The SRC/NF-κB-AKT/NOS3 axis as a key mediator of Kaempferol's protective effects against oxidative stress-induced osteoclastogenesis.

BACKGROUND: Osteoclasts are integral to the advancement of osteoporosis (OP), and their generation under conditions of oxidative stress (OS) involves various pathways. However, the specific mechanism through which the natural antioxidant kaempferol (KAE) mitigates the influence of OS on osteoclasts remains somewhat uncertain. This study aims to evaluate the effect of KAE on osteoclast formation under OS and explore its possible mechanism. METHODS: Zebrafish were used to observe the effects of KAE on OP and OS. OP and OS "double disease targets" network pharmacology were used to predict the action target and mechanism of KAE on OP under OS. The effects of KAE on osteoclast differentiation induced by OS were evaluated using RWA264.7 cells induced by LPS. To elucidate the potential mechanism, we detected the expression of related factors and target genes during induction. RESULTS: The presence of KAE exhibited potential in improving the conditions of OP and OS in zebrafish. KAE can reduce the OS of RAW 264.7 cells stimulated by LPS, inhibit the formation of osteoclasts, and change the level of related factors of OS, and reduce the increase of TRAP. The utilization of network pharmacology and target gene expression assay revealed that KAE exerted a down-regulatory effect on the expression of proto-oncogene tyrosine protein kinase (SRC), nuclear factor kappa-B (NF-κB), Serine/Threonine Kinase-1 (AKT1), Nitric Oxide Synthase 3 (NOS3) and Matrix Metallopeptidase-2 (MMP2). CONCLUSION: Based on the results of this study, KAE may effectively mitigate OS and impede the formation of osteoclasts through the SRC/NF-κB-AKT/NOS3 axis.

CD169+ Macrophages Mediate the Immune Response of Allergic Rhinitis Through the Keap1/Nrf2/HO-1 Axis.

CD169+ macrophages are a newly defined macrophage subpopulation that can recognize and bind with other cells through related ligands, playing an essential role in antigen presentation and immune tolerance. However, its role in Allergic Rhinitis (AR) is still unclear. To investigate the characteristics of CD169+ macrophages in AR, this work first detects their expression patterns in the nasal mucosa of clinical patients. These results show a significant increase in CD169+ macrophages in the nasal mucosa of patients with AR. Subsequently, this work establishes an animal AR model using CD169 transgenic mice and compared the advantages of the two models. Moreover, this work also demonstrates the effects of CD169 knockout on eosinophils, Th cells, Treg cells, and the migration of dendritic cells (DCs). In addition, this metabolomic data shows that CD169+ macrophages can upregulate alanine production and increase reactive oxygen species (ROS) levels. This process may be mediated through the Keap1/Nrf2/HO-1 signaling pathway. In addition, this work also finds that SLC38A2 plays an essential role in the process of CD169+ macrophages promoting alanine uptake by DCs. This study confirms that CD169+ macrophages can upregulate their internal alanine production and increase ROS levels through the Keap1/Nrf2/HO-1 axis, playing an irreplaceable role in AR.

Using a Dual-Disease Target Mapping Network Pharmacology Approach, Verbascoside Ameliorates Osteoporosis by Activating Estrogen Signaling to Alleviate Oxidative Stress.

BACKGROUND: Verbascoside, a compound classified as a phenylethanol glycoside in Dihuang, has been the subject of modern pharmacological investigations. These studies have revealed its noteworthy antioxidant, anti-inflammatory, memory-enhancing, neuroprotective, antitumor, and various other pharmacological properties. While verbascoside exhibits favorable antioxidant effects, its precise mechanism of action in ameliorating osteoporosis through the treatment of oxidative stress remains unclear. METHODS: This study employed CCK8, ALP, ELISA, and ROS staining techniques to examine the osteoporotic effects of verbascoside on zebrafish and MC3T3-E1 cells. Additionally, this study aimed to investigate the molecular mechanism by which verbascoside improves osteoporosis by mitigating oxidative stress. To identify the common targets of verbascoside in relation to oxidative stress and osteoporosis, network pharmacology and molecular dynamics simulation were employed. The construction of the verbascoside - oxidative stress - osteoporosis - potential target gene network aimed to identify the core targets, while the mechanism of action was elucidated through KEGG analysis, and the accuracy was confirmed by assessing the mRNA expression of the targets. RESULTS: In vivo experiments demonstrated that verbascoside exhibited therapeutic effects on osteoporosis and reduced ROS production in zebrafish. In vitro experiments further revealed that verbascoside enhanced the proliferation and differentiation of MC3T3-E1 cells, thereby improving the oxidative stress status of osteoblasts. Thirteen core targets and estrogen signaling pathways were identified through the application of network pharmacology. The pivotal role of the estrogen signaling pathway in facilitating the ability of verbascoside to mitigate oxidative stressinduced osteoporosis was substantiated by the modulation of target protein mRNA expression. CONCLUSION: The findings underscore the considerable therapeutic potential of verbascoside in ameliorating osteoporosis through the alleviation of oxidative stress, thus establishing it as a promising compound for the treatment of this condition.

