Monocercomonoides exilis is a model species of the amitochondrial eukaryotic group Oxymonadida, which makes it a suitable organism for studying the consequences of mitochondrial loss. Although M. exilis has an endobiotic lifestyle, it can be cultured in vitro in polyxenic conditions alongside an uncharacterized prokaryotic community, while attempts to create axenic cultures have not been successful. In this study, we used metagenomic sequencing, transcriptomics, and metabolomics to characterize the microbial consortium that supports the growth of M. exilis. We assembled genomes for 24 bacterial species and identified at least 30 species in total. M. exilis accounted for less than 1.5% of the DNA reads, while bacterial species dominated the sequence data and shifted in abundance over time. Our metabolic reconstruction and differential gene expression analyses show that the bacterial community relies on organic carbon oxidation, fermentation, and hydrogen production, but does not engage in methanogenesis. We observed rapid depletion of amino acids, nucleotides, glyceraldehyde, lactate, fatty acids, and alcohols in the medium, indicating a reliance on external nutrient recycling. The nitrogen cycle in this community is incomplete, with limited nitrogen fixation and no ammonia oxidation. Despite detailed metabolic profiling, we did not find any direct biochemical connections between M. exilis and the prokaryotes. Several bacterial species produce siderophores to assist themselves and others in the community in acquiring iron. However, M. exilis does not appear to benefit directly from siderophore-mediated iron transport and lacks known iron uptake pathways. This indicates that M. exilis may rely indirectly on the iron metabolism of other bacteria through phagocytosis. Additionally, some bacteria synthesize polyamines like spermidine and phosphatidylcholine, which M. exilis may need but cannot produce on its own. As the culture ages, M. exilis shows changes in gene expression consistent with starvation responses, including the upregulation of carbohydrate storage pathways and processes related to exocytosis. These findings provide new insights into microbial interactions within xenic cultures and emphasize the complex nature of maintaining amitochondriate eukaryotes in vitro.

• Publication type
• Journal Article MeSH

The visual cycle is an important pathway in the retinal pigment epithelium (RPE) which regenerates 11-cis retinal chromophore for the retinal photoreceptors. The central enzyme in the visual cycle is RPE65 retinol isomerase. Expression of RPE65 mRNA and protein levels are significantly lower in RPE cell culture models when compared to native RPE. This limits the use of these models to study the visual cycle. To determine the main drivers of RPE65 regulation we compared the transcriptional profiles of native and cell culture models of RPE with various levels of RPE65 expression. We also compared the levels of RPE65 expression between ARPE-19 cells grown in media supplemented with 1 mM pyruvate (PYR) or 10 mM nicotinamide (NAM). In addition, we performed experiments directed at transcriptional and translational regulation of RPE65. We show that RPE65 mRNA and protein expression is significantly higher in NAM media grown cells than PYR cells. Transfection of cells with a variety of different vectors containing RPE65 ORFs with different promoters, codon optimization, IRES, 3' UTRs, suggest that translational effects are less important than transcriptional status. Importantly, we found that feeding with rod outer segments (ROS) decreases RPE65 expression in NAM grown cells, suggesting that certain primary functions of the RPE (here, visual cycle and phagocytosis) are not positively linked. Analysis of differentially regulated microRNAs (miRs) provides a basis for this downregulation. It appears that the regulation of RPE65 expression in ARPE-19 cells, in particular, is multifactorial, involving primarily metabolic and transcriptional status of the cells, with translation of RPE65 mRNA playing a smaller role.

