HMMR triggers immune evasion of hepatocellular carcinoma by inactivation of phagocyte killing.

Hepatocellular carcinoma (HCC) acquires an immunosuppressive microenvironment, leading to unbeneficial therapeutic outcomes. Hyaluronan-mediated motility receptor (HMMR) plays a crucial role in tumor progression. Here, we found that aberrant expression of HMMR could be a predictive biomarker for the immune suppressive microenvironment of HCC, but the mechanism remains unclear. We established an HMMR-/- liver cancer mouse model to elucidate the HMMR-mediated mechanism of the dysregulated "don't eat me" signal. HMMR knockout inhibited liver cancer growth and induced phagocytosis. HMMRhigh liver cancer cells escaped from phagocytosis via sustaining CD47 signaling. Patients with HMMRhighCD47high expression showed a worse prognosis than those with HMMRlowCD47low expression. HMMR formed a complex with FAK/SRC in the cytoplasm to activate NF-κB signaling, which could be independent of membrane interaction with CD44. Notably, targeting HMMR could enhance anti-PD-1 treatment efficiency by recruiting CD8+ T cells. Overall, our data revealed a regulatory mechanism of the "don't eat me" signal and knockdown of HMMR for enhancing anti-PD-1 treatment.

Recombinant SpTransformer proteins are functionally diverse for binding and phagocytosis by three subtypes of sea urchin phagocytes.

Introduction: The California purple sea urchin, Strongylocentrotus purpuratus, relies solely on an innate immune system to combat the many pathogens in the marine environment. One aspect of their molecular defenses is the SpTransformer (SpTrf) gene family that is upregulated in response to immune challenge. The gene sequences are highly variable both within and among animals and likely encode thousands of SpTrf isoforms within the sea urchin population. The native SpTrf proteins bind foreign targets and augment phagocytosis of a marine Vibrio. A recombinant (r)SpTrf-E1-Ec protein produced by E. coli also binds Vibrio but does not augment phagocytosis. Methods: To address the question of whether other rSpTrf isoforms function as opsonins and augment phagocytosis, six rSpTrf proteins were expressed in insect cells. Results: The rSpTrf proteins are larger than expected, are glycosylated, and one dimerized irreversibly. Each rSpTrf protein cross-linked to inert magnetic beads (rSpTrf::beads) results in different levels of surface binding and phagocytosis by phagocytes. Initial analysis shows that significantly more rSpTrf::beads associate with cells compared to control BSA::beads. Binding specificity was verified by pre-incubating the rSpTrf::beads with antibodies, which reduces the association with phagocytes. The different rSpTrf::beads show significant differences for cell surface binding and phagocytosis by phagocytes. Furthermore, there are differences among the three distinct types of phagocytes that show specific vs. constitutive binding and phagocytosis. Conclusion: These findings illustrate the complexity and effectiveness of the sea urchin innate immune system driven by the natSpTrf proteins and the phagocyte cell populations that act to neutralize a wide range of foreign pathogens.

Collaboration between a cis-interacting natural killer cell receptor and membrane sphingolipid is critical for the phagocyte function.

Inhibitory natural killer (NK) cell receptors recognize MHC class I (MHC-I) in trans on target cells and suppress cytotoxicity. Some NK cell receptors recognize MHC-I in cis, but the role of this interaction is uncertain. Ly49Q, an atypical Ly49 receptor expressed in non-NK cells, binds MHC-I in cis and mediates chemotaxis of neutrophils and type I interferon production by plasmacytoid dendritic cells. We identified a lipid-binding motif in the juxtamembrane region of Ly49Q and found that Ly49Q organized functional membrane domains comprising sphingolipids via sulfatide binding. Ly49Q recruited actin-remodeling molecules to an immunoreceptor tyrosine-based inhibitory motif, which enabled the sphingolipid-enriched membrane domain to mediate complicated actin remodeling at the lamellipodia and phagosome membranes during phagocytosis. Thus, Ly49Q facilitates integrative regulation of proteins and lipid species to construct a cell type-specific membrane platform. Other Ly49 members possess lipid binding motifs; therefore, membrane platform organization may be a primary role of some NK cell receptors.

mTOR signaling is required for phagocyte free radical production, GLUT1 expression, and control of Staphylococcus aureus infection.