Tailoring the pKa of Fluorescence Lifetime Imaging Probes to Visualize Aggrephagy and Resolve Its Microenvironmental Viscosity.

Aggrephagy describes lysosomal transport and degradation of protein aggregates via cellular macroautophagy, a key mechanism to prevent neurodegenerative diseases. Here, we develop a dual-probe method to visualize the aggrephagy process and resolve its viscosity heterogeneity using fluorescence lifetime imaging (FLIM). The dual-probe system consists of (1) a near-infrared lysosomal targeting FLIM probe (Lyso-P1) that is derived from a rhodamine scaffold with a tailored pKa value to accommodate an acidic lysosomal environment and (2) a green BODIPY-based FLIM probe (Agg-P2) that reports on degradation of cellular aggregates via HaloTag. Both probes exhibit acid-resistant, viscosity-dependent fluorescence intensity and lifetime (τ) responses, which are ready for intensity- and FLIM-based imaging. Photochemical, theoretical, and biochemical characterizations reveal the probes' mechanism-of-actions. In cells, we exploit Lyso-P1 and Agg-P2 to simultaneously quantify both lysosomal and protein aggegates' viscosity changes upon the aggrephagy process via FLIM. We reveal orthogonal changes in microenvironmental viscosities and morphological heterogeneity upon various cellular stresses. Overall, we provide an imaging toolset to quantitatively study aggrephay, which may benefit screening of aggrephay modulators for disease intervention.

Mechanisms of Zhixiao Tang on Anti-Inflammatory Multiple Targets and Multiple Components: Metabonomics Combined with Database Mining Technology.

Purpose: Zhixiao Tang (ZXT), a traditional Chinese compound prescription, has been used clinically to treat pneumonia in China. However, the underlying mechanism of ZXT treatment in pneumonia is still unclear. The present study aimed to reveal the potential mechanism of ZXT in pneumonia using a strategy combining metabolomics and network pharmacology. Methods: Initially, the chemical compositions were identified by UPLC-QE-Orbitrap-MS, while the prediction of potential signal pathways was performed through network pharmacology. To assess the anti-inflammatory properties of ZXT in the context of pneumonia, models of 16HBE cells induced by LPS and zebrafish induced by CuSO4 were established to measure levels of inflammatory markers and apoptosis. Subsequently, the differential changes of endogenous metabolites in cells caused by ZXT were examined using metabolomics technology, and the molecular docking analysis of key targets was carried out using Autodock Vina software. Ultimately, the validation of the primary pathways and targets was conducted through quantitative RT-PCR and Western blot techniques. Results: A total of 75 compounds were identified through UPLC-QE-Orbitrap-MS analyses. Network pharmacological analysis shows that it plays an anti-inflammatory role in C-type lectin receptor signaling pathway. After ZXT intervention, the inflammatory factors and apoptosis in cells were significantly reduced. Metabonomics analysis showed that 18 metabolites changed significantly. Four key genes were identified, which exhibited partial compatibility with the findings of network pharmacology. Molecular docking analysis confirmed the substantial affinity of the primary targets for ZXT. Furthermore, ZXT exerted a suppressive effect on neutrophil migration, down-regulated the expression of pro-inflammatory cytokine genes, and inhibited the up-regulation of the Dectin-1/SYK/NF-κB signaling pathway. In vivo cell experiments also yielded consistent experimental outcomes. Conclusion: This study enhances comprehension of the pharmacological mechanism underlying ZXT's efficacy in pneumonia treatment, thereby establishing a scholarly basis for future research and clinical utilization of ZXT in pneumonia management.

