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Science Immunology Jun 2024Immune cells have intensely physical lifestyles characterized by structural plasticity and force exertion. To investigate whether specific immune functions require...
Immune cells have intensely physical lifestyles characterized by structural plasticity and force exertion. To investigate whether specific immune functions require stereotyped mechanical outputs, we used super-resolution traction force microscopy to compare the immune synapses formed by cytotoxic T cells with contacts formed by other T cell subsets and by macrophages. T cell synapses were globally compressive, which was fundamentally different from the pulling and pinching associated with macrophage phagocytosis. Spectral decomposition of force exertion patterns from each cell type linked cytotoxicity to compressive strength, local protrusiveness, and the induction of complex, asymmetric topography. These features were validated as cytotoxic drivers by genetic disruption of cytoskeletal regulators, live imaging of synaptic secretion, and in silico analysis of interfacial distortion. Synapse architecture and force exertion were sensitive to target stiffness and size, suggesting that the mechanical potentiation of killing is biophysically adaptive. We conclude that cellular cytotoxicity and, by implication, other effector responses are supported by specialized patterns of efferent force.
Topics: Animals; Immunological Synapses; Single-Cell Analysis; Mice; T-Lymphocytes, Cytotoxic; Biomechanical Phenomena; Cytotoxicity, Immunologic; Macrophages; Mice, Inbred C57BL
PubMed: 38941478
DOI: 10.1126/sciimmunol.adj2898 -
Journal of Integrative Neuroscience Jun 2024Perioperative neurocognitive disorders (PND) are a group of prevalent neurological complications that often occur in elderly individuals following major or emergency...
Differentially Expressed Proteins in the Serum of Elderly Patients Who Experienced Perioperative Neurocognitive Disorders Following Transurethral Resection of the Prostate.
OBJECTIVE
Perioperative neurocognitive disorders (PND) are a group of prevalent neurological complications that often occur in elderly individuals following major or emergency surgical procedures. The etiologies are not fully understood. This study endeavored to investigate novel targets and prediction methods for the occurrence of PND.
METHODS
A total of 229 elderly patients diagnosed with prostatic hyperplasia who underwent transurethral resection of the prostate (TURP) combined with spinal cord and epidural analgesia were included in this study. The patients were divided into two groups, the PND group and non-PND group, based on the Z-score method. According to the principle of maintaining consistency between preoperative and intraoperative conditions, three patients from each group were randomly chosen for serum sample collection. isobaric tags for relative and absolute quantification (iTRAQ) proteomics technology was employed to analyze and identify the proteins that exhibited differential expression in the serum samples from the two groups. Bioinformatics analysis was performed on the proteins that exhibited differential expression.
RESULTS
Among the 1101 serum proteins analyzed in the PND and non-PND groups, eight differentially expressed proteins were identified in PND patients. Of these, six proteins showed up-regulation, while two proteins showed down-regulation. Further bioinformatics analysis of the proteins that exhibited differential expression revealed their predominant involvement in cellular biological processes, cellular component formation, as well as endocytosis and phagocytosis Additionally, these proteins were found to possess the RING domain of E3 ubiquitin ligase.
CONCLUSION
The iTRAQ proteomics technique was employed to analyze the variation in protein expression in serum samples from patients with PND and those without PND. This study successfully identified eight proteins that exhibited differential expression levels between the two groups. Bioinformatics analysis indicates that proteins exhibiting differential expression are primarily implicated in the biological processes associated with microtubules. Investigating the microtubule formation process as it relates to neuroplasticity and synaptic formation may offer valuable insights for enhancing our comprehension and potential prevention of PND.
CLINICAL TRIAL REGISTRATION
Registered (ChiCTR2000028836). Date (20190306).
Topics: Humans; Male; Aged; Transurethral Resection of Prostate; Proteomics; Prostatic Hyperplasia; Neurocognitive Disorders; Postoperative Cognitive Complications; Perioperative Period; Aged, 80 and over; Blood Proteins; Computational Biology
PubMed: 38940081
DOI: 10.31083/j.jin2306123 -
Frontiers in Cell and Developmental... 2024Zebrafish possess the ability to regenerate dying neurons in response to retinal injury, with both Müller glia and microglia playing integral roles in this response....
