Professional Phagocytes Research Articles

Microglia: Key Players in Retinal Ageing and Neurodegeneration.

Microglia are the resident immune cells of the central nervous system (CNS) and play a key role in maintaining the normal function of the retina and brain. During early development, microglia migrate into the retina, transform into a highly ramified phenotype, and scan their environment constantly. Microglia can be activated by any homeostatic disturbance that may endanger neurons and threaten tissue integrity. Once activated, the young microglia exhibit a high diversity in their phenotypes as well as their functions, which relate to either beneficial or harmful consequences. Microglial activation is associated with the release of cytokines, chemokines, and growth factors that can determine pathological outcomes. As the professional phagocytes in the retina, microglia are responsible for the clearance of pathogens, dead cells, and protein aggregates. However, their phenotypic diversity and phagocytic capacity is compromised with ageing. This may result in the accumulation of protein aggregates and myelin debris leading to retinal neuroinflammation and neurodegeneration. In this review, we describe microglial phenotypes and functions in the context of the young and ageing retina, and the mechanisms underlying changes in ageing. Additionally, we review microglia-mediated retinal neuroinflammation and discuss the mechanisms of microglial involvement in retinal neurodegenerative diseases.

Efferocytosis and Its Role in Inflammatory Disorders.

Efferocytosis is the effective clearance of apoptotic cells by professional and non-professional phagocytes. The process is mechanically different from other forms of phagocytosis and involves the localization, binding, internalization, and degradation of apoptotic cells. Defective efferocytosis has been demonstrated to associate with the pathogenesis of various inflammatory disorders. In the current review, we summarize recent findings with regard to efferocytosis networks and discuss the relationship between efferocytosis and different immune cell populations, as well as describe how efferocytosis helps resolve inflammatory response and modulate immune balance. Our knowledge so far about efferocytosis suggests that it may be a useful target in the treatment of numerous inflammatory diseases.

Harnessing anti-tumor and tumor-tropism functions of macrophages via nanotechnology for tumor immunotherapy.

Reprogramming the immunosuppressive tumor microenvironment by modulating macrophages holds great promise in tumor immunotherapy. As a class of professional phagocytes and antigen-presenting cells in the innate immune system, macrophages can not only directly engulf and clear tumor cells, but also play roles in presenting tumor-specific antigen to initiate adaptive immunity. However, the tumor-associated macrophages (TAMs) usually display tumor-supportive M2 phenotype rather than anti-tumor M1 phenotype. They can support tumor cells to escape immunological surveillance, aggravate tumor progression, and impede tumor-specific Tcell immunity. Although many TAMs-modulating agents have shown great success in therapy of multiple tumors, they face enormous challenges including poor tumor accumulation and off-target side effects. An alternative solution is the use of advanced nanostructures, which not only can deliver TAMs-modulating agents to augment therapeutic efficacy, but also can directly serve as modulators of TAMs. Another important strategy is the exploitation of macrophages and macrophage-derived components as tumor-targeting delivery vehicles. Herein, we summarize the recent advances in targeting and engineering macrophages for tumor immunotherapy, including (1) direct and indirect effects of macrophages on the augmentation of immunotherapy and (2) strategies for engineering macrophage-based drug carriers. The existing perspectives and challenges of macrophage-based tumor immunotherapies are also highlighted.

Understanding antimicrobial peptides and combatting antimicrobial resistance problems

Understanding antimicrobial peptides and combatting antimicrobial resistance problems

Mode of action of virulence factors of intracellular pathogens studies with time-resolved and high-resolution atomic force microscopy

An essential part of the innate immune defenses is the phagocytosis of invading pathogenic microbes by professional phagocytes and their subsequent digestion within the phagosome. It is often forgotten that not all pathogens surrender without a fight—in particular, some species succeed in surviving inside the phagosome. These intracellular pathogens have evolved various individual strategies to counteract the killing mechanisms of the host cell, many of which have been poorly understood.

The possibility of using xenogeneic phagocytes in wound treatment.