• Keywords
• MicroRNAs, Nicotinamide, Pyruvate, RPE65, Retina, Retinal pigment epithelium, Ribosome, Transcription, Translation,
• MeSH
• Cell Line MeSH
• cis-trans-Isomerases * genetics   metabolism   MeSH
• Humans MeSH
• RNA, Messenger genetics   metabolism   MeSH
• Gene Expression Regulation * MeSH
• Retinal Pigment Epithelium * metabolism   cytology   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• cis-trans-Isomerases * MeSH
• RNA, Messenger MeSH
• retinoid isomerohydrolase MeSH Browser

BACKGROUND: Trauma is a leading global cause of mortality, and systemic inflammatory response syndrome (SIRS) remains a significant complication, contributing to adverse outcomes. Neutrophils, as first responders to tissue injury, undergo substantial phenotypic and functional changes following trauma. This study investigates neutrophil subpopulations defined by CD16 and CD62L expression in trauma patients, focusing on their correlation with clinical biomarkers, trauma severity, and functional properties. METHODS: We included 50 non-infectious trauma patients, categorized into SIRS and Non-SIRS groups, and 43 elective surgery patients as controls. Neutrophil subsets were analyzed at two time points (TP1 and TP2) using flow cytometry. Functional assays evaluated phagocytosis, oxidative burst, mitochondrial function, and degranulation. Correlations between neutrophil subpopulations and clinical markers, including lactate, creatine kinase, Injury Severity Score, and Trauma and Injury Severity Score, were examined. RESULTS: Patients with SIRS exhibited higher proportions of banded neutrophils and CD16lowCD62Llow neutrophils at TP1, alongside reduced levels of mature neutrophils. Elevated lactate and creatine kinase levels positively correlated with banded neutrophils and CD16lowCD62Llow neutrophils, while negatively correlating with mature neutrophils CD16highCD62Lhigh and hypersegmented neutrophils CD16highCD62Llow. Hypersegmented neutrophils were more prevalent in Non-SIRS patients at TP1 and in SIRS patients at TP2. Banded neutrophils showed a positive correlation with Injury Severity Score and an inverse correlation with Trauma and Injury Severity Score (TRISS), whereas hypersegmented neutrophils were negatively associated with ISS and positively correlated with TRISS. These correlations likely reflect the pro-inflammatory role of banded neutrophils and the inflammation-resolving function of hypersegmented neutrophils. CD16lowCD62Llow neutrophils displayed impaired phagocytosis, oxidative burst, and degranulation capacity, indicating functional deficiencies. CONCLUSION: This study highlights the dynamic changes in neutrophil subpopulations in trauma and their association with systemic inflammation and clinical severity. Increased banded neutrophils correlate with SIRS and metabolic stress, whereas hypersegmented neutrophils may contribute to resolving inflammation. CD16lowCD62Llow neutrophils exhibit functional impairments, warranting further investigation. Monitoring neutrophil subpopulations could aid in identifying trauma patients at risk for non-infectious SIRS and guide therapeutic interventions.

• Keywords
• ISS, SIRS, TRISS, creatine kinase, lactate, neutrophils, trauma,
• MeSH
• Biomarkers MeSH
• Adult MeSH
• Phagocytosis MeSH
• L-Selectin metabolism   MeSH
• Middle Aged MeSH
• Humans MeSH
• Neutrophils * immunology   metabolism   MeSH
• Wounds and Injuries * immunology   blood   MeSH
• Receptors, IgG metabolism   MeSH
• Respiratory Burst MeSH
• Aged MeSH
• Systemic Inflammatory Response Syndrome * immunology   etiology   blood   MeSH
• Check Tag
• Adult MeSH
• Middle Aged MeSH
• Humans MeSH
• Male MeSH
• Aged MeSH
• Female MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Biomarkers MeSH
• L-Selectin MeSH
• Receptors, IgG MeSH

Efficient treatment of osteomyelitis caused by Staphylococcus aureus is a great clinical challenge due to bacterial resistance and immune evasion issues. Macrophages play a crucial role in the fight against S. aureus but suffer from deficiencies in function in the infectious milieu leading to persistent infection. Here, a strategy of exploiting aged neutrophil membrane (aNM) is developed to camouflage 2D MnPSe3 nanosheets (MPS NSs), denoted as aNM@MPS, to mediate in situ macrophage engineering, thereby potentiating macrophages to eradicate refractory osteomyelitis. When administered systematically, the biofunctional aNM@MPS ensures selectivity for osteomyelitis lesions, enhanced bone marrow retention, and subsequent phagocytosis by macrophages. In the mouse model of osteomyelitis, the aNM@MPS enables dysfunctional macrophages to digest intracellular bacteria by generating highly toxic hydroxyl radicals and sequentially reprogramming bactericidal immunity through manganese ion-mediated immune activation, which synergistically terminates persistent infection-initiated pathological cascades and subsequently reestablish host-directed bactericidal potency, thereby conferring a satisfactory osteoprotective effect. These findings demonstrate that macrophages in the skeletal infectious milieu can be precisely remodeled via the lesion-macrophage dual-targeting metalloimmunotherapy strategy, which holds potential for osteomyelitis treatment.

