Macrophage-expressed Gene Research Articles

D-Xylose Ameliorates Non-Alcoholic Fatty Liver Disease by Targeting Macrophage-expressed LYZ Gene.

This study investigates the therapeutic effects of D-Xylose, a natural sugar, on non-alcoholic fatty liver disease (NAFLD), focusing on the expression of the lysozyme gene (LYZ) in macrophages. Using the single-cell dataset GSE136103 for NAFLD, researchers analyzed macrophage populations and other groups utilizing the Seurat package in R, while a differential analysis was performed on the NAFLD dataset GSE61260 using the limma package. Both in vitro and in vivo models, including cell culture, mouse models, RT-qPCR, Western blot, ELISA, and histopathological analyses, were employed to examine the effect of D-Xylose on lipid accumulation, LYZ expression, blood lipid levels, and inflammatory responses. The study found a significant upregulation of LYZ in free fatty acid (FFA)-treated cells and mouse liver tissues, with a subsequent reduction after D-Xylose intervention. Treatment with D-Xylose and Amlodipine led to a notable decrease in lipid accumulation, as evidenced by reduced triglyceride and cholesterol levels. D-Xylose demonstrated a greater improvement in lipid metabolism than Amlodipine. Additionally, D-Xylose significantly mitigated inflammatory responses, reducing levels of inflammatory markers such as IL1R, IL6, MYS8, TNF, NF-κB, and IL-1. Furthermore, D-Xylose administration significantly reduced liver weight and liver index, with a positive impact on serum liver function and blood lipid levels. The findings suggest that D-Xylose could be a therapeutic intervention for NAFLD by targeting LYZ expression in macrophages, thereby modulating lipid metabolism and inflammatory responses.

Pichia pastoris composition expressed aerolysin mutant of Aeromonas veronii as an oral vaccine evaluated in zebrafish (Danio rerio)

Vaccines are one of the most practical means to stop the spreading of Aeromonas veronii in aquaculture. In this study, virulence factor aerolysin mutant NTaer which has lost its hemolytic activity was used as a target antigen. Pichia pastoris constitutive secretory expression NTaer (GS115-NTaer) was used as a potential safe oral vaccine to evaluate its effectiveness on zebrafish immunity. The result shows that vaccination of GS115- NTaer for four weeks did not affect the growth performance of the host, while eliciting an effective immune protective response. Compared with the control group, the GS115-NTaer could significantly up-regulate the relative expression level of the intestinal tight junction protein 1α (TJP1α) gene, and significantly increased the contents of lysozyme (LYZ), complement C3 and C4 in the gut, indicating that the innate immune response of the fish was activated. The relative gene expression levels of macrophage-expressed gene 1 (MPEG1) and T cell receptor (TCR-α) in the gut, and MPEG1, CD4, CD8, TCR-α, GATA3, and T-bet in the spleen were all increased significantly, indicating that the cellular immune response of the fish was activated. Furthermore, the contents of serum IgM and intestinal mucosa IgZ antibodies were significantly increased, which showed that humoral immunity was also activated. Moreover, inoculation with GS115-NTaer significantly changed the structure of gut microbiota. In particular, the relative ratio of (Firmicutes + Fusobacteriota + Bacteroidota)/Proteobacteria was significantly higher than that of the control and GS115 groups. Lastly, the vaccinated fish were challenged with A. veronii, and the relative percent survival of GS115 and the GS115-NTear groups was 14.28% and 33.43%. This improvement of immunity was not only due to the specific immune response but also attributed to the improvement of innate immunity and the gut microbiota which was demonstrated by the germ-free zebrafish model. Collectively, this study provides information on the effectiveness of GS115-NTear as an oral vaccine for the green prevention and control of A. veronii infection in fish aquaculture.