Mammalian target of rapamycin (mTOR) is a key regulator of metabolism in the mammalian cell. Here, we show the essential role for mTOR signaling in the immune response to bacterial infection. Inhibition of mTOR during infection with Staphylococcus aureus revealed that mTOR signaling is required for bactericidal free radical production by phagocytes. Mechanistically, mTOR supported glucose transporter GLUT1 expression, potentially through hypoxia-inducible factor 1α, upon phagocyte activation. Cytokine and chemokine signaling, inducible nitric oxide synthase, and p65 nuclear translocation were present at similar levels during mTOR suppression, suggesting an NF-κB-independent role for mTOR signaling in the immune response during bacterial infection. We propose that mTOR signaling primarily mediates the metabolic requirements necessary for phagocyte bactericidal free radical production. This study has important implications for the metabolic requirements of innate immune cells during bacterial infection as well as the clinical use of mTOR inhibitors.IMPORTANCESirolimus, everolimus, temsirolimus, and similar are a class of pharmaceutics commonly used in the clinical treatment of cancer and the anti-rejection of transplanted organs. Each of these agents suppresses the activity of the mammalian target of rapamycin (mTOR), a master regulator of metabolism in human cells. Activation of mTOR is also involved in the immune response to bacterial infection, and treatments that inhibit mTOR are associated with increased susceptibility to bacterial infections in the skin and soft tissue. Infections caused by Staphylococcus aureus are among the most common and severe. Our study shows that this susceptibility to S. aureus infection during mTOR suppression is due to an impaired function of phagocytic immune cells responsible for controlling bacterial infections. Specifically, we observed that mTOR activity is required for phagocytes to produce antimicrobial free radicals. These results have important implications for immune responses during clinical treatments and in disease states where mTOR is suppressed.

Deletion of Tmem268 in mice suppresses anti-infectious immune responses by downregulating CD11b signaling.

Transmembrane protein 268 (TMEM268) is a novel, tumor growth-related protein first reported by our laboratory. It interacts with the integrin subunit ß4 (ITGB4) and plays a positive role in the regulation of the ITGB4/PLEC signaling pathway. Here, we investigated the effects and mechanism of TMEM268 in anti-infectious immune response in mice. Tmem268 knockout in mice aggravated cecal ligation and puncture-induced sepsis, as evidenced by higher bacterial burden in various tissues and organs, congestion, and apoptosis. Moreover, Tmem268 deficiency in mice inhibited phagocyte adhesion and migration, thus decreasing phagocyte infiltration at the site of infection and complement-dependent phagocytosis. Further findings indicated that TMEM268 interacts with CD11b and inhibits its degradation via the endosome-lysosome pathway. Our results reveal a positive regulatory role of TMEM268 in ß2 integrin-associated anti-infectious immune responses and signify the potential value of targeting the TMEM268-CD11b signaling axis for the maintenance of immune homeostasis and immunotherapy for sepsis and related immune disorders.

Whole-transcriptome sequencing of phagocytes reveals a ceRNA network contributing to natural resistance to tuberculosis infection.