• Keywords
• biofunctional nanosheets, macrophage engineering, metalloimmunotherapy, osteomyelitis, phagolysosomal killing,
• MeSH
• Anti-Bacterial Agents pharmacology   chemistry   MeSH
• Phagocytosis drug effects   MeSH
• Humans MeSH
• Macrophages * metabolism   drug effects   MeSH
• Manganese * chemistry   pharmacology   MeSH
• Disease Models, Animal MeSH
• Mice, Inbred C57BL MeSH
• Mice MeSH
• Nanostructures * chemistry   MeSH
• Neutrophils metabolism   MeSH
• Osteomyelitis * drug therapy   microbiology   pathology   therapy   MeSH
• RAW 264.7 Cells MeSH
• Staphylococcal Infections * drug therapy   MeSH
• Staphylococcus aureus drug effects   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Anti-Bacterial Agents MeSH
• Manganese * MeSH

Exposure to air pollution is associated with neurological diseases. Traffic is a major source of air pollution, consisting of a complex mixture of ultrafine particles, that can invade the brain and induce a microglia-mediated inflammatory response. However, the exact mechanisms of how traffic-related particles impact human microglia remain poorly understood. This study investigates the effects of diesel exhaust particles (DEPs) on human induced pluripotent stem cell-derived microglia-like cells (iMGL). We exposed iMGLs to three different DEPs and studied the impact on the iMGL transcriptome and functionality, focusing on cytokine secretion, mitochondrial respiration, lysosomal function, and phagocytosis. A20 particles were collected from a heavy-duty engine run with petroleum diesel. For A0, the same engine was run with renewable diesel. E6 was produced with a modern 2019 model diesel passenger car run with renewable diesel. RNAseq revealed activation of the cytokine storm pathway and inhibition of the autophagy pathway in iMGLs after exposure to particles derived from older diesel emission technology (A20, A0). Particles from the modern diesel engine technology (E6) did not alter microglial transcriptome after 24 h exposure. A20 and A0 exposure led to impaired lysosomal functions in iMGLs. In contrast, E6 did not cause major alterations in microglia functions. In addition, we show that response to particles is more pronounced in human iMGLs compared to mouse primary microglia. To conclude, particles from older emission technology impair phago-lysosomal functions of iMGLs, but modern alternatives with filtration do not induce drastic changes in the functionality of iMGLs.

• Keywords
• Air pollution, Diesel, Human microglia, Lysosome, iPSC,
• MeSH
• Induced Pluripotent Stem Cells MeSH
• Air Pollutants * toxicity   MeSH
• Humans MeSH
• Lysosomes * drug effects   MeSH
• Microglia * drug effects   MeSH
• Particulate Matter * toxicity   MeSH
• Vehicle Emissions * toxicity   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Air Pollutants * MeSH
• Particulate Matter * MeSH
• Vehicle Emissions * MeSH