The immunomodulatory role of paracrine signalling factor VSIG4 in peritoneal metastases

Peritoneal metastasis (PM), the regional progression of intra-abdominal malignancies, is a common sequelae of colorectal cancer (CRC). Immunotherapy is slated to be effective in generating long-lasting anti-tumour response as it utilizes the specificity and memory of the immune system. In the tumour microenvironment, tumour associated macrophages (TAMs) are posited to create an anti-inflammatory pro-tumorigenic environment. In this paper, we aimed to identify immunomodulatory factors associated with colorectal PM (CPM). A publicly available colorectal single cell database (GSE183916) was analysed to identify possible immunological markers that are associated with the activation of macrophages in cancers. Immunohistochemical analysis for V-set and immunoglobin containing domain 4 (VSIG4) expression was performed on tumour microarrays (TMAs) of tumours of colorectal origin (n = 211). Expression of VSIG4 in cell-free ascites obtained from CPM patients (n = 39) was determined using enzyme-linked immunosorbent assay (ELISA). CD163-positive TAMs cluster expression was extracted from a publicly available single cell database and evaluated for the top 100 genes. From these macrophage-expressed genes, VSIG4, a membrane protein produced by the M2 macrophages, mediates the up-regulation of anti-inflammatory and down-regulation of pro-inflammatory macrophages, contributing to an overall anti-inflammatory state. CRC TMA IHC staining showed that low expression of VSIG4 in stromal tissues of primary CRC are associated with poor prognosis (p = 0.0226). CPM ascites also contained varying concentrations of VSIG4, which points to a possible role of VSIG4 in the ascites. The contribution of VSIG4 to CPM development can be further evaluated for its potential as an immunotherapeutic agent.

Accumulation and Phagocytosis of Fluorescently Visualized Macrophages Against Edwardsiella piscicida Infection in Established mpeg1.1-Transgenic Japanese Medaka Oryzias latipes.

Intracellular bacteria such as those belonging to the genus Edwardsiella can survive and proliferate within macrophages. However, the detailed mechanisms underlying the host macrophage immune response and pathogen evasion strategies remain unknown. To advance the field of host macrophage research, we successfully established transgenic (Tg) Japanese medaka Oryzias latipes that possessesfluorescently visualized macrophages. As a macrophage marker, the macrophage-expressed gene 1.1 (mpeg1.1) was selected because of its predominant expression across various tissues in medaka. To validate the macrophage characteristics of the fluorescently labeled cells, May-Grünwald Giemsa staining and peroxidase staining were conducted. The labeled cells exhibited morphological features consistent with those of monocyte/macrophage-like cells and tested negative for peroxidase activity. Through co-localization studies, the fluorescently labeled cells co-localized with E. piscicida in the intestines and kidneys of infected medaka larvae, confirming the ingestion of bacteria through phagocytosis. In addition, the labeled cells expressed macrophage markers but lacked a neutrophil marker. These results suggested that the fluorescently labeled cells of Tg[mpeg1.1:mCherry/mAG] medaka were monocytes/macrophages, which will be useful for future studies aimed at understanding the mechanisms of macrophage-mediated bacterial infections.

The evolutionary diversification and antimicrobial potential of MPEG1 in Metazoa

Macrophage-expressed gene 1 (MPEG1) is an ancient immune effector known to exist in Cnidaria, Mollusca, Actinopterygii, and Mammalia. In this study, we examined the evolution and antibacterial potential of MPEG1 across Metazoa. By unbiased data-mining, MPEG1 orthologs were found in 11 of 34 screened phyla. In invertebrates, MPEG1 is present in the major phyla and exhibits intensive duplication. In vertebrates, class-based clades were formed by the major, generic MPEG1 (gMPEG1) in each class. However, there is a minority of unique MPEG1 (uMPEG1) from 71 species of 4 classes that clustered into a separate clade detached from all major class-based clades. gMPEG1 and uMPEG1 exhibit strong genomic collinearity and are surrounded by high-density transposons. gMPEG1 and uMPEG1 transcript expressions were most abundant in immune organs, but differed markedly in tissue specificity. Systematic analysis identified an antimicrobial peptide (AMP)-like segment in the C-terminal (CT) tail of MPEG1. Peptides based on the AMP-like regions of 35 representative MPEG1 were synthesized. Bactericidal activities were displayed by all peptides. Together these results suggest transposon-propelled evolutionary diversification of MPEG1 in Metazoa that has likely led to functional specialisation. This study also reveals a possible antimicrobial mechanism mediated directly and solely by the CT tail of MPEG1.