NRhFluors: Quantitative Revealing the Interaction between Protein Homeostasis and Mitochondria Dysfunction via Fluorescence Lifetime Imaging.

Degenerative diseases are closely related to the changes of protein conformation beyond the steady state. The development of feasible tools for quantitative detection of changes in the cellular environment is crucial for investigating the process of protein conformational variations. Here, we have developed a near-infrared AIE probe based on the rhodamine fluorophore, which exhibits dual responses of fluorescence intensity and lifetime to local viscosity changes. Notably, computational analysis reveals that NRhFluors fluorescence activation is due to inhibition of the RACI mechanism in viscous environment. In the chemical regulation of rhodamine fluorophores, we found that variations of electron density distribution can effectively regulate CI states and achieve fluorescence sensitivity of NRhFluors. In addition, combined with the AggTag method, the lifetime of probe A9-Halo exhibits a positive correlation with viscosity changes. This analytical capacity allows us to quantitatively monitor protein conformational changes using fluorescence lifetime imaging (FLIM) and demonstrate that mitochondrial dysfunction leads to reduced protein expression in HEK293 cells. In summary, this work developed a set of near-infrared AIE probes activated by the RACI mechanism, which can quantitatively detect cell viscosity and protein aggregation formation, providing a versatile tool for exploring disease-related biological processes and therapeutic approaches.

Versatile Fluorescence Lifetime-Based Copper Probe to Quantify Mitochondrial Membrane Potential and Reveal Its Interaction with Protein Aggregation.

Mitochondria play a crucial role in maintaining cellular homeostasis, and the depolarization of mitochondrial membrane potential (MMP) is an important signal of apoptosis. Additionally, protein misfolding and aggregation are closely related to diseases including neurodegenerative diseases, diabetes, and cancers. However, the interaction between MMP changes and disease-related protein aggregation was rarely studied. Herein, we report a novel "turn-on" fluorescent probe MitoRhB that specifically targets to mitochondria for Cu2+ detection in situ. The fluorescence lifetime (τ) of MitoRhB exhibits a positive correlation with MMP changes, allowing us to quantitatively determine the relative MMP during SOD1 (A4 V) protein aggregation. Finally, we found that (1) the increasing concentrations of copper will accelerate the depolarization of mitochondria and reduce MMP; (2) the depolarization of mitochondria can intensify the degree of protein aggregation, suggesting a new routine of copper-induced cell death mediated through abnormal MMP depolarization and protein aggregation.

Integrated cell metabolomics and network pharmacology approach deciphers the anti-testosterone deficiency mechanisms of Bushen Zhuanggu Tang.

Testicular dysfunction is distinguished by a deficiency in testosterone levels, which can be attributed to the occurrence of oxidative stress injury in Leydig cells. The empirical prescription known as Bushen Zhuanggu Tang, developed by a highly experienced traditional Chinese medicine practitioner with six decades of clinical expertize, aligns with the traditional Chinese medicine principle of "kidney governing bone". Researchers have demonstrated that the administration of BSZGT can effectively enhance testosterone production. The objective of this study is to investigate the potential anti-testicular dysfunction effects of BSZGT and elucidate its underlying mechanism in an in vitro setting. Specifically, the impact of oxidative stress induced by H2O2 on the activity and testosterone levels of Leydig cells (TM3) was examined. Furthermore, the utilization of UPLC-QE-Qrbitrap-MS enabled the identification of the involvement of BSZGT in various metabolic pathways, including arginine biosynthesis, amino acyl-tRNA biosynthesis, Alanine, aspartate and glutamine metabolism, and Citrate Cycle, through the modulation of 25 distinct metabolites. Additionally, a network pharmacological analysis was conducted to investigate the pivotal protein targets associated with the therapeutic effects of BSZGT. The findings demonstrate the identification of six key proteins (CYP19A1, CYP1B1, ALOX5, ARG1, XDH, and MPO) that play a significant role in augmenting testicular function through their involvement in the ovarian steroid production pathway. In summary, our study presents a comprehensive research methodology that combines cell metabonomics and network pharmacology to enhance the discovery of new therapeutic agents for TD.

Exploring Asthma Mechanism of Belamcanda Chinensis by "Dose-effect Weighted Coefficient" Network Pharmacology and Experiment Validation.