Bordetella pertussis infects human upper airways and deploys an array of immunosuppressive virulence factors, among which the adenylate cyclase toxin (CyaA) plays a prominent role in disarming host phagocytes. CyaA binds the complement receptor-3 (CR3 aka αMβ2 integrin CD11b/CD18 or Mac-1) of myeloid cells and delivers into their cytosol an adenylyl cyclase enzyme that hijacks cellular signaling through unregulated conversion of cytosolic ATP to cAMP. We found that the action of as little CyaA as 22 pM (4 ng/mL) blocks macrophage colony-stimulating factor (M-CSF)-driven transition of migratory human CD14+ monocytes into macrophages. Global transcriptional profiling (RNAseq) revealed that exposure of monocytes to 22 pM CyaA for 40 hours in culture with 20 ng/mL of M-CSF led to upregulation of genes that exert negative control of monocyte to macrophage differentiation (e.g., SERPINB2, DLL1, and CSNK1E). The sustained CyaA action yielded downregulation of numerous genes involved in processes crucial for host defense, such as myeloid cell differentiation, chemotaxis of inflammatory cells, antigen presentation, phagocytosis, and bactericidal activities. CyaA-elicited signaling also promoted deacetylation and trimethylation of lysines 9 and 27 of histone 3 (H3K9me3 and H3K27me3) and triggered the formation of transcriptionally repressive heterochromatin patches in the nuclei of CyaA-exposed monocytes. These effects were partly reversed by the G9a methyltransferase inhibitor UNC 0631 and by the pleiotropic HDAC inhibitor Trichostatin-A, revealing that CyaA-elicited epigenetic alterations mediate transcriptional reprogramming of monocytes and play a role in CyaA-triggered block of monocyte differentiation into bactericidal macrophage cells.IMPORTANCETo proliferate on host airway mucosa and evade elimination by patrolling sentinel cells, the whooping cough agent Bordetella pertussis produces a potently immunosubversive adenylate cyclase toxin (CyaA) that blocks opsonophagocytic killing of bacteria by phagocytes like neutrophils and macrophages. Indeed, chemotactic migration of CD14+ monocytes to the infection site and their transition into bactericidal macrophages, thus replenishing the exhausted mucosa-patrolling macrophages, represents one of the key mechanisms of innate immune defense to infection. We show that the cAMP signaling action of CyaA already at a very low toxin concentration triggers massive transcriptional reprogramming of monocytes that is accompanied by chromatin remodeling and epigenetic histone modifications, which block the transition of migratory monocytes into bactericidal macrophage cells. This reveals a novel layer of toxin action-mediated hijacking of functional differentiation of innate immune cells for the sake of mucosal pathogen proliferation and transmission to new hosts.

• Keywords
• Bordetella pertussis, RTX toxins, cyclic AMP, differentiation, epigenetics, macrophages, monocytes,
• MeSH
• Adenylate Cyclase Toxin * metabolism   MeSH
• Bordetella pertussis * pathogenicity   enzymology   MeSH
• Cell Differentiation * drug effects   MeSH
• Macrophage Colony-Stimulating Factor MeSH
• Cells, Cultured MeSH
• Humans MeSH
• Macrophages * drug effects   cytology   MeSH
• Monocytes * drug effects   cytology   physiology   MeSH
• Cellular Reprogramming * MeSH
• Chromatin Assembly and Disassembly * drug effects   MeSH
• Signal Transduction MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Adenylate Cyclase Toxin * MeSH
• Macrophage Colony-Stimulating Factor MeSH

Prostate cancer (PCa) ranks as the second leading cause of cancer-related deaths among men in the United States. Prostate-specific membrane antigen (PSMA) represents a well-established biomarker of PCa, and its levels correlate positively with the disease progression, culminating at the stage of metastatic castration-resistant prostate cancer. Due to its tissue-specific expression and cell surface localization, PSMA shows superior potential for precise imaging and therapy of PCa. Antibody-based immunotherapy targeting PSMA offers the promise of selectively engaging the host immune system with minimal off-target effects. Here we report on the design, expression, purification, and characterization of a bispecific engager, termed 5D3-CP33, that efficiently recruits macrophages to the vicinity of PSMA-positive cancer cells mediating PCa death. The engager was engineered by fusing the anti-PSMA 5D3 antibody fragment to a cyclic peptide 33 (CP33), selectively binding the Fc gamma receptor I (FcγRI/CD64) on the surface of phagocytes. Functional parts of the 5D3-CP33 engager revealed a nanomolar affinity for PSMA and FcγRI/CD64 with dissociation constants of KD = 3 nM and KD = 140 nM, respectively. At a concentration as low as 0.3 nM, the engager was found to trigger the production of reactive oxygen species by U937 monocytic cells in the presence of PSMA-positive cells. Moreover, flow cytometry analysis demonstrated antibody-dependent cell-mediated phagocytosis of PSMA-positive cancer cells by U937 monocytes when exposed to 0.15 nM 5D3-CP33. Our findings illustrate that 5D3-CP33 effectively and specifically activates monocytes upon PSMA-positive target engagement, resulting in the elimination of tumor cells. The 5D3-CP33 engager can thus serve as a promising lead for developing new immunotherapy tools for the efficient treatment of PCa.