Giant Triton Snail Charonia tritonis Macrophage-Expressed Gene 1 Protein Ct-Mpeg1: Molecular Identification, Expression Analysis, and Antimicrobial Activity

Macrophage-expressed gene 1 proteins (Mpeg1/Perforin-2 (PRF2)) are a family of pore-forming proteins (PFPs) which can form pores and destroy the cell membrane of invading pathogens. However, little information is available regarding the function of Mpeg1 in the giant triton snail Charonia tritonis. In this study, a homolog of Mpeg1 (Ct-Mpeg1) was identified in C. tritonis. The predicted protein of Ct-Mpeg1 contains several structural features known in Mpegs, including a membrane attack complex/perforin (MACPF) domain and single transmembrane region. The Ct-Mpeg1 gene was constitutively expressed in almost all tissues examined except in the proboscis, with the highest expression level observed in the mantle. As a typical pore-forming protein, Ct-Mpeg1 has antibacterial activities against Vibrio (including Vibrio alginolyticus and Vibrio parahaemolyticus). In addition, rCt-Mpeg1 challenge to V. alginolyticus represses the expression of most outer membrane protein synthesis-related genes and genes involved in the TCA cycle pathway, which will lead to reduced outer membrane protein synthesis and less energy capacity. This is the first report to characterize the macrophage-expressed gene 1 protein in C. tritonis, and these results suggest that macrophage-expressed gene 1 protein Ct-Mpeg1 is an important immune molecule of C. tritonis that is involved in the bacterial infection resistance of Vibrio, and this study may provide crucial basic data for the understanding of the innate immunity system of C. tritonis.

Linking erythropoietin to Treg-dependent allograft survival through myeloid cells.

Erythropoietin (EPO) has multiple nonerythropoietic functions, including immune modulation, but EPO’s effects in transplantation remain incompletely understood. We tested the mechanisms linking EPO administration to prolongation of murine heterotopic heart transplantation using WT and conditional EPO receptor–knockout (EPOR-knockout) mice as recipients. In WT controls, peritransplant administration of EPO synergized with CTLA4-Ig to prolong allograft survival (P < 0.001), reduce frequencies of donor-reactive effector CD8+ T cells in the spleen (P < 0.001) and in the graft (P < 0.05), and increase frequencies and total numbers of donor-reactive Tregs (P < 0.01 for each) versus CTLA4-Ig alone. Studies performed in conditional EPOR-knockout recipients showed that each of these differences required EPOR expression in myeloid cells but not in T cells. Analysis of mRNA isolated from spleen monocytes showed that EPO/EPOR ligation upregulated macrophage-expressed, antiinflammatory, regulatory, and pro-efferocytosis genes and downregulated selected proinflammatory genes. Taken together, the data support the conclusion that EPO promotes Treg-dependent murine cardiac allograft survival by crucially altering the phenotype and function of macrophages. Coupled with our previous documentation that EPO promotes Treg expansion in humans, the data support the need for testing the addition of EPO to costimulatory blockade-containing immunosuppression regimens in an effort to prolong human transplant survival.

Fragmentation of tissue-resident macrophages during isolation confounds analysis of single-cell preparations from mouse hematopoietic tissues.

Mouse hematopoietic tissues contain abundant tissue-resident macrophages that support immunity, hematopoiesis, and bone homeostasis. A systematic strategy to characterize macrophage subsets in mouse bone marrow (BM), spleen, and lymph node unexpectedly reveals that macrophage surface marker staining emanates from membrane-bound subcellular remnants associated with unrelated cells. Intact macrophages are not present within these cell preparations. The macrophage remnant binding profile reflects interactions between macrophages and other cell types invivo. Depletion of CD169+ macrophages invivo eliminates F4/80+ remnant attachment. Remnant-restricted macrophage-specific membrane markers, cytoplasmic fluorescent reporters, and mRNA are all detected in non-macrophage cells including isolated stem and progenitor cells. Analysis of RNA sequencing (RNA-seq) data, including publicly available datasets, indicates that macrophage fragmentation is a general phenomenon that confounds bulk and single-cell analysis of disaggregated hematopoietic tissues. Hematopoietic tissue macrophage fragmentation undermines the accuracy of macrophage exvivo molecular profiling and creates opportunity for misattribution of macrophage-expressed genes to non-macrophage cells.

