Early antitumor activity of oral Langerhans cells is compromised by a carcinogen

Early diagnosis of oral squamous cell carcinoma (OSCC) remains an unmet clinical need. Therefore, elucidating the initial events of OSCC preceding tumor development could benefit OSCC prognosis. Here, we define the Langerhans cells (LCs) of the tongue and demonstrate that LCs protect the epithelium from carcinogen-induced OSCC by rapidly priming αβT cells capable of eliminating γH2AX+ epithelial cells, whereas γδT and natural killer cells are dispensable. The carcinogen, however, dysregulates the epithelial resident mononuclear phagocytes, reducing LC frequencies, while dendritic cells (DCs), macrophages, and plasmacytoid DCs (pDCs) populate the epithelium. Single-cell RNA-sequencing analysis indicates that these newly differentiated cells display an immunosuppressive phenotype accompanied by an expansion of T regulatory (Treg) cells. Accumulation of the Treg cells was regulated, in part, by pDCs and precedes the formation of visible tumors. This suggests LCs play an early protective role during OSCC, yet the capacity of the carcinogen to dysregulate the differentiation of mononuclear phagocytes facilitates oral carcinogenesis.

CCR2 promotes monocyte recruitment and intestinal inflammation in mice lacking the interleukin-10 receptor

Macrophages are a heterogeneous population of mononuclear phagocytes abundantly distributed throughout the intestinal compartments that adapt to microenvironmental specific cues. In adult mice, the majority of intestinal macrophages exhibit a mature phenotype and are derived from blood monocytes. In the steady-state, replenishment of these cells is reduced in the absence of the chemokine receptor CCR2. Within the intestine of mice with colitis, there is a marked increase in the accumulation of immature macrophages that demonstrate an inflammatory phenotype. Here, we asked whether CCR2 is necessary for the development of colitis in mice lacking the receptor for IL10. We compared the development of intestinal inflammation in mice lacking IL10RA or both IL10RA and CCR2. The absence of CCR2 interfered with the accumulation of immature macrophages in IL10R-deficient mice, including a novel population of rounded submucosal Iba1+ cells, and reduced the severity of colitis in these mice. In contrast, the absence of CCR2 did not reduce the augmented inflammatory gene expression observed in mature intestinal macrophages isolated from mice lacking IL10RA. These data suggest that both newly recruited CCR2-dependent immature macrophages and CCR2-independent residual mature macrophages contribute to the development of intestinal inflammation observed in IL10R-deficient mice.

Hypoxia Inhibits Subretinal Inflammation Resolution Thrombospondin-1 Dependently

Hypoxia is potentially one of the essential triggers in the pathogenesis of wet age-related macular degeneration (wetAMD), characterized by choroidal neovascularization (CNV) which is driven by the accumulation of subretinal mononuclear phagocytes (MP) that include monocyte-derived cells. Here we show that systemic hypoxia (10% O2) increased subretinal MP infiltration and inhibited inflammation resolution after laser-induced subretinal injury in vivo. Accordingly, hypoxic (2% O2) human monocytes (Mo) resisted elimination by RPE cells in co-culture. In Mos from hypoxic mice, Thrombospondin 1 mRNA (Thbs1) was most downregulated compared to normoxic animals and hypoxia repressed Thbs-1 expression in human monocytes in vitro. Hypoxic ambient air inhibited MP clearance during the resolution phase of laser-injury in wildtype animals, but had no effect on the exaggerated subretinal MP infiltration observed in normoxic Thbs1−/−-mice. Recombinant Thrombospondin 1 protein (TSP-1) completely reversed the pathogenic effect of hypoxia in Thbs1−/−-mice, and accelerated inflammation resolution and inhibited CNV in wildtype mice. Together, our results demonstrate that systemic hypoxia disturbs TSP-1-dependent subretinal immune suppression and promotes pathogenic subretinal inflammation and can be therapeutically countered by local recombinant TSP-1.