Metamorphosis in the insect larva is associated with disintegration, engulf and digestion of larval tissues. These processes are accompanied by a significant shift in physiological parameters like high activity of hydrolytic enzymes and decrease of pH. In the way, the metamorphosing larva resembles the processes occurring in the wound at the stage of inflammation. Based on this thesis, we put forward the idea of the possibility of using insect phagocytes in the wound treatment. The search for a suitable insect cell line and the study of its properties were the purpose of the work. The abilities of insect phagocytes to retain viability and functional activity under conditions physiological for humans were also investigated. We found that blue blowfly Calliphora vicina larvae had histolysocytes, a specialized population of professional phagocytes involved in the histolysis. In vitro, histolysocytes possess high phagocytic activity to fragments of vertebrate soft tissues and debris. These cells retain viability and functional activity for a long time under conditions that are physiological for vertebrate cells. Moreover histolysocytes can realize the humoral control over the bacteria through the synthesis of antimicrobial peptides. So histolysocytes have the potential to be used as xenogeneic phagocytes in the wound treatment. The data obtained allow proceeding to experiments on laboratory animals for studying the effect of such therapy on the wound healing process.

Neutrophils in Extravascular Body Fluids: Cytological-Energy Analysis Enables Rapid, Reliable and Inexpensive Detection of Purulent Inflammation and Tissue Damage.

The simultaneous cytological and metabolic investigation of various extravascular body fluids (EBFs) provides clinically relevant information about the type and intensity of the immune response in particular organ systems. The oxidative burst of professional phagocytes with the concomitant production of reactive oxygen species consumes a large amount of oxygen and is the cause of switch to the development of anaerobic metabolism. We assessed the relationships between percentages of neutrophils, aerobic and anaerobic metabolism, and tissue damage via the determination of aspartate aminotransferase catalytic activities (AST) in cerebrospinal fluid (CSF), pleural effusions (PE), abdominal effusions (AE), and synovial fluids (SF). EBFs with 0.0–20.0% neutrophils: 83.0% aerobic and 1.3% strongly anaerobic cases with median of AST = 13.8 IU/L in CSF; 68.0% aerobic and 9.0% strongly anaerobic cases with median of AST = 20.4 IU/L in PE; 77.5% aerobic and 10.5% strongly anaerobic cases with median of AST = 18.0 IU/L in AE; 64.1% aerobic and 7.7% strongly anaerobic cases with median of AST = 13.8 IU/L in SF. EBFs with 80.0–100.0% neutrophils: 4.2% aerobic and 73.7% strongly anaerobic cases with median of AST = 19.2 IU/L in CSF; 7.4% aerobic and 77.3% strongly anaerobic cases with median of AST = 145.2 IU/L in PE; 11.8% aerobic and 73.7% strongly anaerobic cases with median of AST = 61.8 IU/L in AE; 25.5% aerobic and 38.2% strongly anaerobic cases with median of AST = 37.2 IU/L in SF. The significant presence of neutrophils, concomitant strong anaerobic metabolism, and elevated AST in various EBFs are reliable signs of damaging purulent inflammation.

VASP boosts protrusive activity of macroendocytic cups and drives phagosome rocketing after internalization

Ena/VASP proteins are powerful actin polymerases that drive the processive elongation of actin filaments. Members of this protein family have been implicated in a variety of important cellular processes including axon guidance, cell migration and adhesion. However, the specific function of these proteins in macroendocytosis, comprising macropinocytosis and phagocytosis remain rather poorly understood. Here, we used the professional phagocyte Dictyostelium discoideum to address the function and dynamics of its only family member VASP in macroendocytosis. Confocal time-lapse imaging revealed that VASP localized prominently in a circumferential narrow band at the advancing rim of the phagocytic cup followed by its aperture-like convergence upon particle internalization. Loss of VASP resulted in substantial defects in both, macropinocytosis of bulk fluid and phagocytosis of yeast particles. Consistently, VASP-deficiency coincided with diminished speed of the protruding rim and an impaired internalization rate. Most intriguingly, after cup closure, VASP condensed at the distal side of internalized phagosomes and initiated localized de-novo actin assembly to propel the phagosome by an actin-rich comet deeper into the cell, resembling intracellular movement of rocketing Listeria cells. In line with these findings, travelled distance and speed of rocketing phagosomes in VASP-deficient cells were markedly impaired.

