A Comparative Analysis of Innate Immune Responses and the Structural Characterization of Spike from SARS-CoV-2 Gamma Variants and Subvariants.

The SARS-CoV-2 P.1 variant, responsible for an outbreak in Manaus, Brazil, is distinguished by 12 amino acid differences in the S protein, potentially increasing its ACE-2 affinity and immune evasion capability. We investigated the innate immune response of this variant compared to the original B.1 strain, particularly concerning cytokine production. Blood samples from three severe COVID-19 patients were analyzed post-infection with both strains. Results showed no significant difference in cytokine production of mononuclear cells and neutrophils for either variant. While B.1 had higher cytopathogenicity, neither showed viral replication in mononuclear cells. Structural analyses of the S protein highlighted physicochemical variations, which might be linked to the differences in infectivity between the strains. Our studies point to the increased infectivity of P.1 could stem from altered immunogenicity and receptor-binding affinity.

Bittersweet Sugars: How Unusual Glycan Structures May Connect Epithelial-to-Mesenchymal Transition and Multidrug Resistance in Cancer.

Cancer cells are characterized by metabolic reprogramming, which enables their survival in of-ten inhospitable conditions. A very well-documented example that has gained attraction in re-cent years and is already considered a hallmark of transformed cells is the reprogramming of carbohydrate metabolism. Such a feature, in association with the differential expression of en-zymes involved in the biosynthesis of glycoconjugates, generically known as glycosyltransfer-ases, contributes to the expression of structurally atypical glycans when compared to those ex-pressed in healthy tissues. The latest studies have demonstrated that glycophenotypic alterations are capable of modulating multifactorial events essential for the development and/or progres-sion of the disease. Herein, we will address the importance of glycobiology in modern medi-cine, focusing on the ability of unusual/truncated O-linked glycans to modulate two complex and essential phenomena for cancer progression: the acquisition of the multidrug resistance (MDR) phenotype and the activation of molecular pathways associated with the epithelial-mesenchymal transition (EMT) process, an event deeply linked with cancer metastasis.

Increased Trypanosoma cruzi Growth during Infection of Macrophages Cultured on Collagen I Matrix.

The interactions between cell and cellular matrix confers plasticity to each body tissue, influencing the cellular migratory capacity. Macrophages rely on motility to promote their physiological function. These phagocytes are determinant for the control of invasive infections, and their immunological role largely depends on their ability to migrate and adhere to tissue. Therefore, they interact with the components of the extracellular matrix through their adhesion receptors, conferring morphological modifications that change their shape during migration. Nevertheless, the need to use in vitro cell growth models with the conditioning of three-dimensional synthetic matrices to mimic the dynamics of cell-matrix interaction has been increasingly studied. This becomes more important to effectively understand the changes occurring in phagocyte morphology in the context of infection progression, such as in Chagas disease. This disease is caused by the intracellular pathogen Trypanosoma cruzi, capable of infecting macrophages, determinant cells in the anti-trypanosomatid immunity. In the present study, we sought to understand how an in vitro extracellular matrix model interferes with T. cruzi infection in macrophages. Using different time intervals and parasite ratios, we evaluated the cell morphology and parasite replication rate in the presence of 3D collagen I matrix. Nevertheless, microscopy techniques such as scanning electron microscopy were crucial to trace macrophage-matrix interactions. In the present work, we demonstrated for the first time that the macrophage-matrix interaction favors T. cruzi in vitro replication and the release of anti-inflammatory cytokines during macrophage infection, in addition to drastically altering the morphology of the macrophages and promoting the formation of migratory macrophages.

Special Issue: "Parasitic Infection and Host Immunity": Editorial.

Parasite-host interactions depend on a complex interplay between the metabolism of the parasite, their antigens, and the host immune response system [...].

Increased expression of the pathological O-glycosylated form of oncofetal fibronectin in the multidrug resistance phenotype of cancer cells.