Transient, flexible gene editing in zebrafish neutrophils and macrophages for determination of cell-autonomous functions.

ABSTRACTZebrafish are an important model for studying phagocyte function, but rigorous experimental systems to distinguish whether phagocyte-dependent effects are neutrophil or macrophage specific have been lacking. We have developed and validated transgenic lines that enable superior demonstration of cell-autonomous neutrophil and macrophage genetic requirements. We coupled well-characterized neutrophil- and macrophage-specific Gal4 driver lines with UAS:Cas9 transgenes for selective expression of Cas9 in either neutrophils or macrophages. Efficient gene editing, confirmed by both Sanger and next-generation sequencing, occurred in both lineages following microinjection of efficacious synthetic guide RNAs into zebrafish embryos. In proof-of-principle experiments, we demonstrated molecular and/or functional evidence of on-target gene editing for several genes (mCherry, lamin B receptor, trim33) in either neutrophils or macrophages as intended. These new UAS:Cas9 tools provide an improved resource for assessing individual contributions of neutrophil- and macrophage-expressed genes to the many physiological processes and diseases modelled in zebrafish. Furthermore, this gene-editing functionality can be exploited in any cell lineage for which a lineage-specific Gal4 driver is available.This article has an associated First Person interview with the first author of the paper.

Effects of microplastics (MPs) and tributyltin (TBT) alone and in combination on bile acids and gut microbiota crosstalk in mice

Microplastics (MPs) and tributyltin (TBT) are both potential environmental pollutants that enter organisms through the food chain and affect bodily functions. However, the effects and mechanisms of MPs and TBT exposure (especially the co-exposure of both pollutants) on mammals remain unclear. In this study, Ф5μm MPs (5MP) was administered alone or in combination with TBT to investigate the health risk of oral exposure in mice. All three treatments induced inflammation in the liver, altered gut microbiota composition and disturbed fecal bile acids profiles. In addition to decreasing triglyceride (TG) and increasing aspartate aminotransferase (AST) and macrophage-expressed gene 1 (Mpeg1), 5MP induced hepatic cholestasis by stimulating the expression of the cholesterol hydroxylase enzymes CYP8B1 and CYP27A1, and inhibiting multidrug resistance-associated protein 2 and 3 (MRP2, MRP3), and bile-salt export pump (BSEP) to prevent bile acids for entering the blood and bile. Correspondingly, 5MP treatment decreased 7-ketolithocholic acid (7-ketoLCA) and taurocholic acid (TCA), which were positively correlated with decreased Bacteroides and Marvinbryantia and negatively correlated with increased Bifidobacterium. In addition, TBT increased interferon γ (IFNγ) and Mpeg1 levels to induce inflammation, accompanied by decreased 7-ketoLCA, tauro-alpha-muricholic acid (T-alpha-MCA) and alpha-muricholic acid (alpha-MCA) levels, which were negatively related to Coriobacteriaceae_UCG-002 and Bifidobacterium. Co-exposure to 5MP and TBT also decreased TG and induced bile acids accumulation in the liver due to inhibited BSEP, which might be attributed to the co-regulation of decreased T-alpha-MCA and Harryflintia. In conclusion, the administration of 5MP and TBT alone and in combination could cause gut microbiome dysbiosis and subsequently alter bile acids profiles, while the combined exposure of 5MP and TBT weakened the toxic effects of 5MP and TBT alone.