Eosinophil efferocytosis during emphysematous eosinophilic lymphangitis in the bovine rumen: histopathological evaluation with special focus on cellular kinetics of professional phagocytes

Eosinophil efferocytosis during emphysematous eosinophilic lymphangitis in the bovine rumen: histopathological evaluation with special focus on cellular kinetics of professional phagocytes

Myelin and non-myelin debris contribute to foamy macrophage formation after spinal cord injury

Tissue damage after spinal cord injury (SCI) elicits a robust inflammatory cascade that fails to resolve in a timely manner, resulting in impaired wound healing and cellular regeneration. This inflammatory response is partly mediated by infiltrating immune cells, including macrophages. As professional phagocytes, macrophages initially play an important role in debris clearance at the injury site, which would be necessary for proper tissue regeneration. After SCI, most macrophages become filled with lipid droplets due to excessive uptake of lipid debris, assuming a “foamy” phenotype that is associated with a proinflammatory state. Myelin has been assumed to be the main source of lipid that induces foamy macrophage formation after injury given its abundance in the spinal cord. This assumption has led to the widespread use of purified myelin treatment to model foamy macrophage formation in vitro. However, the assumption that myelin is necessary for foamy macrophage formation remains untested. To this end, we developed a novel foamy macrophage assay utilizing total spinal cord homogenate to include all sources of lipid present at the injury site. Using the myelin basic protein knockout (MBP KO, i.e., Shiverer) mice that lack myelin, we investigated lipid accumulation in foamy macrophages. Primary macrophages treated with myelin-deficient spinal cord homogenate still formed large lipid droplets typically observed in foamy macrophages, although to a lesser degree than cells treated with normal homogenate. Similarly, MBP KO mice subjected to contusive spinal cord injury also formed foamy macrophages that exhibited reduced lipid content and associated with improved histological outcomes and reduced immune cell infiltration. Therefore, the absence of myelin does not preclude foamy macrophage formation, indicating that myelin is not the only major source of lipid that contributes this pathology, even though myelin may alter certain aspects of its inflammatory profile.

The in vitro effects of synthetic complement peptide C3a during Brucella abortus 544 infection in a murine professional phagocyte RAW264.7 cell line

The in vitro effects of synthetic complement peptide C3a during Brucella abortus 544 infection in a murine professional phagocyte RAW264.7 cell line

Characterization and mode of action analysis of black soldier fly (Hermetia illucens) larva-derived hemocytes.

With the growing importance of the black soldier fly (Hermetia illucens) for both sustainable food production and waste management as well as for science, a great demand of understanding its immune system arises. Here, we present the first description of the circulating larval hemocytes with special emphasis on uptake of microorganisms and distinguishing hemocyte types. With histological, zymographic, and cytometric methods and with a set of hemocyte binding lectins and antibodies, the hemocytes of H. illucens are identified as plasmatocytes, crystal cells, and putative prohemocytes. Total hemocyte counts (THC) are determined, and methods for THC determination are compared. Approximately 1100 hemocytes per microliter hemolymph are present in naive animals, while hemocyte density decreases dramatically shortly after wounding, indicating a role of hemocytes in response to wounding (and immune response in general). The determination of the relative abundance of each hemocyte type (differential hemocyte count, DHC) revealed that plasmatocytes are highly abundant, whereas prohemocytes and crystal cells make up only a small percentage of the circulating cells. Plasmatocytes are not only the most abundant but also the professional phagocytes in H. illucens. They rapidly engulf and take up bacteria both in vivo and in vitro, indicating a very potent cellular defense against invading pathogens. Larger bioparticles such as yeasts are also removed from circulation by phagocytosis, but slower than bacteria. This is the first analysis of the potent cellular immune response in the black soldier fly, and a first toolbox that helps to identify hemocyte (types) is presented.

The Cellular Innate Immune Response of the Invasive Pest Insect Drosophila suzukii against Pseudomonas entomophila Involves the Release of Extracellular Traps

Drosophila suzukii is a neobiotic invasive pest that causes extensive damage to fruit crops worldwide. The biological control of this species has been unsuccessful thus far, in part because of its robust cellular innate immune system, including the activity of professional phagocytes known as hemocytes and plasmatocytes. The in vitro cultivation of primary hemocytes isolated from D. suzukii third-instar larvae is a valuable tool for the investigation of hemocyte-derived effector mechanisms against pathogens such as wasp parasitoid larvae, bacteria, fungi and viruses. Here, we describe the morphological characteristics of D. suzukii hemocytes and evaluate early innate immune responses, including extracellular traps released against the entomopathogen Pseudomonas entomophila and lipopolysaccharides. We show for the first time that D. suzukii plasmatocytes cast extracellular traps to combat P. entomophila, along with other cell-mediated reactions, such as phagocytosis and the formation of filopodia.

