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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 -
Haematologica Jun 2024Macrophages are one of the key mediators of the therapeutic effects exerted by monoclonal antibodies, such as the anti-CD19 antibody tafasitamab, approved in combination...
Macrophages are one of the key mediators of the therapeutic effects exerted by monoclonal antibodies, such as the anti-CD19 antibody tafasitamab, approved in combination with lenalidomide for the treatment of relapsed or refractory (r/r) diffuse large B cell lymphoma (DLBCL). However, antibody-dependent cellular phagocytosis (ADCP) in the tumor microenvironment can be counteracted by increased expression of the inhibitory receptor SIRPα on macrophages and its ligand, the immune checkpoint molecule CD47 on tumor cells. The aim of this study was to investigate the impact of the CD47-SIRPα axis on tafasitamabmediated phagocytosis and explore the potential of anti-CD47 blockade to enhance its antitumor activity. Elevated expression of both SIRPα and CD47 was observed in DLBCL patient-derived lymph node biopsies compared to healthy controls. CRISPR-mediated CD47 overexpression impacted tafasitamab-mediated ADCP in vitro and increased expression of SIRPα on macrophages correlated with decreased ADCP activity of tafasitamab against DLBCL cell lines. Combination of tafasitamab and an anti-CD47 blocking antibody enhanced ADCP activity of in vitro generated macrophages. Importantly, tafasitamab-mediated phagocytosis was elevated in combination with CD47 blockade using primary DLBCL cells and patient-derived lymphoma-associated macrophages (LAMs) in an autologous setting. Furthermore, lymphoma cells with low CD19 expression were efficiently eliminated by the combination treatment. Finally, combined treatment of tafasitamab and an anti-CD47 antibody resulted in enhanced tumor volume reduction and survival benefit in lymphoma xenograft mouse models. These findings provide evidence that CD47 blockade can enhance the phagocytic potential of tumor targeting immunotherapies such as tafasitamab and suggest there is value in exploring the combination in the clinic.
PubMed: 38934068
DOI: 10.3324/haematol.2023.284795 -
Nanoscale Advances Jun 2024Tumors pose a significant threat to human health, and their occurrence and fatality rates are on the rise each year. Accurate tumor diagnosis is crucial in preventing...
Tumors pose a significant threat to human health, and their occurrence and fatality rates are on the rise each year. Accurate tumor diagnosis is crucial in preventing untimely treatment and late-stage metastasis, thereby reducing mortality. To address this, we have developed a novel type of hybrid nanogel called γ-FeO@PNIPAM/PAm/CTS, which contains iron oxide nanoparticles and poly(-isopropyl acrylamide)/polyacrylamide/chitosan. The rationale for this study relies on the concept that thermosensitive PNIPAM has the ability to contract when exposed to elevated temperature conditions found within tumors. This contraction leads to a dense clustering of the high-loading γ-FeO nanoparticles within the nanogel, thus greatly enhancing the capabilities of MRI. Additionally, the amino groups in chitosan on the particle surface can be converted into ammonium salts under mildly acidic conditions, allowing for an increase in the charge of the nanogel specifically at the slightly acidic tumor site. Consequently, it promotes the phagocytosis of tumor cells and effectively enhances the accumulation and retention of nanogels at the tumor site. The synthesis of the hybrid nanogels involves a surfactant-free emulsion copolymerization process, where vinyl-modified γ-FeO superparamagnetic nanoparticles are copolymerized with the monomers in the presence of chitosan. We have optimized various reaction parameters to achieve a high loading content of the superparamagnetic nanoparticles, reaching up to 60%. The achieved value of 517.74 mM S significantly surpasses that of the clinical imaging contrast agent Resovist (approximately 151 mM S). To assess the performance of these magnetic nanogels, we conducted experiments using Cal27 oral tumors and 4T1 breast tumors in animal models. The nanogels exhibited temperature- and pH-sensitivity, enabling magnetic targeting and enhancing diagnosis through MRI. The results demonstrated the potential of these hybrid nanogels as contrast agents for magnetic targeting in biomedical applications.
PubMed: 38933853
DOI: 10.1039/d4na00014e -
The Journal of the Association of... Jun 2024Dehydration, often unnoticed, can occur in adults due to heat-related illnesses, viral infections, and upper respiratory tract infections, leading to deficits in fluids,... (Review)
Review
Dehydration, often unnoticed, can occur in adults due to heat-related illnesses, viral infections, and upper respiratory tract infections, leading to deficits in fluids, electrolytes, and energy (FEE), even without diarrhea. This fluid deficit, in turn, impairs immunity at cellular and molecular levels. Physiologically, the immune system rapidly reacts to threats by adjusting its structural permeability, allowing water and small solutes to be passed, facilitated by proteins like aquaporins (AQPs). These AQPs are also essential for immune cell communication and adjustments of the cytoskeleton for functions like chemotaxis and phagocytosis. Apart from hydration, maintaining an optimal intake of essential micronutrients, including vitamins C and E, copper, zinc, and selenium, is also critical for supporting immune function and acting as antioxidants. Transient alterations in immune function caused by acute stressors or nondiarrheal conditions heighten vulnerability to coexisting illnesses, particularly cold and upper respiratory tract infections, while also elevating the likelihood of recurrent health issues. Supplementing fluids with known FEE content according to quality standards and enhanced taste is recommended during acute illness and until complete recovery. Thus, dehydration, caused by various acute conditions, can impair immunity and delay recovery. Prioritizing hydration and nutrition in healthcare and general settings is imperative, even when diarrhea is absent.