Spatial and molecular profiling of the classic Hodgkin lymphoma microenvironment reveals an immunosuppressive mononuclear phagocyte network

Although a lymph node infiltrated by classic Hodgkin lymphoma is mostly composed of non-neoplastic immune cells, the malignant Hodgkin Reed-Sternberg cells (HRSC) successfully suppress an anti-tumor immune response, to create a cancer-permissive microenvironment. Accordingly, unleashing the dormant immune cells, for example by checkpoint inhibition, has been a central focus of recent therapeutic advances for this disease. Here, we profiled the global immune cell composition of normal and diseased lymph nodes by single-cell RNA sequencing, as a basis for interrogating the immediate vicinity of HRSC, first regionally and then at cellular resolution. Our analyses revealed specific immune cells and functional states associated with HRSC. Most prominently, we discovered a non-random spatial association of immunoregulatory mononuclear phagocytes positioned around HRSC, which express the immune checkpoints PD-L1, TIM-3, and the tryptophan-catabolizing protein IDO1. These findings provide a basis for rational targeting and activation of the anti-tumor immune response in classic Hodgkin lymphoma.

Inherent P2X7 Receptors Regulate Macrophage Functions during Inflammatory Diseases

Macrophages are mononuclear phagocytes which derive either from blood-borne monocytes or reside as resident macrophages in peripheral (Kupffer cells of the liver, marginal zone macrophages of the spleen, alveolar macrophages of the lung) and central tissue (microglia). They occur as M1 (pro-inflammatory; classic) or M2 (anti-inflammatory; alternatively activated) phenotypes. Macrophages possess P2X7 receptors (Rs) which respond to high concentrations of extracellular ATP under pathological conditions by allowing the non-selective fluxes of cations (Na+, Ca2+, K+). Activation of P2X7Rs by still higher concentrations of ATP, especially after repetitive agonist application, leads to the opening of membrane pores permeable to ~900 Da molecules. For this effect an interaction of the P2X7R with a range of other membrane channels (e.g., P2X4R, transient receptor potential A1 [TRPA1], pannexin-1 hemichannel, ANO6 chloride channel) is required. Macrophage-localized P2X7Rs have to be co-activated with the lipopolysaccharide-sensitive toll-like receptor 4 (TLR4) in order to induce the formation of the inflammasome 3 (NLRP3), which then activates the pro-interleukin-1β (pro-IL-1β)-degrading caspase-1 to lead to IL-1β release. Moreover, inflammatory diseases (e.g., rheumatoid arthritis, Crohn’s disease, sepsis, etc.) are generated downstream of the P2X7R-induced upregulation of intracellular second messengers (e.g., phospholipase A2, p38 mitogen-activated kinase, and rho G proteins). In conclusion, P2X7Rs at macrophages appear to be important targets to preserve immune homeostasis with possible therapeutic consequences.

Treatment with IgM-enriched intravenous immunoglobulins (IgM-IVIg) enhances clearance of stroke-associated bacterial lung infection

Post-stroke infection is a common complication of stroke that is associated with increased mortality and morbidity. We previously found that experimental stroke induces an ablation of multiple sub-populations of B cells and reduced levels of IgM antibody that coincide with the development of spontaneous bacterial pneumonia. Reduced circulating IgM concentrations were also observed in acute stroke patients. The loss of IgM antibody after stroke could be an important determinant of infection susceptibility and highlights this pathway as an important target for intervention. We treated mice with a low (replacement), dose of IgM-enriched intravenous immunoglobulin (IgM-IVIg) prior to and 24 h after experimental stroke induced by middle cerebral artery occlusion (MCAO) or sham surgery, then recovered mice for 2 d or 5 d. The effect of treatment on lung bacterial burden, lung pathology, brain infarct volume, antibody levels and both lung and systemic cellular and cytokine immune profiles was determined. Treatment with IgM-IVIg enhanced bacterial clearance from the lung after MCAO and improved pathology but did not impact infarct volume. IgM-IVIg treatment induced immunomodulatory effects systemically including rescue of splenic plasma B cell numbers and endogenous mouse IgM and IgA circulating immunoglobulin concentrations that were reduced by MCAO, and treatment also reduced concentrations of pro-inflammatory cytokines in the lung. The effects of MCAO and IgM-IVIg treatment on the immune system were tissue specific as no impact on B cells or mouse immunoglobulins were found within the lung. However, the presence of human immunoglobulins from the IgM-IVIg treatment led to increased total lung immunoglobulin concentration. IgM-IVIg treatment did not increase the number of lung mononuclear phagocytes or directly modulate macrophage phagocytic capacity but enhanced their capability to phagocytose Staphylococcus aureus bioparticles in vitro by increasing opsonisation. Low dose IgM-IVIg contributes to increased clearance of spontaneous lung bacteria after MCAO likely via increasing availability of antibody in the lung to enhance phagocytic activity. Immunomodulatory effects of IgM-IVIg treatment, including reduced pro-inflammatory cytokine production, may also contribute to reduced levels of damage in the lung after MCAO. IgM-IVIg shows promise as an antibacterial and immunomodulatory agent to use in the treatment of post-stroke infection.