Tuberculosis (TB) is a major fatal infectious disease globally, exhibiting high morbidity rates and impacting public health and other socio-economic factors. However, some individuals are resistant to TB infection and are referred to as "Resisters". Resisters remain uninfected even after exposure to high load of Mycobacterium tuberculosis (Mtb). To delineate this further, this study aimed to investigate the factors and mechanisms influencing the Mtb resistance phenotype. We assayed the phagocytic capacity of peripheral blood mononuclear cells (PBMCs) collected from Resisters, patients with latent TB infection (LTBI), and patients with active TB (ATB), following infection with fluorescent Mycobacterium bovis Bacillus Calmette-Guérin (BCG). Phagocytosis was stronger in PBMCs from ATB patients, and comparable in LTBI patients and Resisters. Subsequently, phagocytes were isolated and subjected to whole transcriptome sequencing and small RNA sequencing to analyze transcriptional expression profiles and identify potential targets associated with the resistance phenotype. The results revealed that a total of 277 mRNAs, 589 long non-coding RNAs, 523 circular RNAs, and 35 microRNAs were differentially expressed in Resisters and LTBI patients. Further, the endogenous competitive RNA (ceRNA) network was constructed from differentially expressed genes after screening. Bioinformatics, statistical analysis, and quantitative real-time polymerase chain reaction were used for the identification and validation of potential crucial targets in the ceRNA network. As a result, we obtained a ceRNA network that contributes to the resistance phenotype. TCONS_00034796-F3, ENST00000629441-DDX43, hsa-ATAD3A_0003-CYP17A1, and XR_932996.2-CERS1 may be crucial association pairs for resistance to TB infection. Overall, this study demonstrated that the phagocytic capacity of PBMCs was not a determinant of the resistance phenotype and that some non-coding RNAs could be involved in the natural resistance to TB infection through a ceRNA mechanism.

Characterization of intestinal mononuclear phagocyte subsets in young ruminants at homeostasis and during Cryptosporidium parvum infection.

Introduction: Cryptosporidiosis is a poorly controlled zoonosis caused by an intestinal parasite, Cryptosporidium parvum, with a high prevalence in livestock (cattle, sheep, and goats). Young animals are particularly susceptible to this infection due to the immaturity of their intestinal immune system. In a neonatal mouse model, we previously demonstrated the importance of the innate immunity and particularly of type 1 conventional dendritic cells (cDC1) among mononuclear phagocytes (MPs) in controlling the acute phase of C. parvum infection. These immune populations are well described in mice and humans, but their fine characterization in the intestine of young ruminants remained to be further explored. Methods: Immune cells of the small intestinal Peyer's patches and of the distal jejunum were isolated from naive lambs and calves at different ages. This was followed by their fine characterization by flow cytometry and transcriptomic analyses (q-RT-PCR and single cell RNAseq (lamb cells)). Newborn animals were infected with C. parvum, clinical signs and parasite burden were quantified, and isolated MP cells were characterized by flow cytometry in comparison with age matched control animals. Results: Here, we identified one population of macrophages and three subsets of cDC (cDC1, cDC2, and a minor cDC subset with migratory properties) in the intestine of lamb and calf by phenotypic and targeted gene expression analyses. Unsupervised single-cell transcriptomic analysis confirmed the identification of these four intestinal MP subpopulations in lamb, while highlighting a deeper diversity of cell subsets among monocytic and dendritic cells. We demonstrated a weak proportion of cDC1 in the intestine of highly susceptible newborn lambs together with an increase of these cells within the first days of life and in response to the infection. Discussion: Considering cDC1 importance for efficient parasite control in the mouse model, one may speculate that the cDC1/cDC2 ratio plays also a key role for the efficient control of C. parvum in young ruminants. In this study, we established the first fine characterization of intestinal MP subsets in young lambs and calves providing new insights for comparative immunology of the intestinal MP system across species and for future investigations on host-Cryptosporidium interactions in target species.

Exogenous non-coding dsDNA-dependent trans-activation of phagocytes augments anti-tumor immunity.

Stimulator of interferon genes (STING)-dependent signaling is requisite for effective anti-microbial and anti-tumor activity. STING signaling is commonly defective in cancer cells, which enables tumor cells to evade the immunosurveillance system. We evaluate here whether intrinsic STING signaling in such tumor cells could be reconstituted by creating recombinant herpes simplex viruses (rHSVs) that express components of the STING signaling pathway. We observe that rHSVs expressing STING and/or cGAS replicate inefficiently yet retain in vivo anti-tumor activity, independent of oncolytic activity requisite on the trans-activation of extrinsic STING signaling in phagocytes by engulfed microbial dsDNA species. Accordingly, the in vivo effects of virotherapy could be simulated by nanoparticles incorporating non-coding dsDNA species, which comparably elicit the trans-activation of phagocytes and augment the efficacy of established cancer treatments including checkpoint inhibition and radiation therapy. Our results help elucidate mechanisms of virotherapeutic anti-tumor activity as well as provide alternate strategies to treat cancer.