BACKGROUND: Asthma is a chronic inflammatory disease of the airways that seriously endangers human health. Belamcanda chinensis (BC), a traditional Chinese medicine, has been used to counteract asthma as it has been shown to possess anti-inflammatory and regulatory immunity properties. OBJECTIVE: The study aimed to investigate the mechanisms of action of BC in the treatment of asthma; a "dose-effect weighted coefficient" network pharmacology method was established to predict potential active compounds. METHODS: Information on the components and content of BC was obtained by UPLC-QEOrbitrap- MS spectrometry. Based on BC content, oral bioavailability, and molecular docking binding energy, dose-effect weighting coefficients were constructed. With the degree greater than average as the index, a protein-protein interaction (PPI) database was used to obtain the core key targets for asthma under dose-effect weighting. GO function and KEGG pathway analyses of the core targets were performed using DAVID software. Finally, MTT and ELISA assays were used to assess the effects of active components on 16HBE cell proliferation. RESULTS: The experimental results using the 16HBE model demonstrated BC to have a potential protective effect on asthma. Network pharmacology showed SYK, AKT1, and ALOX5 to be the main key targets, and Fc epsilon RI as the promising signaling pathway. Eight components, such as tectoridin, mangiferin, luteolin, and isovitexin were the main active compounds, Finally, we analyzed the LPS-induced 16HBE proliferation of each active ingredient. Based on the activity verification study, all five predicted components promoted the proliferation of 16HBE cells. These five compounds can be used as potential quality markers for asthma. CONCLUSION: This study provides a virtual and practical method for the simple and rapid screening of active ingredients in natural products.

MGA-seq: robust identification of extrachromosomal DNA and genetic variants using multiple genetic abnormality sequencing.

Genomic abnormalities are strongly associated with cancer and infertility. In this study, we develop a simple and efficient method - multiple genetic abnormality sequencing (MGA-Seq) - to simultaneously detect structural variation, copy number variation, single-nucleotide polymorphism, homogeneously staining regions, and extrachromosomal DNA (ecDNA) from a single tube. MGA-Seq directly sequences proximity-ligated genomic fragments, yielding a dataset with concurrent genome three-dimensional and whole-genome sequencing information, enabling approximate localization of genomic structural variations and facilitating breakpoint identification. Additionally, by utilizing MGA-Seq, we map focal amplification and oncogene coamplification, thus facilitating the exploration of ecDNA's transcriptional regulatory function.

Single cell RNA sequencing unravels mechanisms underlying senescence-like phenotypes of alveolar macrophages.

Alveolar macrophages (AMs) are resident innate immune cells that play vital roles in maintaining lung physiological functions. However, the effects of aging on their dynamics, heterogeneity, and transcriptional profiles remain to be fully elucidated. Through single cell RNA sequencing (scRNA-seq), we identified CBFß as an indispensable transcription factor that ensures AM self-renewal. Intriguingly, despite transcriptome similarities of proliferating cells, AMs from aged mice exhibited reduced embryonic stem cell-like features. Aged AMs also displayed compromised DNA repair abilities, potentially leading to obstructed cell cycle progression and an elevation of senescence markers. Consistently, AMs from aged mice exhibited impaired self-renewal ability and reduced sensitivity to GM-CSF. Decreased CBFß was observed in the cytosol of AMs from aged mice. Similar senescence-like phenotypes were also found in human AMs. Taken together, these findings suggest that AMs in aged hosts demonstrate senescence-like phenotypes, potentially facilitated by the abrogated CBF ß activity.

A transcriptomics and network pharmacology approach to elucidate the mechanism of action of geniposide on carbon tetrachloride-induced liver injury in rats.