Cytosolic detection of phagosomal bacteria-Mechanisms underlying PAMP exodus from the phagosome into the cytosol.

The metazoan innate immune system senses bacterial infections by detecting highly conserved bacterial molecules, termed pathogen-associated molecular patterns (PAMPs). PAMPs are detected by a variety of host pattern recognition receptors (PRRs), whose function is to coordinate downstream immune responses. PRR activities are, in part, regulated by their subcellular localizations. Accordingly, professional phagocytes can detect extracellular bacteria and their PAMPs via plasma membrane-oriented PRRs. Conversely, phagocytosed bacteria and their PAMPs are detected by transmembrane PRRs oriented toward the phagosomal lumen. Even though PAMPs are unable to passively diffuse across membranes, phagocytosed bacteria are also detected by PRRs localized within the host cell cytosol. This phenomenon is explained by phagocytosis of bacteria that specialize in phagosomal escape and cytosolic residence. Contrary to this cytosolic lifestyle, most bacteria studied to date spend their entire intracellular lifestyle contained within phagosomes, yet they also stimulate cytosolic PRRs. Herein, we will review our current understanding of how phagosomal PAMPs become accessible to cytosolic PRRs, as well as highlight knowledge gaps that should inspire future investigations.

Single-Cell Transcriptome Profiling Identifies Phagocytosis-Related Dual-Feature Cells in A Model of Acute Otitis Media in Rats.

BackgroundThe molecular mechanisms of acute otitis media (AOM) development, and the intercellular crosstalk within the multicellular ecosystem of AOM, are not clear.MethodsWe established a model of AOM in rats (with normal rats as controls) and undertook single-cell RNA sequencing (scRNA-seq) for the middle-ear mucosa (MEM). Cell clustering and trajectory analyses were undertaken using Seurat and Monocle 2 packages in R software. Pathway analyses were done by gene set enrichment analysis (GSEA). Cell–cell interactions were inferred by CellChat. Cell scores were calculated to identify cells with dual-feature.ResultsA total of 7023 cells from three samples of inflamed MEM and 5258 cells from three samples of healthy MEM underwent scRNA-seq, which identified 20 cell clusters belonging to eight major cell types. After exposure to lipopolysaccharide, the MEM underwent significant conversion of cell types characterized by rapid infiltration of macrophages and neutrophils. M2 macrophages seemed to play a key part in inflammatory intercellular crosstalk, which facilitated the maintenance and proliferation of macrophages, cell chemotaxis, and regulation of the proinflammatory activities of cytokines. Three rare cell clusters with phagocytosis-related dual-feature were also identified. They coexisted with professional phagocytes in the MEM, and displayed distinct immunoregulatory functions by maintaining a normal immune microenvironment or influencing inflammation progression.ConclusionsMacrophages might be the “master” initiators and regulators of the inflammatory response of the MEM to external stimuli. And their functions are fulfilled by a specific polarization status (M2) and sophisticated intercellular crosstalk via certain signaling pathways. Besides, the coexistence of professional phagocytes and non-professional phagocytes as well as their interplay in the MEM provides new clues for deciphering the underlying pathogenic mechanisms of AOM.

Phagocytic microglia in development: Are they what they eat?

Microglia, the innate immune cells of the brain, are indispensable for proper brain development. As professional phagocytes, microglia engulf other cells within distinct developmental niches to sculpt the architecture of the brain. Here, I highlight the age-, brain region-, and substrate-dependent diversity of developmental phagocytosis, and pose the idea that phagocytosis may, in turn, drive changes in microglia phenotype. Ultimately, phagocytosis might be just as important for shaping microglia function as it is for shaping the brain.

Bacterial minicells to the rescue: cyto-Immunotherapy for the treatment of late stage cancers with minimal to no toxicity.