The Hippo kinases control inflammatory Hippo signaling and restrict bacterial infection in phagocytes.

The Hippo kinases MST1 and MST2 initiate a highly conserved signaling cascade called the Hippo pathway that limits organ size and tumor formation in animals. Intriguingly, pathogens hijack this host pathway during infection, but the role of MST1/2 in innate immune cells against pathogens is unclear. In this report, we generated Mst1/2 knockout macrophages to investigate the regulatory activities of the Hippo kinases in immunity. Transcriptomic analyses identified differentially expressed genes (DEGs) regulated by MST1/2 that are enriched in biological pathways, such as systemic lupus erythematosus, tuberculosis, and apoptosis. Surprisingly, pharmacological inhibition of the downstream components LATS1/2 in the canonical Hippo pathway did not affect the expression of a set of immune DEGs, suggesting that MST1/2 control these genes via alternative inflammatory Hippo signaling. Moreover, MST1/2 may affect immune communication by influencing the release of cytokines, including TNFα, CXCL10, and IL-1ra. Comparative analyses of the single- and double-knockout macrophages revealed that MST1 and MST2 differentially regulate TNFα release and expression of the immune transcription factor MAF, indicating that the two homologous Hippo kinases individually play a unique role in innate immunity. Notably, both MST1 and MST2 can promote apoptotic cell death in macrophages upon stimulation. Lastly, we demonstrate that the Hippo kinases are critical factors in mammalian macrophages and single-cell amoebae to restrict infection by Legionella pneumophila, Escherichia coli, and Pseudomonas aeruginosa. Together, these results uncover non-canonical inflammatory Hippo signaling in macrophages and the evolutionarily conserved role of the Hippo kinases in the anti-microbial defense of eukaryotic hosts. IMPORTANCE: Identifying host factors involved in susceptibility to infection is fundamental for understanding host-pathogen interactions. Clinically, individuals with mutations in the MST1 gene which encodes one of the Hippo kinases experience recurrent infection. However, the impact of the Hippo kinases on innate immunity remains largely undetermined. This study uses mammalian macrophages and free-living amoebae with single- and double-knockout in the Hippo kinase genes and reveals that the Hippo kinases are the evolutionarily conserved determinants of host defense against microbes. In macrophages, the Hippo kinases MST1 and MST2 control immune activities at multiple levels, including gene expression, immune cell communication, and programmed cell death. Importantly, these activities controlled by MST1 and MST2 in macrophages are independent of the canonical Hippo cascade that is known to limit tissue growth and tumor formation. Together, these findings unveil a unique inflammatory Hippo signaling pathway that plays an essential role in innate immunity.

CADHERIN-11 regulation of myeloid phagocytes and autoimmune inflammation in murine lupus.

BACKGROUND AND OBJECTIVE: Understanding the regulation of efferocytosis by myeloid phagocytes is important in identifying novel targets in systemic lupus erythematosus (SLE). Cadherin-11 (CDH11), a cell adhesion molecule, is implicated in inflammatory arthritis and fibrosis and recently been shown to regulate macrophage phagocytosis. The extent and mechanism of this regulation is unknown. Our objective was to examine the extent to which CDH11 regulates myeloid phagocytes and contributes to autoimmunity and tissue inflammation. METHODS: We analyzed efferocytosis in macrophages and dendritic cells (DCs) from WT and Cdh11-/- mice and investigated the mechanisms in vitro. We investigated the role of CDH11 in disease development in vivo using the pristane induced lupus model. To translate the clinical relevance of CDH11 in human disease, we measured serum CDH11 levels in two independent pediatric SLE (pSLE) cohorts and healthy controls. RESULTS: Using bone marrow derived macrophages (BMDMs) and DCs (BMDCs), we found impaired efferocytosis in phagocytes from Cdh11-/- mice, mediated by downregulated efferocytosis receptor expression and RhoGTPase activation. Specifically, loss of CDH11 downregulated Mertk expression and Rac1 activation in BMDMs, and integrin αVß3 expression and Cdc42 activation in BMDCs, highlighting distinct pathways. In vivo, Cdh11-/- mice displayed defective efferocytosis and increased accumulation of apoptotic debris in pristane-induced lupus. Further, Cdh11-/- mice had enhanced systemic inflammation and autoimmune inflammation with increased anti-dsDNA autoantibodies, splenomegaly, type I interferons, and inflammatory cytokines. Paradoxically, at the tissue level, Cdh11-/- mice were protected against glomerulonephritis, indicating a dual role in murine lupus. Finally, SLE patients had increased serum CDH11 compared to controls. CONCLUSION: This study highlights a novel role of CDH11 in regulating myeloid cells and efferocytosis and its potential as a contributor to development in autoimmunity murine lupus. Despite the increase in autoimmunity, Cdh11-/- mice developed decreased tissue inflammation and damage.

