Differential structure and immunomodulatory functions of lipophosphoglycan between Leishmania spp.

Leishmaniasis is a collective term for several tropical, neglected diseases caused by protozoans of the species Leishmania, 20 of which causing disease in humans ranging from localised self-healing lesions to chronic manifestations which affect the skin or inner organs. Although millions of infections are accounted for annually, treatment options are scarce and limited to medication associated with heavy side-effects and increasing antibiotic resistance. Case studies point towards immunotherapy as effective alternative treatment relying on immunomodulatory properties of e.g., the Bacillus Calmette-Guérin vaccine. Leishmania parasites are also known to modulate the immune system, yet the underlying macromolecules and surface molecules remain widely under characterised. With this short review, we aim to provide a complete summary of the existing literature describing one of the most expressed surface molecule on Leishmania spp, lipophosphoglycan (LPG), which shows great variability between different lifecycle stages and different Leishmania spp. Complete characterisation of LPG may aid to improve treatment and aid the development of vaccination strategies, and open new avenues to exploit the immunomodulatory properties of LPG in unrelated conditions.

Leishmania braziliensis enhances monocyte responses to promote anti-tumor activity.

Innate immune cells can undergo long-term functional reprogramming after certain infections, a process called trained immunity (TI). Here, we focus on antigens of Leishmania braziliensis, which induced anti-tumor effects via trained immunity in human monocytes. We reveal that monocytes exposed to promastigote antigens of L. braziliensis develop an enhanced response to subsequent exposure to Toll-like receptor (TLR)2 or TLR4 ligands. Mechanistically, the induction of TI in monocytes by L. braziliensis is mediated by multiple pattern recognition receptors, changes in metabolism, and increased deposition of H3K4me3 at the promoter regions of immune genes. The administration of L. braziliensis exerts potent anti-tumor capabilities by delaying tumor growth and prolonging survival of mice with non-Hodgkin lymphoma. Our work reveals mechanisms of TI induced by L. braziliensis in vitro and identifies its potential for cancer immunotherapy.

Opposing Effects of Interleukin-36γ and Interleukin-38 on Trained Immunity.

Trained immunity is the process of long-term functional reprogramming (a de facto innate immune memory) of innate immune cells such as monocytes and macrophages after an exposure to pathogens, vaccines, or their ligands. The induction of trained immunity is mediated through epigenetic and metabolic mechanisms. Apart from exogenous stimuli, trained immunity can be induced by endogenous compounds such as oxidized LDL, urate, fumarate, but also cytokines including IL-1α and IL-1ß. Here, we show that also recombinant IL-36γ, a pro-inflammatory cytokine of the IL-1-family, is able to induce trained immunity in primary human monocytes, demonstrated by higher cytokine responses and an increase in cellular metabolic pathways both regulated by epigenetic histone modifications. These effects could be inhibited by the IL-36 receptor antagonist as well as by IL-38, an anti-inflammatory cytokine of the IL-1 family which shares its main receptor with IL-36 (IL-1R6). Further, we demonstrated that trained immunity induced by IL-36γ is mediated by NF-κB and mTOR signaling. The inhibitory effect of IL-38 on IL-36γ-induced trained immunity was confirmed in experiments using bone marrow of IL-38KO and WT mice. These results indicate that exposure to IL-36γ results in long-term pro-inflammatory changes in monocytes which can be inhibited by IL-38. Recombinant IL-38 could therefore potentially be used as a therapeutic intervention for diseases characterized by exacerbated trained immunity.

Effect of exogenous IL-37 on immune cells from a patient carrying a potential IL37 loss-of-function variant: A case study.

INTRODUCTION: Chronic inflammatory or autoimmune diseases are commonly treated with immunosuppressive medication such as NSAIDs, corticosteroids, or antibodies against specific cytokines (TNF, IL-1 IL-17, IL-23, etc.) or signalling cascades (e.g. JAK-STAT inhibitors). Using sequencing data to locate genetic mutations in relevant genes allows the identification of alternative targets in a patient-tailored therapy setting. Interleukin (IL)-37 is an anti-inflammatory cytokine with broad effects on innate and adaptive immune cell function. Dysfunctional IL-37 expression or signalling is linked to various autoinflammatory disorders. The administration of recombinant IL-37 to hyperinflammatory patients that are non-responsive to standard treatment bears the potential to alleviate symptoms. METHODS: In this case study, the (hyper)responsiveness of immune cell subsets was investigated in a single patient with a seronegative autoimmune disorder who carries a heterozygous stop-gain variant in IL37 (IL37 Chr2(GRCh37):g.113670640G > A NM_014439.3:c.51G > A p.(Trp17*)). As the patient has been non-responsive to blockage of TNF or IL-1 by Etanercept or Anakinra, respectively, additional in-vitro experiments were set out to elucidate whether treatment with recombinant IL-37 could normalise observed immune cell functions. FINDINGS: Characterisation of immune cell function showed no elevated overall production of acute-phase pro-inflammatory cytokines by patient PBMCs and neutrophils at baseline or upon stimulation. T-cell responses were elevated, as was the metabolic activity and IL-1Ra production of PBMCs at baseline. The identified stop-gain variant in IL37 does not result in the absence of the protein in circulation. In line with this, treatment with recombinant IL-37 did overall not dampen immune responses with the exception of the complete suppression of IL-17. CONCLUSION: The heterozygous stop-gain variant in IL37 (IL37 NM_014439.3:c.51G > A p.(Trp17*)) is not of functional relevance as we observed no clear pro-inflammatory phenotype in immune cells of a patient carrying this variant.

