: LINC01686 expression was upregulated in THP-1 cells following LPS stimulation. Suppression of LINC01686 enhanced LPS-induced expression of IL-6 and IL-8, mediated through increased production of reactive oxygen species. Moreover, LINC01686 knockdown upregulated the expression and activation of IκB-ζ, STAT1, and downregulated A20 expression. Transfection with decoy RNAs reversed the effects of LINC01686 suppression on A20, STAT1, IL-6, and IL-8 expression, highlighting the role of LINC01686 in sponging miR-18a-5p and regulating A20 expression. CONCLUSION: This study provides the first evidence that LINC01686 plays a critical role in modulating LPS-induced inflammatory responses in THP-1 cells by sponging miR-18a-5p, thereby regulating the expression and activation of A20 and STAT1. These findings shed light on the complex regulatory mechanisms involving lncRNAs in immune responses and offer potential therapeutic targets for inflammatory diseases.

Clinical Relevance of Plasma Prolylcarboxypeptidase Level in Patients with Idiopathic Acute Optic Neuritis.

Objectives: This study evaluated the plasma concentration of prolylcarboxypeptidase (PRCP) and its clinical relevance in patients with idiopathic acute optic neuritis (ON). Methods: We investigated the expression of PRCP in the optic nerves of experimental autoimmune optic neuritis (EAON)-induced mice. Peripheral blood samples were collected from ON patients (n = 20) and healthy controls (n = 20). ELISA was used to measure the plasma PRCP levels. We performed measurements of visual acuity and the mean thicknesses of the macular ganglion cell layer plus inner plexiform layer (GCL+IPL) at diagnosis and 6 months after diagnosis. Results: The PRCP mRNA expression in EAON-induced mice was markedly higher than that in naïve mice. The mean plasma PRCP level was significantly higher in patients with ON than in controls. Plasma PRCP levels were negatively correlated with logMAR visual acuity at 6 months after diagnosis and differences in macular GCL+IPL thickness during an ON attack. A plasma PRCP level of 49.98 (pg/mL) predicted the recurrence of ON with a 75% sensitivity and 87.5% specificity. Conclusions: Patients with idiopathic acute ON had higher plasma PRCP levels, and this was positively correlated with final visual outcome and well-preserved macular GCL+IPL thickness during an ON attack. The increase in plasma PRCP level may reflect its compensatory secretion to counteract neuroinflammation in ON patients.

Lipocalin-2 as a mediator of neuroimmune communication.

Lipocalin-2, a neutrophil gelatinase-associated lipocalin, is a 25-kDa secreted protein implicated in a broad range of inflammatory diseases affecting the brain and periphery. It is a pleotropic protein expressed by various immune and nonimmune cells throughout the body. Importantly, the surge in lipocalin-2 levels in disease states has been associated with a myriad of undesirable effects, further exacerbating the ongoing pathological processes. In the brain, glial cells are the principal source of lipocalin-2, which plays a definitive role in determining their functional phenotypes. In different central nervous system pathologies, an increased expression of glial lipocalin-2 has been linked to neurotoxicity. Lipocalin-2 mediates a crosstalk between central and peripheral immune cells under neuroinflammatory conditions. One intriguing aspect is that elevated lipocalin-2 levels in peripheral disorders, such as cancer, metabolic conditions, and liver diseases, potentially incite an inflammatory activation of glial cells while disrupting neuronal functions. This review comprehensively summarizes the influence of lipocalin-2 on the exacerbation of neuroinflammation by regulating various cellular processes. Additionally, this review explores lipocalin-2 as a mediator of neuroimmune crosstalk in various central nervous system pathologies and highlights the role of lipocalin-2 in carrying inflammatory signals along the neuroimmune axis.

Modulating effects of heat-killed and live Limosilactobacillus reuteri PSC102 on the immune response and gut microbiota of cyclophosphamide-treated rats.