Genome-wide investigation and expression analysis of MACPF gene family reveals its immune role in response to bacterial challenge of Manila clam

In this study, 18 MACPF genes (RpMACPF) were identified and classed into three types (Macrophage-expressed gene 1, Apextrin, and MACPF domain contain protein) based on gene structure and phylogenetic relationship in R. philippinarum. The length of RpMACPF proteins varied from 287 to 785 amino acids. The molecular weights and Theoretical PI values ranged from 3.2 kDa to 8.7 kDa and 4.7 to 8.6, respectively. RNA-seq data analysis revealed that 14 of 18 RpMACPF genes were highly expressed at the pediveliger larvae stage indicate RpMACPF might contribute to the early development and metamorphosis processes of the R. philippinarum. Besides, we found RpMACPF genes were significantly regulated by pathogen-associated molecular patterns (PAMPs) and Vibrio parahemolyticus, which indicates RpMACPF genes may play significant roles in response to invading pathogens. The results obtained in this work will provide valuable insight into the immune function of MACPF gene in R. philippinarum.

Distinct macrophage phenotypes and redox environment during the fin fold regenerative process in zebrafish.

In contrast to mammals, zebrafish (Danio rerio) has the ability to regenerate injured sites such as different tissues present in the fin. It is known that cells of the innate immune system play essential roles in regeneration; however, some aspects of the molecular mechanisms by which these cells orchestrate regeneration remain unknown. This study aimed to evaluate the infiltration dynamics of neutrophils and macrophages in the regenerative process of fin fold in regard to the influence of the redox environment and oxidative pathways. Fin fold amputation was performed on transgenic larvae for macrophage-expressed gene 1 (mpeg1), lysozyme (lyz), myeloperoxidase (mpo) and tumour necrosis factor alpha (TNFα) at 3days post-fertilization, followed by confocal microscopy imaging and measurement of the activities of oxidant and antioxidant enzymes. We observed initially an increase in the number of neutrophils (lyz:DsRed+/mpx:GFP+) and then macrophages (mpeg1+) in the injury site followed by a decrease in neutrophils at 7days post-amputation (dpa). Moreover, macrophages switch from a pro-inflammatory to an anti-inflammatory profile throughout the process, while the activity of superoxide dismutase (SOD) increased at 1 dpa and catalase (CAT) at 5 dpa. Higher levels of lipid peroxidation were also detected during regeneration. Despite oxidative stress, there is, therefore, an antioxidant response throughout the regeneration of the caudal fin. The present work can contribute to future studies on the development of cell therapies, achieving greater effectiveness in the treatment of diseases related to the formation of fibrotic tissue.

Ancient but Not Forgotten: New Insights Into MPEG1, a Macrophage Perforin-Like Immune Effector.

Macrophage-expressed gene 1 [MPEG1/Perforin-2 (PRF2)] is an ancient metazoan protein belonging to the Membrane Attack Complex/Perforin (MACPF) branch of the MACPF/Cholesterol Dependent Cytolysin (CDC) superfamily of pore-forming proteins (PFPs). MACPF/CDC proteins are a large and extremely diverse superfamily that forms large transmembrane aqueous channels in target membranes. In humans, MACPFs have known roles in immunity and development. Like perforin (PRF) and the membrane attack complex (MAC), MPEG1 is also postulated to perform a role in immunity. Indeed, bioinformatic studies suggest that gene duplications of MPEG1 likely gave rise to PRF and MAC components. Studies reveal partial or complete loss of MPEG1 causes an increased susceptibility to microbial infection in both cells and animals. To this end, MPEG1 expression is upregulated in response to proinflammatory signals such as tumor necrosis factor α (TNFα) and lipopolysaccharides (LPS). Furthermore, germline mutations in MPEG1 have been identified in connection with recurrent pulmonary mycobacterial infections in humans. Structural studies on MPEG1 revealed that it can form oligomeric pre-pores and pores. Strikingly, the unusual domain arrangement within the MPEG1 architecture suggests a novel mechanism of pore formation that may have evolved to guard against unwanted lysis of the host cell. Collectively, the available data suggest that MPEG1 likely functions as an intracellular pore-forming immune effector. Herein, we review the current understanding of MPEG1 evolution, regulation, and function. Furthermore, recent structural studies of MPEG1 are discussed, including the proposed mechanisms of action for MPEG1 bactericidal activity. Lastly limitations, outstanding questions, and implications of MPEG1 models are explored in the context of the broader literature and in light of newly available structural data.