Geniposide, the main active component of Fructus Gardeniae (FG), is known to confer protection against liver diseases. Herein we explored the hepatoprotective effects of geniposide and elucidated its molecular mechanism by transcriptome RNA-seq and network pharmacology. Liver injury was modeled by intraperitoneally injecting CCl4 (0.15% prepared with refined peanut oil) at a dose of 1.5 mL/kg thrice a week; from the second week, rats were administered geniposide (20 mg/kg or 40 mg/kg) by gavage for 6 weeks. Serum and liver samples were then collected to assess liver function indicators and inflammatory factors and to observe pathological changes in the liver. The Illumina HiSeq 4000 platform was used to obtain transcriptome data from the liver tissue of rats after geniposide administration. Core targets and pathways related to the liver protection mechanism of geniposide were further analyzed by integrating transcriptomics and network pharmacology. Differentially expressed genes (DEGs), core targets, and signaling pathways were identified by methods such as q-PCR, molecular docking, and Western blotting. We found that after geniposide administration, the levels of aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, and inflammatory factors decreased in the model group, and liver injury cells be effectively repaired. RNA-seq data analysis showed that compared to control group, the model group reversed 1,451 DEGs; further, compared to model group, geniposide reversed 511 DEGs. Eight key targets, including PIK3R1, ACOX3, and EGF, were found through further analyses. Geniposide was determined to mainly regulate the PPAR signaling pathway, apoptosis signaling pathway, and MAPK signaling pathway in liver tissues. To summarize, the protective and restorative effects of geniposide on rat liver may seem to be related to its efficacy in inhibiting the activation of inflammatory pathways, thereby reducing cell apoptosis. Our findings should serve as the basis for the development of functional foods or drugs to prevent and treat liver diseases.

UPLC-QE-Orbitrap-Based Cell Metabolomics and Network Pharmacology to Reveal the Mechanism of N-Benzylhexadecanamide Isolated from Maca (Lepidium meyenii Walp.) against Testicular Dysfunction.

Testicular dysfunction (TDF) is characterized by testosterone deficiency and is caused by oxidative stress injury in Leydig cells. A natural fatty amide named N-benzylhexadecanamide (NBH), derived from cruciferous maca, has been shown to promote testosterone production. Our study aims to reveal the anti-TDF effect of NBH and explore its potential mechanism in vitro. This study examined the effects of H2O2 on cell viability and testosterone levels in mouse Leydig cells (TM3) under oxidative stress. In addition, cell metabolomics analysis based on UPLC-Q-Exactive-MS/MS showed that NBH was mainly involved in arginine biosynthesis, aminoacyl-tRNA biosynthesis, phenylalanine, tyrosine and tryptophan biosynthesis, the TCA cycle and other metabolic pathways by affecting 23 differential metabolites, including arginine and phenylalanine. Furthermore, we also performed network pharmacological analysis to observe the key protein targets in NBH treatment. The results showed that its role was to up-regulate ALOX5, down-regulate CYP1A2, and play a role in promoting testicular activity by participating in the steroid hormone biosynthesis pathway. In summary, our study not only provides new insights into the biochemical mechanisms of natural compounds in the treatment of TDF, but also provides a research strategy that integrates cell metabolomics and network pharmacology in order to promote the screening of new drugs for the treatment of TDF.

H1N1 influenza virus infection through NRF2-KEAP1-GCLC pathway induces ferroptosis in nasal mucosal epithelial cells.

Influenza A virus can induce nasal inflammation by stimulating the death of nasal mucosa epithelium, however, the mechanism is not clear. In this study, to study the causes and mechanisms of nasal mucosa epithelial cell death caused by Influenza A virus H1N1, we isolated and cultured human nasal epithelial progenitor cells (hNEPCs) and exposed them to H1N1 virus after leading differentiation. Then we performed high-resolution untargeted metabolomics and RNAseq analysis of human nasal epithelial cells (hNECs) infected with H1N1 virus. Surprisingly, H1N1 virus infection caused the differential expression of a large number of ferroptosis related genes and metabolites in hNECs. Furthermore, we have observed a significant reduction in Nrf2/KEAP1 expression, GCLC expression, and abnormal glutaminolysis. By constructing overexpression vector of GCLC and the shRNAs of GCLC and Keap1, we determined the role of NRF2-KEAP1-GCLC signaling pathway in H1N1 virus-induced ferroptosis. In addition, A glutaminase antagonist, JHU-083, also demonstrated that glutaminolysis can regulate the NRF2-KEAP1-GCLC signal pathway and ferroptosis. According to this study, H1N1 virus can induce the ferroptosis of hNECs via the NRF2-KEAP1-GCLC signal pathway and glutaminolysis, leading to nasal mucosal epithelial inflammation. This discovery is expected to provide an attractive therapeutic target for viral-induced nasal inflammation.

Zika virus replication on endothelial cells and invasion into the central nervous system by inhibiting interferon ß translation.