• MeSH
• Antigens, Surface * immunology   metabolism   MeSH
• Glutamate Carboxypeptidase II * immunology   metabolism   MeSH
• Immunotherapy methods   MeSH
• Humans MeSH
• Macrophages immunology   MeSH
• Monocytes * immunology   metabolism   MeSH
• Cell Line, Tumor MeSH
• Prostatic Neoplasms * immunology   therapy   pathology   MeSH
• Antibodies, Bispecific * immunology   pharmacology   MeSH
• Receptors, IgG metabolism   immunology   MeSH
• Check Tag
• Humans MeSH
• Male MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Antigens, Surface * MeSH
• FOLH1 protein, human MeSH Browser
• Glutamate Carboxypeptidase II * MeSH
• Antibodies, Bispecific * MeSH
• Receptors, IgG MeSH

In the current study, the effects of dietary fulvic acid supplementation at levels of 0.5, 1 and 2% were examined in white-leg shrimp, Litopenaeus vannamei. A significant increase in the weight of the shrimp was observed in the group treated with 2% fulvic acid in comparison to the control group. This may have been associated with an increased digestive efficiency, with the food conversion ratio reducing from 2.4 to 1.9, and increased hepatopancreatic amylase, protease, and lipase enzyme activities. Enhanced activity of hemolymph superoxide dismutase was suggestive of an enhanced immune capacity, while hemolymph cell count increased by 16.4 and 13.6% in shrimp receiving diets supplemented with 1 and 2% fulvic acid, respectively. Additionally, the number of large granular cells increased by 37.3% and 40.8% relative to the control in these two groups. Furthermore, the lysozyme activity increased in shrimp receiving dietary supplementation of 1% and 2% fulvic acid by 16.7% and 24.7%, respectively. Phenol oxidase activity, which activates phagocytosis and encapsulation of invading pathogens, increased in all groups supplemented with fulvic acid, with the highest activity in the 1% fulvic acid group. Overall the present results suggest that fulvic acid is a promising feed additive for white-leg shrimp super-intensive culture.

• Keywords
• Fulvic acid, Growth performance, Humic substances, Immunostimulants, Pacific white shrimp,
• MeSH
• Antioxidants * metabolism   pharmacology   MeSH
• Benzopyrans * pharmacology   MeSH
• Hemolymph metabolism   drug effects   MeSH
• Animal Feed analysis   MeSH
• Penaeidae * drug effects   immunology   metabolism   MeSH
• Dietary Supplements MeSH
• Digestion drug effects   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Antioxidants * MeSH
• Benzopyrans * MeSH
• fulvic acid MeSH Browser