Macrophage-specific protein perforin-2 is associated with poor neurological recovery and reduced survival after sudden cardiac arrest

BackgroundBiomarkers involved in inflammation and stress response were implicated in patients who were successfully resuscitated from out of hospital cardiac arrest (sR-OHCA). Here we report that macrophage-expressed gene, perforin-2, an evolutionarily conserved protein with membrane attack domain, is associated with poor neurological outcomes and mortality after sR-OHCA. ObjectivesTo examine the association between circulating perforin-2 protein measured within 6-h of sR-OHCA, mortality and neurological outcomes. MethodsWe prospectively enrolled 144 sR-OHCA patients from 4 different tertiary care centers. We measured perforin-2 and other conventional clinical biomarkers and compared between survivors vs. non-survivors. The neurological outcomes were dichotomized as poor or good according to the cereberal performance score. ResultsAt the end of the hospital stay, 45% of the patients had died and 46% had poor neurological outcomes. Serum perforin-2 levels were significantly higher in patients with poor neurological recovery, compared to the ones with good neurological recovery (ng/mL, 13.7 ± 45.9 vs. 1.2 ± 7.0, p = 0.01). There were no differences in other routinely measured biomarkers and left ventricular ejection fraction. On multivariate logistic regression, elevated perforin-2 (OR: 12.78, 95% CI: 1.0–17.8, p = 0.02), comatose on presentation (OR: 27.82, 95% CI: 0.2–19.5, p = 0.02) and non-shockable rhythm (OR: 17.04, 95% CI: 0.7–15.7, p = 0.01) were the significant predictors of poor neurological outcome. ConclusionsThis study reports a novel macrophage-expressed circulating biomarker perforin-2 to be strongly associated with reduced survival and poor neurological outcomes in sR-OHCA. These data can guide clinicians to prognosticate survival and neurological outcomes in sR-OHCA, and also form the basis for future therapeutic approaches.

Abstract 131: Elevated Serum MPEG-1 Protein is Associated With Reduced Survival and Poor Neurological Recovery in Resuscitated Out-Of-Hospital Sudden Cardiac Arrest

Background: For decades, bio-markers involved in inflammation and stress response were implicated in patients who were successfully resuscitated from out of hospital cardiac arrest (sR-OHCA). Here we report that increased serum levels of Macrophage-expressed gene 1 (MPEG1), an evolutionary conserved protein with membrane attack complex domain, is associated with increased mortality and poor neurological outcomes after sR-OHCA. Objectives: To examine the association between circulating MPEG-1 protein measured within the first 6-hours of sR-OHCA, and inpatient survival and neurological outcomes. Methods: We prospectively enrolled 144 sR-OHCA patients from 4 different tertiary care centers. We measured MPEG-1 protein, and other conventional clinical bio-markers including troponin-I and CK-MB in the sera obtained within the first 6-hours of resuscitation. The clinical covariates during hospitalization (till the time death or discharge) were obtained via medical record review. We compared MPEG-1 protein, troponin-I, and CK-MB between survivors vs . non-survivors, and neurological outcomes dichotomized as poor or good according to cerebral performance score (CPS). Results: At the end of the hospital stay, 47% of the patients had poor neurological status based on the CPS score of ≥ 4. Within these patients, 95% had inhospital mortality. The mean serum MPEG-1 levels were significantly higher in patients with poor neurological status, compared to the ones with good neurological recovery (ng/ml, 15.17 ± 48.04 vs 1.15 ± 6.73, p=0.017). There were no differences in other conventionally measured bio-markers, such as peak troponin , CK-MB or left ventricular ejection fraction. Elevated MPEG-1 of more than 8.8 ng/ml had significantly higher odds of predicting poor neurological outcome (OR: 8.37, 95% CI: 1.79-39.14, p=0.006). Conclusions: This study reports a novel macrophage-expressed circulating bio-marker strongly associated with reduced survival and poor neurological outcomes in sR-OHCA. These data can guide clinicians to prognosticate survival and neurological outcomes in sR-OHCA, and also form the basis for future therapeutic approaches.