The blood-brain barrier (BBB) is one of the tightest physical barriers to prevent pathogens from invading the central nervous system (CNS). However, the mechanism by which Zika virus (ZIKV) crossing the BBB remains unresolved. We found ZIKV induced high morbidity and mortality in newborn mice, accompanied by inflammatory injury on CNS. ZIKV was found to replicate primarily in the cortex and hippocampus in neonatal mouse brains. An in vitro model revealed that ZIKV had no impact on hBMECs permeability but led to endothelial activation, as shown by the enhancement of adhesion molecules expression and F-actin redistribution. ZIKV replication in hBMECs might be associated with the suppression of IFN-ß translation via inhibiting RPS6 phosphorylation. On the other hand, ZIKV infection induced IFN-stimulated genes (ISGs), activated the mitogen-activated protein kinase (MAPK) signaling pathway, and promoted chemokine secretion. This study provides an understanding of virus replication and transmigration across the BBB during ZIKV infection.

Screening of the Active Compounds against Neural Oxidative Damage from Ginseng Phloem Using UPLC-Q-Exactive-MS/MS Coupled with the Content-Effect Weighted Method.

The neuroprotective properties of ginsenosides have been found to reverse the neurological damage caused by oxidation in many neurodegenerative diseases. However, the distribution of ginsenosides in different tissues of the main root, which was regarded as the primary medicinal portion in clinical practice was different, the specific parts and specific components against neural oxidative damage were not clear. The present study aims to screen and determine the potential compounds in different parts of the main root in ginseng. Comparison of the protective effects in the main root, phloem and xylem of ginseng on hydrogen peroxide-induced cell death of SH-SY5Y neurons was investigated. UPLC-Q-Exactive-MS/MS was used to quickly and comprehensively characterize the chemical compositions of the active parts. Network pharmacology combined with a molecular docking approach was employed to virtually screen for disease-related targets and potential active compounds. By comparing the changes before and after Content-Effect weighting, the compounds with stronger anti-nerve oxidative damage activity were screened out more accurately. Finally, the activity of the selected monomer components was verified. The results suggested that the phloem of ginseng was the most effective part. There were 19 effective compounds and 14 core targets, and enriched signaling pathway and biological functions were predicted. After Content-Effect weighting, compounds Ginsenosides F1, Ginsenosides Rf, Ginsenosides Rg1 and Ginsenosides Rd were screened out as potential active compounds against neural oxidative damage. The activity verification study indicated that all four predicted ginsenosides were effective in protecting SH-SY5Y cells from oxidative injury. The four compounds can be further investigated as potential lead compounds for neurodegenerative diseases. This also provides a combined virtual and practical method for the simple and rapid screening of active ingredients in natural products.

Multiple Death Pathways of Neutrophils Regulate Alveolar Macrophage Proliferation.

Alveolar macrophage (AM) proliferation and self-renewal play an important role in the lung tissue microenvironment. However, the impact of immune cells, especially the neutrophils, on AM homeostasis or function is not well characterized. In this study, we induced in vivo migration of neutrophils into bronchoalveolar lavage (BAL) fluid and lung using CXCL1, and then co-cultured these with AMs in vitro. Neutrophils in the BAL (BAL-neutrophils), rather than neutrophils of bone marrow (BM-neutrophils), were found to inhibit AM proliferation. Analysis of publicly available data showed high heterogeneity of lung neutrophils with distinct molecular signatures of BM- and blood-neutrophils. Unexpectedly, BAL-neutrophils from influenza virus PR8-infected mice (PR8-neutrophils) did not inhibit the proliferation of AMs. Bulk RNA sequencing further revealed that co-culture of AMs with PR8-neutrophils induced IFN-α and -γ responses and inflammatory response, and AMs co-cultured with BAL-neutrophils showed higher expression of metabolism- and ROS-associated genes; in addition, BAL-neutrophils from PR8-infected mice modulated AM polarization and phagocytosis. BAL-neutrophil-mediated suppression of AM proliferation was abrogated by a combination of inhibitors of different neutrophil death pathways. Collectively, our findings suggest that multiple cell death pathways of neutrophils regulate the proliferation of AMs. Targeting neutrophil death may represent a potential therapeutic strategy for improving AM homeostasis during respiratory diseases.

Altered gene expression in human brain microvascular endothelial cells in response to the infection of influenza H1N1 virus.