Topics: Humans; Micronutrients; Dehydration; Fluid Therapy
PubMed: 38932732
DOI: 10.59556/japi.72.0546 -
Nutrients Jun 2024The immune system is affected by the dietary products humans intake. Immune system regulation by nutrition has uses in the clinical context, but it can also benefit... (Randomized Controlled Trial)
Randomized Controlled Trial
Prospective, Randomized, Double-Blind Parallel Group Nutritional Study to Evaluate the Effects of Routine Intake of Fresh vs. Pasteurized Yogurt on the Immune System in Healthy Adults.
The immune system is affected by the dietary products humans intake. Immune system regulation by nutrition has uses in the clinical context, but it can also benefit healthy populations by delaying or preventing the emergence of immune-mediated chronic illnesses. In this study, the purpose was to describe and compare the modulator effects on the immune system of the routine ingestion of fresh vs. pasteurized yogurt. A unicentral, prospective, randomized, double-blind, parallel group 8-week nutritional study was carried out comparing the ingestion of 125 g of the products in healthy adults three times a day. A complete battery of in vitro tests on the activity of the immune system, processes and phenomena was performed. Exclusive immune-modulatory effects of fresh yogurt with respect to base line were found in terms of increased systemic IgM (primary immune responses), increased synthesis of IFN-gamma upon stimulation (Th1) and increased peripheral T cells (mainly "naive" CD4s). In the three interventions, we observed an increased phagocytic activity and burst test in granulocytes, together with increased secretion of IL-6, IL-1 β and IL-8 (pro-inflammatory) and increased CD16 expression (FcR favoring phagocytosis) in granulocytes. Overall, it is concluded that regardless of bacteria being alive or thermally inactivated, yogurt has common effects on the innate system, but the presence of live bacteria is necessary to achieve a potentiating effect on the specific immune response.
Topics: Yogurt; Humans; Double-Blind Method; Adult; Male; Female; Prospective Studies; Pasteurization; Phagocytosis; Cytokines; Young Adult; Immunoglobulin M; Interferon-gamma; Middle Aged; Granulocytes; Immune System; Receptors, IgG
PubMed: 38931322
DOI: 10.3390/nu16121969 -
Medicina (Kaunas, Lithuania) Jun 2024: As modulators of the tumor microenvironment, macrophages have been extensively studied for their potential in developing anticancer strategies, particularly in...
: As modulators of the tumor microenvironment, macrophages have been extensively studied for their potential in developing anticancer strategies, particularly in regulating macrophage polarization towards an antitumorigenic (M1) phenotype rather than a protumorigenic (M2) one in various experimental models. Here, we evaluated the effect of PD98059, a mitogen-activated protein kinase kinase MAPKK MEK1-linked pathway inhibitor, on the differentiation and polarization of THP-1 monocytes in response to phorbol-12-myristate-13-acetate (PMA) under various culture conditions for tumor microenvironmental application. : Differentiation and polarization of THP-1 were analyzed by flow cytometry and RT-PCR. Polarized THP-1 subsets with different treatment were compared by motility, phagocytosis, and so on. : Clearly, PMA induced THP-1 differentiation occurs in adherent culture conditions more than nonadherent culture conditions by increasing CD11b expression up to 90%, which was not affected by PD98059 when cells were exposed to PMA first (post-PD) but inhibited when PD98059 was treated prior to PMA treatment (pre-PD). CD11b THP-1 cells treated with PMA and PMA-post-PD were categorized into M0 (HLA-DR and CD206), M1 (HLA-DR and CD206), and M2 (HLA-DR and CD206), resulting in an increased population of M1 macrophages. The transcription levels of markers of macrophage differentiation and polarization confirmed the increased M1 polarization of THP-1 cells with post-PD treatment rather than with PMA-only treatment. The motility and cytotoxicity of THP-1 cells with post-PD treatment were higher than THP-1 cells with PMA, suggesting that post-PD treatment enhanced the anti-tumorigenicity of THP-1 cells. Confocal microscopy and flow cytometry showed the effect of post-PD treatment on phagocytosis by THP-1 cells. : We have developed an experimental model of macrophage polarization with THP-1 cells which will be useful for further studies related to the tumor microenvironment.
Topics: Humans; Macrophages; Tetradecanoylphorbol Acetate; Flavonoids; THP-1 Cells; Cell Differentiation; Monocytes; Flow Cytometry; Phagocytosis
PubMed: 38929626
DOI: 10.3390/medicina60061009 -
Animals : An Open Access Journal From... Jun 2024MafB is a transcription factor that regulates macrophage differentiation. Macrophages are a traditional feature of the hamster Harderian gland (HG); however, studies...
MafB is a transcription factor that regulates macrophage differentiation. Macrophages are a traditional feature of the hamster Harderian gland (HG); however, studies pertaining to MafB expression in the HG are scant. Here, the full-length cDNA of the gene in hamsters was cloned and sequenced. Molecular characterization revealed that MafB encodes a protein containing 323 amino acids with a DNA-binding domain, a transactivation domain, and a leucine zipper domain. qPCR assays indicated that MafB was expressed in different tissues of both sexes. The highest relative expression levels in endocrine tissues were identified in the pancreas. Gonadectomy in male hamsters was associated with significantly higher mRNA levels in the HG; replacement with dihydrotestosterone restored mRNA expression. The HG in male hamsters contained twofold more MafB mRNA than the HG of female hamsters. Adrenals revealed similar mRNA relative expression levels during the estrous cycle. The estrous phase was associated with higher mRNA levels in the ovary. A significantly up-regulated expression and sexual dimorphism of MafB was found in the pancreas. Therefore, MafB in the HG may play an active role in the macrophage differentiation required for phagocytosis activity and intraocular repair. Additionally, sex steroids appear to strongly influence the MafB expression in the HG and pancreas. These studies highlight the probable biological importance of MafB in immunological defense and pancreatic β cell regulation.
PubMed: 38929347
DOI: 10.3390/ani14121728