The cryo-EM structure of the acid activatable pore-forming immune effector Macrophage-expressed gene 1

Macrophage-expressed gene 1 (MPEG1/Perforin-2) is a perforin-like protein that functions within the phagolysosome to damage engulfed microbes. MPEG1 is thought to form pores in target membranes, however, its mode of action remains unknown. We use cryo-Electron Microscopy (cryo-EM) to determine the 2.4 Å structure of a hexadecameric assembly of MPEG1 that displays the expected features of a soluble prepore complex. We further discover that MPEG1 prepore-like assemblies can be induced to perforate membranes through acidification, such as would occur within maturing phagolysosomes. We next solve the 3.6 Å cryo-EM structure of MPEG1 in complex with liposomes. These data reveal that a multi-vesicular body of 12 kDa (MVB12)-associated β-prism (MABP) domain binds membranes such that the pore-forming machinery of MPEG1 is oriented away from the bound membrane. This unexpected mechanism of membrane interaction suggests that MPEG1 remains bound to the phagolysosome membrane while simultaneously forming pores in engulfed bacterial targets.

Telmisartan inhibits Ang II-induced MMP-9 expression in macrophages in stabilizing atheromatous plaque.

To investigate the effects of telmisartan on matrix metalloproteinase-9 (MMP-9) expression in macrophages induced by angiotensin II (Ang II) and its mechanism. THP-1 cells were adopted for research, and phorbol-12-myristate-13-acetate (PMA) was utilized to induce THP-1 cells to be transformed into macrophages, with Ang II as a stimulating factor and telmisartan as a therapeutic drug. Cell counting kit-8 (CCK8) and lactate dehydrogenase (LDH) were applied to detect cell viability and toxicity. Enzyme-linked immunosorbent assay (ELISA) was performed to measure the MMP-9 release level. Polymerase Chain Reaction (PCR) and Western blotting were conducted to detect the expressions of MMP-9 messenger ribonucleic acid (mRNA) and protein, respectively. The mechanism of action was further studied, and the activity of cyclooxygenase-2 (COX2)/macrophage-expressed gene 1 (mPEG1) pathway was determined via PCR and Western blotting. The 1 mM Ang II could remarkably activate the synthesis and release of MMP-9 as well as the COX2/mPEG1 pathway in macrophages. However, telmisartan could effectively repress the Ang II-induced MMP-9 synthesis and release in the macrophages, and suppress the COX2/mPEG1 pathway in the macrophages activated by Ang II. Telmisartan can inhibit the activation of MMP-9 in the macrophages by suppressing the COX2/mPEG1 pathway.

C-type lectin 17A and macrophage-expressed receptor genes are magnified by fungal β-glucan after Vibrio parahaemolyticus infection in Totoaba macdonaldi cells

C-type lectins are a principal carbohydrate recognition mechanism as glucans on cell surfaces. This study identified and investigated molecular characterization and immune roles of a novel c-type lectin 17A from Totoaba macdonaldi (TmCLEC17A), which were described in head-kidney leukocytes after immunostimulation with fungal β-glucan 197A and Vibrio parahaemolyticus infection. This nucleotide sequence from totoaba was acquired using NGS and bioinformatics tools. Its full-length cDNA sequence consisted of 1128 bp (including the stop codon) and an open reading frame (ORF) of 771 bp encoding a 256 amino acid protein, 5´-UTR of 48 bp and 3´-UTR of 309 bp. The TmCLEC17A protein revealed a C-terminal-C-type lectin (CTL, also named carbohydrate-recognition domain, CRD), a N-terminal trans-membrane domain and a coiled coil motif, showing the highest similarity (80%) and identity (96%) with Larimichthys crocea. Fungal β-glucan 197A plus V. parahaemolyticus enhanced transcriptions of CLEC17A and TLR2 significantly besides the macrophage receptors, such as macrophage mannose receptor 1 and macrophage colony-stimulating factor 1 receptor 2. In addition, natural resistance-associated macrophage protein 2 was significantly up-regulated in leukocytes challenged with live V. parahaemolyticus. Overall, these results indicated that CLEC17A might be implicated in T. macdonaldi innate immunity as a pattern recognition receptor; fungal β-glucan 197A could stimulate cellular immune mechanisms in head-kidney leukocytes; and it could be used as potential immunostimulant in fish aquaculture.