Influenza viruses not only cause respiratory illness, but also have been reported to elicit neurological manifestations following acute viral infection. The central nervous system (CNS) has a specific defense mechanism against pathogens structured by cerebral microvasculature lined with brain endothelial cells to form the blood-brain barrier (BBB). To investigate the response of human brain microvascular endothelial cells (hBMECs) to the Influenza A virus (IAV), we inoculated the cells with the A/WSN/33 (H1N1) virus. We then conducted an RNAseq experiment to determine the changes in gene expression levels and the activated disease pathways following infection. The analysis revealed an effective activation of the innate immune defense by inducing the pattern recognition receptors (PRRs). Along with the production of proinflammatory cytokines, we detected an upregulation of interferons and interferon-stimulated genes, such as IFN-ß/λ, ISG15, CXCL11, CXCL3 and IL-6, etc. Moreover, infected hBMECs exhibited a disruption in the cytoskeletal structure both on the transcriptomic and cytological levels. The RNAseq analysis showed different pathways and candidate genes associated with the neuroactive ligand-receptor interaction, neuroinflammation, and neurodegenerative diseases, together with a predicted activation of the neuroglia. Likewise, some genes linked with the mitochondrial structure and function displayed a significantly altered expression. En masse, this data supports that hBMECs could be infected by the IAV, which induces the innate and inflammatory immune response. The results suggest that the influenza virus infection could potentially induce a subsequent aggravation of neurological disorders. Supplementary Information: The online version contains supplementary material available at 10.1186/s44149-022-00053-9.

JEV Infection Induces M-MDSC Differentiation Into CD3+ Macrophages in the Brain.

Japanese encephalitis virus (JEV) is one of the most important members of the flavivirus family. It is a typical zoonotic pathogen that has caused substantial social and economic losses worldwide. The relation between JEV-induced immunosuppression and inflammatory responses has not been thoroughly investigated. In this study, cells infiltrating the brain tissue of JEV-infected mice were mainly identified as monocytic myeloid-derived suppressor cells (M-MDSCs), which subsequently differentiated into CD3+ macrophages. Co-culture with T cells showed that both splenic M-MDSCs and brain infiltrated M-MDSCs isolated from JEV-infected mice inhibited T cell proliferation through ARG1 and iNOS. The splenectomy model revealed that JEV-induced M-MDSCs were mainly derived from bone marrow and migrated to the spleen and central nervous system (CNS). The results of the transcriptome analysis and IRF7-deficient mice indicated that the ZBP1-IRF7 signaling pathway stimulated by JEV RNA played a central role in the induction of M-MDSCs. M-MDSCs migrated into the CNS through the chemokine CCL2/N-CCL2 derived from astrocytes and brain infiltrated M-MDSCs differentiated into CD3+ macrophages through a mechanism mediated by M-CSF, IL-6 and IFN-γ in the brain microenvironment. These findings provide evidence for the mechanism that JEV regulates the differentiation of M-MDSCs and thereby exacerbates pathogenicity, which represents a potential therapeutic target for Japanese encephalitis (JE).

TFAM-Dependent Mitochondrial Metabolism Is Required for Alveolar Macrophage Maintenance and Homeostasis.

Alveolar macrophages (AMs) are major lung tissue-resident macrophages capable of proliferating and self-renewal in situ. AMs are vital in pulmonary antimicrobial immunity and surfactant clearance. The mechanisms regulating AM compartment formation and maintenance remain to be fully elucidated currently. In this study, we have explored the roles of mitochondrial transcription factor A (TFAM)-mediated mitochondrial fitness and metabolism in regulating AM formation and function. We found that TFAM deficiency in mice resulted in significantly reduced AM numbers and impaired AM maturation in vivo. TFAM deficiency was not required for the generation of AM precursors nor the differentiation of AM precursors into AMs, but was critical for the maintenance of AM compartment. Mechanistically, TFAM deficiency diminished gene programs associated with AM proliferation and self-renewal and promoted the expression of inflammatory genes in AMs. We further showed that TFAM-mediated AM compartment impairment resulted in defective clearance of cellular debris and surfactant in the lung and increased the host susceptibility to severe influenza virus infection. Finally, we found that influenza virus infection in AMs led to impaired TFAM expression and diminished mitochondrial fitness and metabolism. Thus, our data have established the critical function of TFAM-mediated mitochondrial metabolism in AM maintenance and function.