Changes in protein glycosylation are a hallmark of transformed cells and modulate numerous phenomena associated with cancer progression, such as the acquisition of multidrug resistance (MDR) phenotype. Different families of glycosyltransferases and their products have already been described as possible modulators of the MDR phenotype. Among the glycosyltransferases intensively studied in cancer research, UDP-N-acetyl-d-galactosamine:polypeptide N-acetylgalactosaminyltransferase-6 (pp-GalNAc-T6), which is widely expressed in many organs and tissues, stands out. Its influence in several events associated with kidney, oral, pancreatic, renal, lung, gastric and breast cancer progression has already been described. However, its participation in the MDR phenotype has never been studied. Here, we demonstrate that human breast adenocarcinoma MCF-7 MDR cell lines, generated by chronic exposure to doxorubicin, in addition to exhibiting increased expression of proteins belonging to the ABC superfamily (ABCC1 and ABCG2), and anti-apoptotic proteins (Blcl-2 and Bcl-xL), also present high expression of pp-GalNAc-T6, the enzyme currently proposed as the main responsible for the biosynthesis of oncofetal fibronectin (onf-FN), a major extracellular matrix component expressed by cancer cells and embryonic tissues, but absent in healthy cells. Our results show that onf-FN, which is generated by the addition of a GalNAc unit at a specific threonine residue inside the type III homology connective segment (IIICS) domain of FN, is strongly upregulated during the acquisition of the MDR phenotype. Also, the silencing of pp-GalNAc-T6, not only compromises the expression of the oncofetal glycoprotein, but also made the MDR cells more sensitive to all anticancer drugs tested, partially reversing the MDR phenotype. Taken together, our results demonstrate for the first time the upregulation of the O-glycosylated oncofetal fibronectin, as well as the direct participation of pp-GalNAc-T6 during the acquisition of a MDR phenotype in a breast cancer model, giving credence to the hypothesis that in transformed cells, glycosyltransferases and/or their products, such as unusual extracellular matrix glycoproteins can be used as potential therapeutic targets for the treatment of cancer.

The Blessed Union of Glycobiology and Immunology: A Marriage That Worked.

In this article, we discuss the main aspects regarding the recognition of cell surface glycoconjugates and the immunomodulation of responses against the progression of certain pathologies, such as cancer and infectious diseases. In the first part, we talk about different aspects of glycoconjugates and delve deeper into the importance of N-glycans in cancer immunotherapy. Then, we describe two important lectin families that have been very well studied in the last 20 years. Examples include the sialic acid-binding immunoglobulin (Ig)-like lectins (siglecs), and galectins. Finally, we discuss a topic that needs to be better addressed in the field of glycoimmunology: the impact of oncofetal antigens on the cells of the immune system. New findings in this area are of great importance for advancement, especially in the field of oncology, since it is already known that cellular interactions mediated by carbohydrate-carbohydrate and/or carbohydrate proteins are able to modulate the progression of different types of cancer in events that compromise the functionality of the immune responses.

Non-self glycan structures as possible modulators of cancer progression: would polysaccharides from Cryptococcus spp. impact this phenomenon?

Invasive fungal infections (IFI) are responsible for a large number of annual deaths. Most cases are closely related to patients in a state of immunosuppression, as is the case of patients undergoing chemotherapy. Cancer patients are severely affected by the worrisome proportions that an IFI can take during cancer progression, especially in an already immunologically and metabolically impaired patient. There is scarce knowledge about strategies to mitigate cancer progression in these cases, beyond conventional treatment with antifungal drugs with a narrow therapeutic range. However, in recent years, ample evidence has surfaced describing the possible interferences that IFI may have both on the progression of pre-existing cancers and in the induction of newly transformed cells. The leading gambit for modulation of tumor progression comes from the ability of fungal virulence factors to modulate the host's immune system, since they are found in considerable concentrations in the tumor microenvironment during infection. In this context, cryptococcosis is of particular concern, since the main virulence factor of the pathogenic yeast is its polysaccharide capsule, which carries constituents with high immunomodulatory properties and cytotoxic potential. Therefore, we open a discussion on what has already been described regarding the progression of cryptococcosis in the context of cancer progression, and the possible implications that fungal glycan structures may take in both cancer development and progression.

Neuraminidase is a host-directed approach to regulate neutrophil responses in sepsis and COVID-19.