Myeloid Immune Cells CARriying a New Weapon Against Cancer

Chimeric antigen receptor (CAR) engineering for T cells and natural killer cells (NK) are now under clinical evaluation for the treatment of hematologic cancers. Although encouraging clinical results have been reported for hematologic diseases, pre-clinical studies in solid tumors have failed to prove the same effectiveness. Thus, there is a growing interest of the scientific community to find other immune cell candidate to express CAR for the treatment of solid tumors and other diseases. Mononuclear phagocytes may be the most adapted group of cells with potential to overcome the dense barrier imposed by solid tumors. In addition, intrinsic features of these cells, such as migration, phagocytic capability, release of soluble factors and adaptive immunity activation, could be further explored along with gene therapy approaches. Here, we discuss the elements that constitute the tumor microenvironment, the features and advantages of these cell subtypes and the latest studies using CAR-myeloid immune cells in solid tumor models.

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

β‐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.

Increasing the Therapeutic Efficacy of Extracellular Vesicles From the Antigen-Specific Antibody and Light Chain Perspective

Due to their exceptional properties, extracellular vesicles (EVs) receive special attention as next generation biotherapeutics and vehicles for drug delivery. However, despite having many advantages over cell-based therapies, EVs usually exert lower therapeutic efficacy. This results from a number of hurdles that are faced by the EV-based approaches. Administered EVs could be rapidly cleared by the mononuclear phagocytes as well as can randomly distribute within various tissues, making tissue penetration and cell targeting insufficient. However, recent research findings imply that these limitations could be overcome with the use of antigen-specific antibodies and light chains. Major histocompatibility complex (MHC) class II-expressing EVs have been shown to form aggregates after co-incubation with antigen-specific antibodies, which greatly enhanced their biological efficacy. On the other hand, EVs could be coated with antibody light chains of chosen specificity to direct them towards desired target cell population. Both findings open up a promising perspective to achieve the highest efficacy of the EV-based approaches. Herein we discuss the opportunities for enhancing extracellular vesicle’s biological activity by using specific antibodies and light chains in the context of the challenges faced by such therapeutic approach.

Immunotherapy With 5, 15-DPP Mediates Macrophage M1 Polarization and Modulates Subsequent Mycobacterium tuberculosis Infectivity in rBCG30 Immunized Mice

Tuberculosis (TB) is a significant and continuing problem worldwide, with a death toll of around 1.5 million human lives annually. BCG, the only vaccine against TB, offers a varied degree of protection among human subjects in different regions and races of the world. The majority of the population living near the tropics carries a varying degree of tolerance against BCG due to the widespread prevalence of non-tuberculous mycobacteria (NTM). Interestingly, ≈90% of the Mycobacterium tuberculosis (Mtb) infected population restrain the bacilli on its own, which strengthens the notion of empowering the host immune system to advance the protective efficacy of existing mycobacterial vaccines. In general, Mtb modulates IL-10/STAT3 signaling to skew host mononuclear phagocytes toward an alternatively activated, anti-inflammatory state that helps it thrive against hostile immune advances. We hypothesized that modulating the IL-10/STAT3 driven anti-inflammatory effects in mononuclear cells may improve the prophylactic ability of TB vaccines. This study investigated the immunotherapeutic ability of a porphyrin based small molecule inhibitor of IL-10/STAT3 axis, 5, 15-diphenyl porphyrin (DPP), in improving anti-TB immunity offered by second generation recombinant BCG30 (rBCG30-ARMF-II®) vaccine in mice. The DPP therapy potentiated vaccine induced anti-TB immunity by down-modulating anti-inflammatory responses, while simultaneously up-regulating pro-inflammatory immune effector responses in the immunized host. The employed DPP based immunotherapy led to the predominant activation/proliferation of pro-inflammatory monocytes/macrophages/DCs, the concerted expansion of CD4+/CD8+ effector and central memory T cells, alongside balanced Th17 and Treg cell amplification, and conferred augmented resistance to aerosol Mtb challenge in rBCG30 immunized BALB/c mice.