Swarna Bhasma Induces Antigen-Presenting Abilities of Macrophages and Helps Antigen Experienced CD4+ T Cells to Acquire Th1 Phenotypes Against Leishmania donovani Antigens.

In leishmaniasis, the protective immunity is largely mediated by proinflammatory cytokine producing abilities of T cells and an efficient parasite killing by phagocytic cells. Notwithstanding a substantial progress that has been made during last decades, the mechanisms or factors involved in establishing protective immunity against Leishmania are not identified. In ancient Indian literature, metallic "bhasma," particularly that of "swarna" or gold (fine gold particles), is indicated as one of the most prominent metal-based therapeutic medicine, which is known to impart protective and curative properties in various health issues. In this work, we elucidated the potential of swarna bhasma (SB) on the effector properties of phagocytes and antigen-activated CD4+ T cells in augmenting the immunogenicity of L. donovani antigens. The characterization of SB revealing its shape, size, composition, and measurement of cytotoxicity established the physiochemical potential for its utilization as an immunomodulator. The activation of macrophages with SB enhanced their capacity to produce nitric oxide and proinflammatory cytokines, which eventually resulted in reduced uptake of parasites and their proliferation in infected cells. Further, in Leishmania-infected animals, SB administration reduced the generation of IL-10, an anti-inflammatory cytokine, and enhanced pro-inflammatory cytokine generation by antigen activated CD4+ T cells with increased frequency of double (IFNγ+/TNFα+) and triple (IFNγ+TNFα+IL-2+) positive cells and abrogated disease pathogeneses at the early days of infection. Our results also suggested that cow-ghee (A2) emulsified preparation of SB, either alone or with yashtimadhu, a known natural immune modulator which enhances the SB's potential in enhancing the immunogenicity of parasitic antigens. These findings suggested a definite potential of SB in enhancing the effector functions of phagocytes and CD4+ T cells against L. donovani antigens. Therefore, more studies are needed to elucidate the mechanistic details of SB and its potential in enhancing vaccine-induced immunity.

Human seminal plasma stimulates the migration of CD11c+ mononuclear phagocytes to the apical side of the colonic epithelium without altering the junctional complexes in an ex vivo human intestinal model.

Introduction: Human immunodeficiency virus type 1 (HIV) transmission mostly occurs through the genital and intestinal mucosae. Although HIV-1 transmission has been extensively investigated, gaps remain in understanding the initial steps of HIV entry through the colonic mucosa. We previously showed that HIV can selectively trigger mononuclear phagocytes (MNP) to migrate within colonic epithelial cells to sample virions. Mucosal exposure to human seminal plasma (HSP), rich in pro- and anti-inflammatory cytokines, chemokines and growth factors, may as well induce alterations of the colonic mucosa and recruit immune cells, hence, affecting pathogen sampling and transmission. Methods: Here, we studied the role of HSP on the paracellular intestinal permeability by analyzing the distribution of two proteins known to play a key role in controlling the intestinal barrier integrity, namely the tight junctions-associated junctional adhesion molecule (JAM-A) and the adherents junction associated protein E-cadherin (E-CAD), by immunofluorescence and confocal microscopy. Also, we evaluated if HSP promotes the recruitment of MNP cells, specifically, the CD11c and CD64 positive MNPs, to the apical side of the human colonic mucosa. At this scope, HSP of HIV-infected and uninfected individuals with known fertility status was tested for cytokines, chemokines and growth factors concentration and used in an ex vivo polarized colonic tissue culture system to mimic as closely as possible the physiological process. Results: HSP showed statistically significant differences in cytokines and chemokines concentrations between the three groups of donors, i.e. HIV infected, or uninfected fertile or randomly identified. Nevertheless, we showed that in the ex vivo tissue culture HSP in general, neither affected the morphological structure of the colonic mucosa nor modulated the paracellular intestinal permeability. Interestingly, CD11c+ MNP cells migrated to the apical surface of the colonic epithelium regardless, if incubated with HIV-infected or -uninfected HSPs, while CD64+ MNP cells, did not change their distribution within the colonic mucosa. Discussion: In conclusion, even if HSP did not perturb the integrity of the human colonic mucosa, it affected the migration of a specific subset of MNPs that express CD11c towards the apical side of the colonic mucosa, which in turn may be involved in pathogen sampling.