Exploratory analysis of interleukin-38 in hospitalized COVID-19 patients.

INTRODUCTION: A major contributor to coronavirus disease 2019 (COVID-19) progression and severity is a dysregulated innate and adaptive immune response. Interleukin-38 (IL-38) is an IL-1 family member with broad anti-inflammatory properties, but thus far little is known about its role in viral infections. Recent studies have shown inconsistent results, as one study finding an increase in circulating IL-38 in COVID-19 patients in comparison to healthy controls, whereas two other studies report no differences in IL-38 concentrations. METHODS: Here, we present an exploratory, retrospective cohort study of circulating IL-38 concentrations in hospitalized COVID-19 patients admitted to two Dutch hospitals (discovery n = 148 and validation n = 184) and age- and sex-matched healthy subjects. Plasma IL-38 concentrations were measured by enzyme-linked immunosorbent assay, disease-related proteins by proximity extension assay, and clinical data were retrieved from hospital records. RESULTS: IL-38 concentrations were stable during hospitalization and similar to those of healthy control subjects. IL-38 was not associated with rates of intensive care unit admission or mortality. Only in men in the discovery cohort, IL-38 concentrations were positively correlated with hospitalization duration. A positive correlation between IL-38 and the inflammatory biomarker d-dimer was observed in men of the validation cohort. In women of the validation cohort, IL-38 concentrations correlated negatively with thrombocyte numbers. Furthermore, plasma IL-38 concentrations in the validation cohort correlated positively with TNF, TNFRSF9, IL-10Ra, neurotrophil 3, polymeric immunoglobulin receptor, CHL1, CD244, superoxide dismutase 2, and fatty acid binding protein 2, and negatively with SERPINA12 and cartilage oligomeric matrix protein. CONCLUSIONS: These data indicate that IL-38 is not associated with disease outcomes in hospitalized COVID-19 patients. However, moderate correlations between IL-38 concentrations and biomarkers of disease were identified in one of two cohorts. While we demonstrate that IL-38 concentrations are not indicative of COVID-19 severity, its anti-inflammatory effects may reduce COVID-19 severity and should be experimentally investigated.

Circulating interleukin-38 concentrations in healthy adults.

Interleukin (IL)-38 is the latest discovered member of the interleukin-1 family, which has anti-inflammatory properties similar to IL-36Ra. Several studies compared circulating IL-38 concentrations in healthy and diseased populations to characterize its role in both auto-immune and inflammatory pathologies, with both higher and lower concentrations being associated with certain diseases. However, in order to use IL-38 as a biomarker, a reference range in healthy adults is needed. To establish a reference IL-38 circulating concentration, accessible data from 25 eligible studies with IL-38 concentrations in healthy adults was collected. To validate the values found in literature, we measured IL-38 concentrations by enzyme-linked immunosorbent assay (ELISA) in several cohorts from our own institute. Additionally, the effect of blood collection techniques, freeze thawing cycles, and hemolysis on IL-38 measurements was assessed. To evaluate the importance of the genetic background of individuals as confounding factor of IL-38 synthesis, we used publicly available eQTL databases with matched data on allele frequencies in individuals of different ethnicities. Mean IL-38 concentrations in the various studies were weighted by their corresponding sample size, resulting in a weighted mean, and weighted upper and lower limits were calculated by mean ± 2 SD. Differences of over 10.000-fold were found in the weighted means between studies, which could not be attributed to the blood collection method or assessment of IL-38 in plasma or serum. Although IL-38 concentrations were markedly higher in Chinese then in European population studies, we could not show an association with the genetic background. From our analysis, a reference range for circulating IL-38 in healthy adults could thus not yet be established.

IL-1 family cytokines as drivers and inhibitors of trained immunity.

Trained immunity is the long-term memory of innate immune cells, characterised by increased pro-inflammatory responses towards homo- and heterologous secondary stimuli. Interleukin (IL)-1 signalling plays an essential role in the induction of trained immunity, also called innate immune memory. As such, certain anti-inflammatory members of the IL-1 family of cytokines (IL-1F) which interfere with the inflammatory process have the potential to regulate the induction of a trained phenotype. The aim of this review is to provide an update on the role of IL-1F members in the context of trained immunity, emphasising the role of anti-inflammatory cytokines from the IL-1F to inhibit the induction of trained immunity, and touching upon their potential as therapeutics in IL-1-driven inflammatory disorders.