BACKGROUND AND PURPOSE: Neutrophil overstimulation plays a crucial role in tissue damage during severe infections. Because pathogen-derived neuraminidase (NEU) stimulates neutrophils, we investigated whether host NEU can be targeted to regulate the neutrophil dysregulation observed in severe infections. EXPERIMENTAL APPROACH: The effects of NEU inhibitors on lipopolysaccharide (LPS)-stimulated neutrophils from healthy donors or COVID-19 patients were determined by evaluating the shedding of surface sialic acids, cell activation, and reactive oxygen species (ROS) production. Re-analysis of single-cell RNA sequencing of respiratory tract samples from COVID-19 patients also was carried out. The effects of oseltamivir on sepsis and betacoronavirus-induced acute lung injury were evaluated in murine models. KEY RESULTS: Oseltamivir and zanamivir constrained host NEU activity, surface sialic acid release, cell activation, and ROS production by LPS-activated human neutrophils. Mechanistically, LPS increased the interaction of NEU1 with matrix metalloproteinase 9 (MMP-9). Inhibition of MMP-9 prevented LPS-induced NEU activity and neutrophil response. In vivo, treatment with oseltamivir fine-tuned neutrophil migration and improved infection control as well as host survival in peritonitis and pneumonia sepsis. NEU1 also is highly expressed in neutrophils from COVID-19 patients, and treatment of whole-blood samples from these patients with either oseltamivir or zanamivir reduced neutrophil overactivation. Oseltamivir treatment of intranasally infected mice with the mouse hepatitis coronavirus 3 (MHV-3) decreased lung neutrophil infiltration, viral load, and tissue damage. CONCLUSION AND IMPLICATIONS: These findings suggest that interplay of NEU1-MMP-9 induces neutrophil overactivation. In vivo, NEU may serve as a host-directed target to dampen neutrophil dysfunction during severe infections.

Expression of O-glycosylated oncofetal fibronectin in alternatively activated human macrophages.

Macrophage (Mϕ) polarization is an essential phenomenon for the maintenance of homeostasis and tissue repair, and represents the event by which Mϕ reach divergent functional phenotypes as a result to specific stimuli and/or microenvironmental signals. Mϕ can be polarized into two main phenotypes, M1 or classically activated and M2 or alternatively activated. These two categories diverge in many aspects, such as secreted cytokines, markers of cell surface, and biological functions. Over the last 10 years, many potential markers have been proposed for both M1 and M2 human Mϕ. However, there is scarce information regarding the glycophenotype adopted by these cells. Here, we show that M2- but not M1-polarized Mϕ expresses high levels of an unusual glycoform of fibronectin (FN), named O-glycosylated oncofetal FN (onf-FN), found in fetal/cancer cells, but not in healthy tissues. The onf-FN expression was confirmed in vitro by Western blot and real-time RT-qPCR in primary and cell line monocyte-derived Mϕ. onf-FN was induced by IL-4 and IL-13, but not by pro-inflammatory stimuli (LPS and INF-γ). RNA and protein analysis clearly demonstrated that it is specifically associated with the M2 polarization. In conclusion, we show by the first time that O-glycosylated onf-FN is expressed by M2-polarized Mϕ.

Humoral response in Leishmaniasis.

Leishmaniasis presents different types of clinical manifestations that can be divided into cutaneous leishmaniasis and visceral leishmaniasis. The host's immune system, associated with genetic and nutritional factors, is strongly involved in the evolution of the disease or parasite escape. Humoral immunity is characterized by the production of antibodies capable of promoting neutralization, opsonization, and activation of the complement system. In this scenario, B lymphocytes produce antibodies that play an important role in Leishmania infection although neglected for a long time. Thus, relevant aspects in the establishment of Leishmania infection will be addressed, highlighting the importance of humoral immunity during the entire process of Leishmania infection.

Pulmonary Fibrosis and Hypereosinophilia in TLR9-/- Mice Infected by Cryptococcus gattii.

Cryptococcus gattii is a worldwide-distributed basidiomycetous yeast that can infect immunocompetent hosts. However, little is known about the mechanisms involved in the disease. The innate immune response is essential to the control of infections by microorganisms. Toll-like receptor 9 (TLR9) is an innate immune receptor, classically described as a non-methylated DNA recognizer and associated with bacteria, protozoa and opportunistic mycosis infection models. Previously, our group showed that TLR9-/- mice were more susceptible to C. gattii after 21 days of infection. However, some questions about the innate immunity involving TLR9 response against C. gattii remain unknown. In order to investigate the systemic cryptococcal infection, we evaluated C57BL/6 mice and C57BL/6 TLR9-/- after intratracheal infection with 104C. gattii yeasts for 21 days. Our data evidenced that TLR9-/- was more susceptible to C. gattii. TLR9-/- mice had hypereosinophilia in pulmonary mixed cellular infiltrate, severe bronchiolitis and vasculitis and type 2 alveolar cell hyperplasia. In addition, TLR9-/- mice developed severe pulmonary fibrosis and areas with strongly birefringent fibers. Together, our results corroborate the hypothesis that TLR9 is important to support the Th1/Th17 response against C. gattii infection in the murine experimental model.