Past history of obesity triggers persistent epigenetic changes in innate immunity and exacerbates neuroinflammation.

Age-related macular degeneration is a prevalent neuroinflammatory condition and a major cause of blindness driven by genetic and environmental factors such as obesity. In diseases of aging, modifiable factors can be compounded over the life span. We report that diet-induced obesity earlier in life triggers persistent reprogramming of the innate immune system, lasting long after normalization of metabolic abnormalities. Stearic acid, acting through Toll-like receptor 4 (TLR4), is sufficient to remodel chromatin landscapes and selectively enhance accessibility at binding sites for activator protein-1 (AP-1). Myeloid cells show less oxidative phosphorylation and shift to glycolysis, ultimately leading to proinflammatory cytokine transcription, aggravation of pathological retinal angiogenesis, and neuronal degeneration associated with loss of visual function. Thus, a past history of obesity reprograms mononuclear phagocytes and predisposes to neuroinflammation.

Gata6+ large peritoneal macrophages: an evolutionarily conserved sentinel and effector system for infection and injury.

There are striking similarities between the sea urchin cavity macrophage-like phagocytes (coelomocytes) and mammalian cavity macrophages in not only their location, but also their behaviors. These cells are crucial for maintaining homeostasis within the cavity following a breach, filling the gap and functioning as a barrier between vital organs and the environment. In this review, we summarize the evolving literature regarding these Gata6+ large peritoneal macrophages (GLPMs), focusing on ontogeny, their responses to perturbations, including their rapid aggregation via coagulation, as well as scavenger receptor cysteine-rich domains and their potential roles in diseases, such as cancer. We challenge the 50-year old phenomenon of the 'macrophage disappearance reaction' (MDR) and propose the new term 'macrophage disturbance of homeostasis reaction' (MDHR), which may better describe this complex phenomenon.

Accumulation of Cytochrome b558 at the Plasma Membrane: Hallmark of Oxidative Stress in Phagocytic Cells.

Neutrophils play a very key role in the human immune defense against pathogenic infections. The predominant players in this role during the activation of neutrophils are the release of cytotoxic agents stored in the granules and secretory vesicles and the massive production of reactive oxygen species (ROS) initiated by the enzyme NADPH oxidase. In addition, in living organisms, cells are continuously exposed to endogenous (inflammations, elevated neutrophil presence in the vicinity) and exogenous ROS at low and moderate levels (travels by plane, radiotherapy, space irradiation, blood banking, etc.). To study these effects, we used ROS induced by gamma radiation from low (0.2 Gy) to high (25 Gy) dose levels on PLB-985 cells from a myeloid cell line differentiated to neutrophil-like cells that are considered a good alternative to neutrophils. We determined a much longer lifetime of PLB-985 cells than that of neutrophils, which, as expected, decreased by increasing the irradiation dose. In the absence of any secondary stimulus, a very low production of ROS is detected with no significant difference between irradiated and non-irradiated cells. However, in phagocytosing cells, irradiation doses above 2 Gy enhanced oxidative burst in PLB-985 cells. Whatever the irradiation dose, NADPH oxidase devoid of its cytosolic regulatory units is observed at the plasma membrane in irradiated PLB-985 cells. This result is different from that observed for irradiated neutrophils in which irradiation also induced a translocation of regulatory subunits suggesting that the signal transduction mechanism or pathway operate differently in both cells.