Zebrafish possess the ability to regenerate dying neurons in response to retinal injury, with both Müller glia and microglia playing integral roles in this response. Resident Müller glia respond to damage by reprogramming and undergoing an asymmetric cell division to generate a neuronal progenitor cell, which continues to proliferate and differentiate into the lost neurons. In contrast, microglia become reactive, phagocytose dying cells, and release inflammatory signals into the surrounding tissue following damage. In recent years, there has been increased attention on elucidating the role that microglia play in regulating retinal regeneration. Here we demonstrate that inflammatory cytokines are differentially expressed during retinal regeneration, with the expression of a subset of pro-inflammatory cytokine genes upregulated shortly after light damage and the expression of a different subset of cytokine genes subsequently increasing. We demonstrate that both cytokine IL-1β and IL-10 are essential for Müller glia proliferation in the light-damaged retina. While IL-1β is sufficient to induce Müller glia proliferation in an undamaged retina, expression of IL-10 in undamaged retinas only induces Müller glia to express gliotic markers. Together, these findings demonstrate the essential role of inflammatory cytokines IL-1β and IL-10 on Müller glia proliferation following light damage in adult zebrafish.
PubMed: 38938553
DOI: 10.3389/fcell.2024.1406330 -
Journal of Translational Medicine Jun 2024Monocyte-derived alveolar macrophages (Mo_AMs) are increasingly recognised as potential pathogenic factors for idiopathic pulmonary fibrosis (IPF). While scRNAseq...
BACKGROUND
Monocyte-derived alveolar macrophages (Mo_AMs) are increasingly recognised as potential pathogenic factors for idiopathic pulmonary fibrosis (IPF). While scRNAseq analysis has proven valuable in the transcriptome profiling of Mo_AMs, the integration analysis of multi-omics may provide additional dimensions of understanding of these cellular populations.
METHODS
We performed multi-omics analysis on 116 scRNAseq, 119 bulkseq and five scATACseq lung tissue samples from IPF. We built a large-scale IPF scRNAseq atlas and conducted the Monocle 2/3 as well as the Cellchat to explore the developmental path and intercellular communication on Mo_AMs. We also reported the difference in metabolisms, tissue repair and phagocytosis between Mo_AMs and tissue-resident alveolar macrophages (TRMs). To determine whether Mo_AMs affected pulmonary function, we projected clinical phenotypes (FVC%pred) from the bulkseq dataset onto the scRNAseq atlas. Finally, we used scATATCseq to uncover the upstream regulatory mechanisms and determine key drivers in Mo_AMs.
RESULTS
We identified three Mo_AMs clusters and the trajectory analysis further validated the origin of these clusters. Moreover, via the Cellchat analysis, the CXCL12/CXCR4 axis was found to be involved in the molecular basis of reciprocal interactions between Mo_AMs and fibroblasts through the activation of the ERK pathway in Mo_AMs. SPP1_RecMacs (RecMacs, recruited macrophages) were higher in the low-FVC group than in the high-FVC group. Specifically, compared with TRMs, the functions of lipid and energetic metabolism as well as tissue repair were higher in Mo_AMs than TRMs. But, TRMs may have higher level of phagocytosis than TRMs. SPIB (PU.1), JUNB, JUND, BACH2, FOSL2, and SMARCC1 showed stronger association with open chromatin of Mo_AMs than TRMs. Significant upregulated expression and deep chromatin accessibility of APOE were observed in both SPP1_RecMacs and TRMs.
CONCLUSION
Through trajectory analysis, it was confirmed that SPP1_RecMacs derived from Monocytes. Besides, Mo_AMs may influence FVC% pred and aggravate pulmonary fibrosis through the communication with fibroblasts. Furthermore, distinctive transcriptional regulators between Mo_AMs and TRMs implied that they may depend on different upstream regulatory mechanisms. Overall, this work provides a global overview of how Mo_AMs govern IPF and also helps determine better approaches and intervention therapies.
Topics: Humans; Idiopathic Pulmonary Fibrosis; Macrophages, Alveolar; Monocytes; Male; Gene Expression Profiling; Female; Receptors, CXCR4; Middle Aged; Phenotype; Lung; Gene Expression Regulation
PubMed: 38937806
DOI: 10.1186/s12967-024-05398-y -
Cell Communication and Signaling : CCS Jun 2024Tumor cells release extracellular vesicles (EVs) that contribute to the polarization of macrophages towards tumor-associated macrophages (TAMs). High expression levels...
BACKGROUND
Tumor cells release extracellular vesicles (EVs) that contribute to the polarization of macrophages towards tumor-associated macrophages (TAMs). High expression levels of the RNA binding protein IGF2BP2/IMP2 are correlated with increased tumor cell proliferation, invasion, and poor prognosis in the clinic. However, there is a lack of understanding of whether IMP2 affects the cargo of cancer cell-derived EVs, thereby modulating macrophage polarization.