In the present study, we investigated the potential immunomodulatory effects of heat-killed (hLR) and live Limosilactobacillus reuteri PSC102 (LR; formerly Lactobacillus reuteri PSC102) in RAW264.7 macrophage cells and Sprague-Dawley rats. RAW264.7 murine macrophage cells were stimulated with hLR and LR for 24 h. Cyclophosphamide (CTX)-induced immunosuppressed Sprague-Dawley rats were orally administered with three doses of hLR (L-Low, M-Medium, and H-High) and LR for 3 weeks. The phagocytic capacity, production of nitric oxide (NO), and expression of cytokines in RAW264.7 cells were measured, and the different parameters of immunity in rats were determined. hLR and LR treatments promoted phagocytic activity and induced the production of NO and the expression of iNOS, TNF-α, IL-1ß, IL-6, and Cox-2 in macrophage cells. In the in vivo experiment, hLR and LR treatments significantly increased the immune organ indices, alleviated the spleen injury, and ameliorated the number of white blood cells, granulocytes, lymphocytes, and mid-range absolute counts in immunosuppressive rats. hLR and LR increased neutrophil migration and phagocytosis, splenocyte proliferation, and T lymphocyte subsets (CD4+, CD8+, CD45RA+, and CD28+). The levels of immune factors (IL-2, IL-4, IL-6, IL-10, IL-12A, TNF-α, and IFN-γ) in the hLR and LR groups were upregulated compared with those in the CTX-treatment group. hLR and LR treatments could also modulate the gut microbiota composition, thereby increasing the relative abundance of Bacteroidetes and Firmicutes but decreasing the level of Proteobacteria. hLR and LR protected against CTX-induced adverse reactions by modulating the immune response and gut microbiota composition. Therefore, they could be used as potential immunomodulatory agents.

The microglial innate immune protein PGLYRP1 mediates neuroinflammation and consequent behavioral changes.

Peptidoglycan recognition protein 1 (PGLYRP1) is a pattern-recognition protein that mediates antibacterial actions and innate immune responses. Its expression and role in neuroinflammatory conditions remain unclear. We observed the upregulation of PGLYRP1 in inflamed human and mouse spinal cord and brain, with microglia being the primary cellular source. Experiments using a recombinant PGLYRP1 protein show that PGLYRP1 potentiates reactive gliosis, neuroinflammation, and consequent behavioral changes in multiple animal models of neuroinflammation. Furthermore, shRNA-mediated knockdown of Pglyrp1 gene expression attenuates this inflammatory response. In addition, we identify triggering receptor expressed on myeloid cell-1 (TREM1) as an interaction partner of PGLYRP1 and demonstrate that PGLYRP1 promotes neuroinflammation through the TREM1-Syk-Erk1/2-Stat3 axis in cultured glial cells. Taken together, our results reveal a role for microglial PGLYRP1 as a neuroinflammation mediator. Finally, we propose that PGLYRP1 is a potential biomarker and therapeutic target in various neuroinflammatory diseases.

Structural reorganization of medullary dorsal horn astrocytes in a rat model of neuropathic pain.

Multiple studies have shown that astrocytes in the medullary dorsal horn (MDH) play an important role in the development of pathologic pain. However, little is known about the structural reorganization of the peripheral astrocytic processes (PAP), the main functional part of the astrocyte, in MDH in neuropathic state. For this, we investigated the structural relationship between PAP and their adjacent presynaptic axon terminals and postsynaptic dendrites in the superficial laminae of the MDH using electron microscopical immunohistochemistry for ezrin, a marker for PAP, and quantitative analysis in a rat model of neuropathic pain following chronic constriction injury of the infraorbital nerve (CCI-ION). We found that, compared to controls, in rats with CCI-ION, (1) the number, % area, surface density, and volume fraction of ezrin-positive (+) PAP, as well as the fraction of synaptic edge apposed by ezrin + PAP and the degree of its coverage of presynaptic axon terminals and postsynaptic dendrites increased significantly, (2) these effects were abolished by administration of the mGluR5 antagonist 2-methyl-6-(phenylethynyl) pyridine (MPEP). These findings indicate that PAP undergoes structural reorganization around the central synapses of sensory afferents following nerve injury, suggest that it may be mediated by mGluR5, and may represent the structural basis for enhancing astrocyte-neuron interaction in neuropathic pain.