Bacterial minicells to the rescue: cyto-Immunotherapy for the treatment of late stage cancers with minimal to no toxicity.

Immune functions of phagocytic blood cells in teleost

AbstractPhagocytosis is a fundamental immune function against foreign microorganisms and abnormal cells, and it is also a bridge between innate and adaptive immunity. Blood as the direct living environment of various immune cells is a crucial site for phagocytosis. In this review, we firstly summarized the morphological structure and cytochemical characteristics of teleost peripheral blood cells, divided them into erythrocytes, leucocytes and thrombocytes, and then systematically elaborated the immune functions of phagocytic blood cells including professional and non‐professional phagocytes. The professional phagocytes consist of monocytes/macrophages and neutrophils. Monocytes/macrophages activate phagocytosis to ingest pathogens, and destroy them through respiratory burst and nitric oxide response to generate antigens, and then antigens are presented to adaptive immune cells. Neutrophils are recruited to inflammatory sites and exert phagocytosis, which is accompanied by degranulation and the production of neutrophil extracellular traps to kill pathogens. Non‐professional phagocytes including thrombocytes, B cells and erythrocytes are indispensable to exercise the phagocytosis, in addition to their main functions in the immune system. Thrombocytes participate in innate immunity and adaptive immunity. B cells carry out antigen presentation and immunoglobulin secretion in adaptive immunity. Erythrocytes perform immune regulation and infection resistance. In general, professional phagocytes play a primary role in phagocytosis and pathogen treatment, whereas non‐professional phagocytes play auxiliary role in phagocytosis and perform their unique immune functions. The functions of these two types of phagocytes jointly constitute the basis of the immune system. This review provides insights into the overall roles of phagocytic blood cells in the teleost immune system.

Intracellular Staphylococcus aureus employs the cysteine protease staphopain A to induce host cell death in epithelial cells.

Staphylococcus aureus is a major human pathogen, which can invade and survive in non-professional and professional phagocytes. Uptake by host cells is thought to contribute to pathogenicity and persistence of the bacterium. Upon internalization by epithelial cells, cytotoxic S. aureus strains can escape from the phagosome, replicate in the cytosol and induce host cell death. Here, we identified a staphylococcal cysteine protease to induce cell death after translocation of intracellular S. aureus into the host cell cytoplasm. We demonstrated that loss of staphopain A function leads to delayed onset of host cell death and prolonged intracellular replication of S. aureus in epithelial cells. Overexpression of staphopain A in a non-cytotoxic strain facilitated intracellular killing of the host cell even in the absence of detectable intracellular replication. Moreover, staphopain A contributed to efficient colonization of the lung in a mouse pneumonia model. In phagocytic cells, where intracellular S. aureus is exclusively localized in the phagosome, staphopain A did not contribute to cytotoxicity. Our study suggests that staphopain A is utilized by S. aureus to exit the epithelial host cell and thus contributes to tissue destruction and dissemination of infection.

Tumor Temporal Proteome Profiling Reveals the Immunological Triple Offensive Induced by Synthetic Anti-Cancer Salmonella.

The engineered “obligate” anaerobic Salmonella typhimurium strain YB1 shows a prominent ability to repress tumor growth and metastasis, which has great potential as a novel cancer immunotherapy. However, the antitumor mechanism of YB1 remains unelucidated. To resolve the proteome dynamics induced by the engineered bacteria, we applied tumor temporal proteome profiling on murine bladder tumors after intravenous injection of either YB1 or PBS as a negative control. Our data suggests that during the two weeks treatment of YB1 injections, the cured tumors experienced three distinct phases of the immune response. Two days after injection, the innate immune response was activated, particularly the complement and blood coagulation pathways. In the meantime, the phagocytosis was initiated. The professional phagocytes such as macrophages and neutrophils were recruited, especially the infiltration of iNOS+ and CD68+ cells was enhanced. Seven days after injection, substantial amount of T cells was observed at the invasion margin of the tumor. As a result, the tumor shrunk significantly. Overall, the temporal proteome profiling can systematically reveal the YB1 induced immune responses in tumor, showing great promise for elucidating the mechanism of bacteria-mediated cancer immunotherapy.