Bimodal Targeting of Human Leukocytes by Fc- and CpG-Decorated Polymersomes to Tune Immune Induction.

The use of well-defined nanovesicles composed of amphiphilic block copolymers (polymersomes) for delivery of adjuvants and antigens is a promising strategy for vaccine development. However, the potency of nanoparticle vaccines depends on efficient interaction with and activation of cells involved in antigen presentation, which can be achieved by targeting cellular receptors. Here, we showed that the Fc fragment display on the polymersome surface resulted in markedly improved interactions with granulocytes, monocytes, and NK cells, while for "naked" polymersomes, virtually no binding to leukocytes was observed. Moreover, CpG-decorated polymersomes were found to also interact with T and/or B cells. Interestingly, whole blood stimulations with Fc fragment and CpG-decorated polymersomes induced interleukin (IL)-6, IL-8, and TNF-α production, while naked polymersomes did not induce any cytokine production. In conclusion, specific immune induction by polymersomes can be controlled using bimodal targeting of different immune receptors, which is an essential feature for targeted vaccine delivery.

The role of IL-32 in Bacillus Calmette-Guérin (BCG)-induced trained immunity in infections caused by different Leishmania spp.

BACKGROUND: Cells of the innate immune system undergo long-term functional reprogramming in response to Bacillus Calmette-Guérin (BCG) exposure via a process called trained immunity, conferring nonspecific protection to unrelated infections. Here, we investigate whether BCG-induced trained immunity is able to protect against infections caused by different Leishmania spp., protozoa that cause cutaneous and mucosal or visceral lesions. METHODS: We used training models of human monocytes with BCG and subsequent infection by L. braziliensis, L. amazonensis and L. infantum, and the vaccination of wild-type and transgenic mice for IL-32γ before in vivo challenge with parasites. RESULTS: We demonstrated that monocytes trained with BCG presented enhanced ability to kill L. braziliensis, L. amazonensis and L. infantum through increased production of reactive oxygen species. Interleukin (IL)-32 appears to play an essential role in the development of trained immunity. Indeed, BCG exposure induced IL-32 production in human primary monocytes, both mRNA and protein. We have used a human IL-32γ transgenic mouse model (IL-32γTg) to study the effect of BCG vaccination in different Leishmania infection models. BCG vaccination decreased lesion size and parasite load in infections caused by L. braziliensis and reduced the spread of L. amazonensis to other organs in both infected wild-type (WT) and IL-32γTg mice. In addition, BCG reduced the parasite load in the spleen, liver and bone marrow of both WT and IL-32γTg mice infected with L. infantum. BCG vaccination increased inflammatory infiltrate in infected tissues caused by different Leishmania spp. In all infections, the presence of IL-32γ was not mandatory, but it increased the protective and inflammatory effects of BCG-induced training. CONCLUSIONS: BCG's ability to train innate immune cells, providing protection against leishmaniasis, as well as the participation of IL-32γ in this process, pave the way for new treatment strategies for this neglected infectious disease.

IL-38 prevents induction of trained immunity by inhibition of mTOR signaling.

Trained immunity is the acquisition of a hyperresponsive phenotype by innate immune cells (such as monocytes and macrophages) after an infection or vaccination, a de facto nonspecific memory dependent on epigenetic and metabolic reprogramming of these cells. We have recently shown that induction of trained immunity is dependent on IL-1ß. Here, we show that recombinant IL-38, an anti-inflammatory cytokine of the IL-1-family, was able to induce long-term inhibitory changes and reduce the induction of trained immunity by ß-glucan in vivo in C57BL/6 mice and ex vivo in their bone marrow cells. IL-38 blocked mTOR signaling and prevented the epigenetic and metabolic changes induced by ß-glucan. In healthy subjects, the IL1F10 associated single nucleotide polymorphism rs58965312 correlated with higher plasma IL-38 concentrations and reduced induction of trained immunity by ß-glucan ex vivo. These results indicate that IL-38 induces long-term anti-inflammatory changes and also inhibits the induction of trained immunity. Recombinant IL-38 could therefore potentially be used as a therapeutic intervention for diseases characterized by exacerbated trained immunity.

Immunotherapeutic Potential of Interleukin-32 and Trained Immunity for Leishmaniasis Treatment.

Neglected tropical diseases annually account for several million infections worldwide. Efficacious treatment for these poorly understood infectious diseases is often limited to ineffective, expensive, and toxic therapies such as the SbV used for leishmaniasis patients. Here, we review the latest discoveries and literature on the molecular pathways, cell types, and immune mediators involved in the immune response to infection with New World Leishmania spp. in humans and their interaction with the adaptive and innate immune system. Novel developments in the field of trained innate immunity and the recently described role of IL-32 are emphasized as potential immunotherapeutic treatments for the management of leishmaniasis.