Innate immune deficiencies are associated with severity and poor prognosis in patients with COVID-19.

COVID-19 can cause acute respiratory distress syndrome, leading to death in many individuals. Evidence of a deleterious role of the innate immune system is accumulating, but the precise mechanisms involved remain unclear. In this study, we investigated the links between circulating innate phagocytes and severity in COVID-19 patients. We performed in-depth phenotyping of neutrophil and monocyte subpopulations and measured soluble activation markers in plasma. Additionally, anti-microbial functions (phagocytosis, oxidative burst, and NETosis) were evaluated on fresh cells from patients. Neutrophils and monocytes had a strikingly disturbed phenotype, and elevated concentrations of activation markers (calprotectin, myeloperoxidase, and neutrophil extracellular traps) were measured in plasma. Critical patients had increased CD13low immature neutrophils, LOX-1 + and CCR5 + immunosuppressive neutrophils, and HLA-DRlow downregulated monocytes. Markers of immature and immunosuppressive neutrophils were strongly associated with severity. Moreover, neutrophils and monocytes of critical patients had impaired antimicrobial functions, which correlated with organ dysfunction, severe infections, and mortality. Together, our results strongly argue in favor of a pivotal role of innate immunity in COVID-19 severe infections and pleads for targeted therapeutic options.

SCARF1-Induced Efferocytosis Plays an Immunomodulatory Role in Humans, and Autoantibodies Targeting SCARF1 Are Produced in Patients with Systemic Lupus Erythematosus.

Deficiency in the clearance of cellular debris is a major pathogenic factor in the emergence of autoimmune diseases. We previously demonstrated that mice deficient for scavenger receptor class F member 1 (SCARF1) develop a lupus-like autoimmune disease with symptoms similar to human systemic lupus erythematosus (SLE), including a pronounced accumulation of apoptotic cells (ACs). Therefore, we hypothesized that SCARF1 will be important for clearance of ACs and maintenance of self-tolerance in humans, and that dysregulation of this process could contribute to SLE. In this article, we show that SCARF1 is highly expressed on phagocytic cells, where it functions as an efferocytosis receptor. In healthy individuals, we discovered that engagement of SCARF1 by ACs on BDCA1+ dendritic cells initiates an IL-10 anti-inflammatory response mediated by the phosphorylation of STAT1 and STAT3. Unexpectedly, there was no significant difference in SCARF1 expression in samples of patients with SLE compared with healthy donor samples. However, we detected anti-SCARF1 autoantibodies in 26% of patients with SLE, which was associated with dsDNA Ab positivity. Furthermore, our data show a direct correlation of the levels of anti-SCARF1 in the serum and defects in the removal of ACs. Depletion of Ig restores efferocytosis in SLE serum, suggesting that defects in the removal of ACs are partially mediated by SCARF1 pathogenic autoantibodies. Our data demonstrate that human SCARF1 is an AC receptor in dendritic cells and plays a role in maintaining tolerance and homeostasis.

CircRNA75 and CircRNA72 Function as the Sponge of MicroRNA-200 to Suppress Coelomocyte Apoptosis Via Targeting Tollip in Apostichopus japonicus.