Triggering receptor expressed on myeloid cells 2 (TREM-2), a glycosylated receptor belonging to the immunoglobin superfamily and especially expressed in the myeloid cell lineage, is frequently explained as a reminiscent receptor for both adaptive and innate immunity regulation. TREM-2 is also acknowledged to influence NK cell differentiation via the PI3K and PLCγ signaling pathways, as well as the partial activation or direct inhibition of T cells. Additionally, TREM-2 overexpression is substantially linked to cell-specific functions, such as enhanced phagocytosis, reduced toll-like receptor (TLR)-mediated inflammatory cytokine production, increased transcription of anti-inflammatory cytokines, and reshaped T cell function. Whereas TREM-2-deficient cells exhibit diminished phagocytic function and enhanced proinflammatory cytokines production, proceeding to inflammatory injuries and an immunosuppressive environment for disease progression. Despite the growing literature supporting TREM-2+ cells in various diseases, such as neurodegenerative disorders and cancer, substantial facets of TREM-2-mediated signaling remain inadequately understood relevant to pathophysiology conditions. In this direction, herein, we have summarized the current knowledge on TREM-2 biology and cell-specific TREM-2 expression, particularly in the modulation of pivotal TREM-2-dependent functions under physiopathological conditions. Furthermore, molecular regulation and generic biological relevance of TREM-2 are also discussed, which might provide an alternative approach for preventing or reducing TREM-2-associated deformities. At last, we discussed the TREM-2 function in supporting an immunosuppressive cancer environment and as a potential drug target for cancer immunotherapy. Hence, summarized knowledge of TREM-2 might provide a window to overcome challenges in clinically effective therapies for TREM-2-induced diseases in humans.

• Keywords
• Cancer, Cell signaling, Cytokines, Immunotherapy, Inflammation,
• MeSH
• Killer Cells, Natural immunology   MeSH
• Phagocytosis MeSH
• Humans MeSH
• Membrane Glycoproteins * metabolism   genetics   MeSH
• Neoplasms immunology   metabolism   genetics   MeSH
• Immunity, Innate MeSH
• Receptors, Immunologic * metabolism   genetics   MeSH
• Signal Transduction MeSH
• T-Lymphocytes immunology   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Review MeSH
• Names of Substances
• Membrane Glycoproteins * MeSH
• Receptors, Immunologic * MeSH
• TREM2 protein, human MeSH Browser

INTRODUCTION: Red blood cells (RBCs), also known as erythrocytes, are underestimated in their role in the immune system. In mammals, erythrocytes undergo maturation that involves the loss of nuclei, resulting in limited transcription and protein synthesis capabilities. However, the nucleated nature of non-mammalian RBCs is challenging this conventional understanding of RBCs. Notably, in bony fishes, research indicates that RBCs are not only susceptible to pathogen attacks but express immune receptors and effector molecules. However, given the abundance of RBCs and their interaction with every physiological system, we postulate that they act in surveillance as sentinels, rapid responders, and messengers. METHODS: We performed a series of in vitro experiments with Cyprinus carpio RBCs exposed to Aeromonas hydrophila, as well as in vivo laboratory infections using different concentrations of bacteria. RESULTS: qPCR revealed that RBCs express genes of several inflammatory cytokines. Using cyprinid-specific antibodies, we confirmed that RBCs secreted tumor necrosis factor alpha (TNFα) and interferon gamma (IFNγ). In contrast to these indirect immune mechanisms, we observed that RBCs produce reactive oxygen species and, through transmission electron and confocal microscopy, that RBCs can engulf particles. Finally, RBCs expressed and upregulated several putative toll-like receptors, including tlr4 and tlr9, in response to A. hydrophila infection in vivo. DISCUSSION: Overall, the RBC repertoire of pattern recognition receptors, their secretion of effector molecules, and their swift response make them immune sentinels capable of rapidly detecting and signaling the presence of foreign pathogens. By studying the interaction between a bacterium and erythrocytes, we provide novel insights into how the latter may contribute to overall innate and adaptive immune responses of teleost fishes.

• Keywords
• Aeromonas hydrophila (A. hydrophila), Cyprinus carpio, bacteria, cytokines, engulfment, inflammation, red blood cell (RBC), teleost fish,
• MeSH
• Aeromonas hydrophila * immunology   MeSH
• Cytokines * metabolism   immunology   MeSH
• Erythrocytes * immunology   metabolism   MeSH
• Phagocytosis immunology   MeSH
• Gram-Negative Bacterial Infections * immunology   MeSH
• Carps * immunology   microbiology   MeSH
• Fish Diseases * immunology   microbiology   MeSH
• Pathogen-Associated Molecular Pattern Molecules immunology   MeSH
• Immunity, Innate MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Cytokines * MeSH
• Pathogen-Associated Molecular Pattern Molecules MeSH