Priming of innate antimycobacterial immunity by heat-killed Listeria monocytogenes induces sterilizing response in the adult zebrafish tuberculosis model.

ABSTRACTMycobacterium tuberculosis remains one of the most problematic infectious agents, owing to its highly developed mechanisms to evade host immune responses combined with the increasing emergence of antibiotic resistance. Host-directed therapies aiming to optimize immune responses to improve bacterial eradication or to limit excessive inflammation are a new strategy for the treatment of tuberculosis. In this study, we have established a zebrafish-Mycobacterium marinum natural host-pathogen model system to study induced protective immune responses in mycobacterial infection. We show that priming adult zebrafish with heat-killed Listeria monocytogenes (HKLm) at 1 day prior to M. marinum infection leads to significantly decreased mycobacterial loads in the infected zebrafish. Using rag1−/− fish, we show that the protective immunity conferred by HKLm priming can be induced through innate immunity alone. At 24 h post-infection, HKLm priming leads to a significant increase in the expression levels of macrophage-expressed gene 1 (mpeg1), tumor necrosis factor α (tnfa) and nitric oxide synthase 2b (nos2b), whereas superoxide dismutase 2 (sod2) expression is downregulated, implying that HKLm priming increases the number of macrophages and boosts intracellular killing mechanisms. The protective effects of HKLm are abolished when the injected material is pretreated with nucleases or proteinase K. Importantly, HKLm priming significantly increases the frequency of clearance of M. marinum infection by evoking sterilizing immunity (25 vs 3.7%, P=0.0021). In this study, immune priming is successfully used to induce sterilizing immunity against mycobacterial infection. This model provides a promising new platform for elucidating the mechanisms underlying sterilizing immunity and to develop host-directed treatment or prevention strategies against tuberculosis.This article has an associated First Person interview with the first author of the paper.

Identification of the macrophage-specific promoter signature in FANTOM5 mouse embryo developmental time course data.

The FANTOM5 consortium used cap analysis of gene expression (CAGE) to analyze the time course of gene expression over development from 11 days postcoitum (dpc) to adult in 16 developing organs and the whole body of the mouse. Every tissue in the body contains a large number of resident macrophages that initially infiltrate the embryo from the yolk sac. These cells contribute to organogenesis, and their functions diversify during development as they acquire tissue-specific adaptations. In each of the FANTOM5 time courses, the expression of known macrophage-specific genes, including CSF1 receptor (Csf1r), epidermal growth factor-like module-containing mucin-like hormone receptor-like 1 (Emr1), and mer receptor tyrosine kinase (Mertk), was readily detectable and increased with time. We reasoned that genes expressed by macrophages would be strongly correlated in their expression with these known markers and might vary between tissues. We used the network analysis tool, Miru, to extract the sets of coexpressed genes from the time course and identified a core set of coexpressed genes attributable to embryonic macrophages, including some, such as dehydrogenase/reductase 3 (Dhrs3), that may have unique functions in development. The FANTOM5 data also detected the appearance of tissue-specific macrophage-expressed genes, such as T cell Ig and mucin domain-containing 4 (Timd4) and V-set and Ig domain-containing 4 (Vsig4) in liver and sialic acid-binding Ig-like lectin 5 (Siglec5) in lung, and confirmed that macrophage content increases with time in each organ as the proliferative phases end, and tissue-specific gene-expression increases. The FANTOM5 data are available on a comprehensive browser (http://fantom.gsc.riken.jp/zenbu/), which provides a resource for the study of macrophage transcriptional regulation and roles in mouse development.