Circular RNAs (circRNAs) act as essential regulators in many biological processes, especially in mammalian immune response. Nonetheless, the functions and mechanisms of circRNAs in the invertebrate immune system are largely unclarified. In our previous work, 261 differentially expressed circRNAs potentially related to the development of Apostichopus japonicus skin ulceration syndrome (SUS), which is a major problem restricting the sea cucumber breeding industry, were identified by genome-wide screening. In this study, via miRanda analysis, both circRNA75 and circrRNA72 were shown to share the miR-200 binding site, a key microRNA in the SUS. The two circRNAs were verified to be increased significantly in LPS-exposed primary coelomocytes, similar to the results of circRNA-seq in sea cucumber under Vibrio splendidus-challenged conditions. A dual-luciferase assay indicated that both circRNA75 and circRNA72 could bind miR-200 in vivo, in which circRNA75 had four binding sites of miR-200 and only one for circRNA72. Furthermore, we found that miR-200 could bind the 3'-UTR of Toll interacting protein (Tollip) to negatively mediate the expression of Tollip. Silencing Tollip increased primary coelomocyte apoptosis. Consistently, inference of circRNA75 and circRNA72 could also downregulate Tollip expression, thereby increasing the apoptosis of primary coelomocytes, which could be blocked by miR-200 inhibitor treatment. Moreover, the rate of si-circRNA75-downregulated Tollip expression was higher than that of si-circRNA72 under an equivalent amount. CircRNA75 and circRNA72 suppressed coelomocyte apoptosis by sponging miR-200 to promote Tollip expression. The ability of circRNA to adsorb miRNA might be positively related to the number of binding sites for miRNA.

Secretome Profiling of Atlantic Salmon Head Kidney Leukocytes Highlights the Role of Phagocytes in the Immune Response to Soluble ß-Glucan.

ß-Glucans (BG) are glucose polymers which are produced in bacteria and fungi but not in vertebrate organisms. Being recognized by phagocytic leukocytes including macrophages and neutrophils through receptors such as dectin-1 and Complement receptor 3 (CR3), the BG are perceived by the innate immune system of vertebrates as foreign substances known as Pathogen Associated Molecular Patterns (PAMPs). The yeast-derived BG has been recognized for its potent biological activity and it is used as an immunomodulator in human and veterinary medicine. The goal of the current study was to characterize the immunostimulatory activity of soluble yeast BG in primary cultures of Atlantic salmon (Salmo salar) head kidney leukocytes (HKLs) in which phagocytic cell types including neutrophils and mononuclear phagocytes predominate. The effect of BG on the secretome of HKL cultures, including secretion of extracellular vesicles (EVs) and soluble protein55s was characterized through western blotting and mass spectrometry. The results demonstrate that, along with upregulation of proinflammatory genes, BG induces secretion of ubiquitinated proteins (UbP), MHCII-containing EVs from professional antigen presenting cells as well as proteins derived from granules of polymorphonuclear granulocytes (PMN). Among the most abundant proteins identified in BG-induced EVs were beta-2 integrin subunits, including CD18 and CD11 homologs, which highlights the role of salmon granulocytes and mononuclear phagocytes in the response to soluble BG. Overall, the current work advances the knowledge about the immunostimulatory activity of yeast BG on the salmon immune system by shedding light on the effect of this PAMP on the secretome of salmon leukocytes.

Identification and Evaluation of Recombinant Outer Membrane Proteins as Vaccine Candidates Against Klebsiella pneumoniae.

Klebsiella pneumoniae found in the normal flora of the human oral and intestinal tract mainly causes hospital-acquired infections but can also cause community-acquired infections. To date, most clinical trials of vaccines against K. pneumoniae have ended in failure. Furthermore, no single conserved protein has been identified as an antigen candidate to accelerate vaccine development. In this study, we identified five outer membrane proteins of K. pneumoniae, namely, Kpn_Omp001, Kpn_Omp002, Kpn_Omp003, Kpn_Omp004, and Kpn_Omp005, by using reliable second-generation proteomics and bioinformatics. Mice vaccinated with these five KOMPs elicited significantly higher antigen-specific IgG, IgG1, and IgG2a. However, only Kpn_Omp001, Kpn_Omp002, and Kpn_Omp005 were able to induce a protective immune response with two K. pneumoniae infection models. These protective effects were accompanied by the involvement of different immune responses induced by KOMPs, which included KOMPs-specific IFN-γ-, IL4-, and IL17A-mediated immune responses. These findings indicate that Kpn_Omp001, Kpn_Omp002, and Kpn_Omp005 are three potential Th1, Th2, and Th17 candidate antigens, which could be developed into multivalent and serotype-independent vaccines against K. pneumoniae infection.