METHODS
EVs were isolated from IMP2-expressing HCT116 parental cells (WT) and CRISPR/Cas9 IMP2 knockout (KO) cells. EVs were characterized according to MISEV guidelines, microRNA cargo was assessed by microRNA-Seq, and the protein cargo was analyzed by proteomics. Primary human monocyte-derived macrophages (HMDMs) were polarized by EVs, and the expression of genes and surface markers was assessed using qPCR and flow cytometry, respectively. Morphological changes of macrophages, as well as the migratory potential of cancer cells, were assessed by the Incucyte system and macrophage matrix degradation potential by zymography. Changes in the metabolic activity of macrophages were quantified using a Seahorse analyzer. For in vivo studies, EVs were injected into the yolk sac of zebrafish larvae, and macrophages were isolated by fluorescence-activated cell sorting.
RESULTS
EVs from WT and KO cells had a similar size and concentration and were positive for 25 vesicle markers. The expression of tumor-promoting genes was higher in macrophages polarized with WT EVs than KO EVs, while the expression of TNF and IL6 was reduced. A similar pattern was observed in macrophages from zebrafish larvae treated in vivo. WT EV-polarized macrophages showed a higher abundance of TAM-like surface markers, higher matrix degrading activity, as well as a higher promotion of cancer cell migration. MicroRNA-Seq revealed a significant difference in the microRNA composition of WT and KO EVs, particularly a high abundance of miR-181a-5p in WT EVs, which was absent in KO EVs. Inhibitors of macropinocytosis and phagocytosis antagonized the delivery of miR-181a-5p into macrophages and the downregulation of the miR-181a-5p target DUSP6. Proteomics data showed differences in protein cargo in KO vs. WT EVs, with the differentially abundant proteins mainly involved in metabolic pathways. WT EV-treated macrophages exhibited a higher basal oxygen consumption rate and a lower extracellular acidification rate than KO EV-treated cells.
CONCLUSION
Our results show that IMP2 determines the cargo of EVs released by cancer cells, thereby modulating the EVs' actions on macrophages. Expression of IMP2 is linked to the secretion of EVs that polarize macrophages towards a tumor-promoting phenotype.
Topics: Humans; Extracellular Vesicles; RNA-Binding Proteins; Animals; Zebrafish; Tumor-Associated Macrophages; HCT116 Cells; MicroRNAs; Cell Movement; Macrophages
PubMed: 38937789
DOI: 10.1186/s12964-024-01701-y -
The Science of the Total Environment Jun 2024Crystalline silica (CS) particles are ubiquitously present in the environment, particularly in occupational settings, and exposure to respirable CS causes silicosis,...
Crystalline silica (CS) particles are ubiquitously present in the environment, particularly in occupational settings, and exposure to respirable CS causes silicosis, imposing a significant disease burden. However, the pathogenesis of silicosis remains unclear. Exposure to external stimuli, such as CS, leads to the accumulation of unfolded proteins and triggers endoplasmic reticulum (ER) stress, disrupting tissue immune homeostasis and accelerating pathological progression. While pulmonary macrophages phagocytose CS particles to initiate the immune response, the role of ER stress in this process is unknown. Herein, we used a murine model of silicosis to simulate the pathological progression from acute inflammation to fibrosis in silicosis and conducted in vivo pharmacological inhibition of ER stress to explore the underlying mechanism. Using flow cytometry, we further classified pulmonary macrophages into monocyte-like macrophages (monocytes), interstitial macrophages (IMs), and alveolar macrophages (AMs). Our results showed that CS-induced ER stress primarily contributed to the augmentation of IMs and thereby exerted a significant impact on pulmonary macrophages. Despite coexpressing M1- and M2-like markers, IMs predominantly exhibited an M1-like polarization state and played a proinflammatory role by expressing the cytokines pro-IL-1β and TNF-α during the pathological progression of silicosis. Additionally, IMs recruited by CS-induced ER stress also exhibited high expression of MHCII and exerted active immunomodulatory effects. Overall, our study demonstrates that ER stress induced by CS particles triggers a proinflammatory immune microenvironment dominated by IMs and reveals novel insights into the pulmonary toxicological effects of CS particles.
PubMed: 38936737
DOI: 10.1016/j.scitotenv.2024.174299 -
Journal of Immunological Methods Jun 2024Antibody-dependent cellular phagocytosis (ADCP) is a cellular process by which antibody-opsonized targets (pathogens or cells) activate the Fc receptors on the surface...
Antibody-dependent cellular phagocytosis (ADCP) is a cellular process by which antibody-opsonized targets (pathogens or cells) activate the Fc receptors on the surface of phagocytes to induce phagocytosis, resulting in internalization and degradation of pathogens or target cells through phagosome acidification. Besides NK cells-mediated antibody-dependent cellular cytotoxicity (ADCC), tumor-infiltrated monocytes and macrophages can directly kill tumor cells in the presence of tumor antigen-specific antibodies through ADCP, representing another attractive strategy for cancer immunotherapy. Even though several methods have been developed to measure ADCP, an automated and high-throughput quantitative assay should offer highly desirable advantages for drug discovery. In this study we established a new ADCP assay to identify therapeutical monoclonal antibodies (mAbs) that facilitate macrophages phagocytosis of live target cells. We used Incucyte, an imaging system for live cell analysis. By labeling the live target cells with a pH sensitive dye (pHrodo), we successfully monitored the ADCP in real time. We demonstrated that our image-based assay is robust and quantitative, suitable for screening and characterization of therapeutical mAbs that directly kill target cells through ADCP. Furthermore, we found different subtypes of macrophages have distinct ADCP activities using both mouse and human primary macrophages differentiated in vitro. By studying various mAbs with mutations in their Fc regions using our assay, we showed that the variants with increased binding to Fc gamma receptors (FcγRs) have enhanced ADCP activities.
PubMed: 38936465
DOI: 10.1016/j.jim.2024.113715 -
PloS One 2024We investigated the interactions of unopsonized and opsonized Mycoplasma mycoides subsp. mycoides (Mmm) with bovine macrophages in vitro. Mmm survived and proliferated...
We investigated the interactions of unopsonized and opsonized Mycoplasma mycoides subsp. mycoides (Mmm) with bovine macrophages in vitro. Mmm survived and proliferated extracellularly on bovine macrophage cell layers in the absence of Mmm-specific antisera. Bovine complement used at non-bactericidal concentrations did neither have opsonizing effect nor promoted intracellular survival, whereas Mmm-specific antisera substantially increased phagocytosis and Mmm killing. A phagocytosis-independent uptake of Mmm by macrophages occurred at a high multiplicity of infection, also found to induce the production of TNF, and both responses were unaffected by non-bactericidal doses of bovine complement. Bovine complement used at higher doses killed Mmm in cell-free cultures and completely abrogated TNF responses by macrophages. These results provide a framework to identify Mmm antigens involved in interactions with macrophages and targeted by potentially protective antibodies and point towards a pivotal role of complement in the control of inflammatory responses in contagious bovine pleuropneumonia.
Topics: Animals; Cattle; Macrophages; Phagocytosis; Complement System Proteins; Mycoplasma; Tumor Necrosis Factor-alpha; Pleuropneumonia, Contagious; Mycoplasma mycoides
PubMed: 38935768
DOI: 10.1371/journal.pone.0305851 -
Cell Reports Jun 2024Oligodendrocyte death is common in aging and neurodegenerative disease. In these conditions, dying oligodendrocytes must be efficiently removed to allow remyelination...
Oligodendrocyte death is common in aging and neurodegenerative disease. In these conditions, dying oligodendrocytes must be efficiently removed to allow remyelination and to prevent a feedforward degenerative cascade. Removal of this cellular debris is thought to primarily be carried out by resident microglia. To investigate the cellular dynamics underlying how microglia do this, we use a single-cell cortical demyelination model combined with longitudinal intravital imaging of dual-labeled transgenic mice. Following phagocytosis, single microglia clear the targeted oligodendrocyte and its myelin sheaths in one day via a precise, rapid, and stereotyped sequence. Deletion of the fractalkine receptor, CX3CR1, delays the microglial phagocytosis of the cell soma but has no effect on clearance of myelin sheaths. Unexpectedly, deletion of the phosphatidylserine receptor, MERTK, has no effect on oligodendrocyte or myelin sheath clearance. Thus, separate molecular signals are used to detect, engage, and clear distinct sub-compartments of dying oligodendrocytes to maintain tissue homeostasis.
PubMed: 38935500
DOI: 10.1016/j.celrep.2024.114385 -
Journal of Pediatric Hematology/oncology Jun 2024
PubMed: 38934621
DOI: 10.1097/MPH.0000000000002907