SARS-CoV-2's Variants of Concern: A Brief Characterization.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) disclose the variants of concern (VOC) including Alpha (B.1.1.7), Beta (B.1.351), Gamma (P1), Delta (B.1.617.2), and Omicron (B.1.1.529). Its spike protein (S) present on the surface of the virus is recognized by the host cell receptor, the angiotensin-2 converting enzyme (ACE2) which promotes their entry into the cell. The mutations presented by VOCs are found in RBD and the N-terminal region of S protein. Therefore, mutations occurring in RBD can modify the biological and immunogenic characteristics of the virus, such as modifying the spike affinity for ACE2, increasing the virus transmissibility, or conferring the ability to escape the immune responses. The raise of a potential new SARS-CoV-2 variant capable of evading the host defenses at the same time maintaining its fitness justifies the importance of continued genetic monitoring of the pandemic coronavirus.

Cellular Stress and Senescence Induction during Trypanosoma cruzi Infection.

Chagas disease (CD) is a neglected tropical disease caused by Trypanosoma cruzi infection that, despite being discovered over a century ago, remains a public health problem, mainly in developing countries. Since T. cruzi can infect a wide range of mammalian host cells, parasite-host interactions may be critical to infection outcome. The intense immune stimulation that helps the control of the parasite's replication and dissemination may also be linked with the pathogenesis and symptomatology worsening. Here, we discuss the findings that support the notion that excessive immune system stimulation driven by parasite persistence might elicit a progressive loss and collapse of immune functions. In this context, cellular stress and inflammatory responses elicited by T. cruzi induce fibroblast and other immune cell senescence phenotypes that may compromise the host's capacity to control the magnitude of T. cruzi-induced inflammation, contributing to parasite persistence and CD progression. A better understanding of the steps involved in the induction of this chronic inflammatory status, which disables host defense capacity, providing an extra advantage to the parasite and predisposing infected hosts prematurely to immunosenescence, may provide insights to designing and developing novel therapeutic approaches to prevent and treat Chagas disease.

Glycobiology of Cancer: Sugar Drives the Show.

Cancer development and progression is associated with aberrant changes in cellular glycosylation. Cells expressing altered glycan-structures are recognized by cells of the immune system, favoring the induction of inhibitory immune processes which subsequently promote tumor growth and spreading. Here, we discuss about the importance of glycobiology in modern medicine, taking into account the impact of altered glycan structures expressed in cancer cells as potential glycobiomarkers of disease, as well as on cancer development and progression.

The Sweet Side of Fungal Infections: Structural Glycan Diversity and Its Importance for Pathogenic Adaptation.

Fungal infections are the most common secondary infections in debilitated individuals in a state of chronic disease or immunosuppression. Despite this, most fungal infections are neglected, mainly due to the lower frequency of their more severe clinical forms in immunocompetent individuals with a healthy background. However, over the past few years, several cases of severe fungal infections in healthy individuals have provoked a change in the epidemiological dynamics of fungal infections around the world, both due to recurrent outbreaks in previously infrequent regions and the greater emergence of more pathogenic fungal variants affecting healthy individuals, such as in the Cryptococcus genus. Therefore, before the arrival of a scenario of prevalent severe fungal infections, it is necessary to assess more carefully what are the real reasons for the increased incidence of fungal infection globally. What are the factors that are currently contributing to this new possible epidemiological dynamic? Could these be of a structural nature? Herein, we propose a discussion based on the importance of the virulence factors of glycoconjugate composition in the adaptation of pathogenic fungal species into the current scenario of increasing severity of these infections.

Eryptosis as a New Insight in Malaria Pathogenesis.

Eryptosis is a programmed cell death-like process that occurs in red blood cells. Although the red blood cells are anucleated, there are similarities between eryptosis and apoptosis, such as increased calcium efflux, calpain activation, phosphatidylserine exposure, cell blebbing and cell shrinkage. Eryptosis occurs physiologically in red blood cells, as a consequence of the natural senescence process of these cells, but it can also be stimulated in pathological situations such as metabolic syndromes, uremic syndromes, polycythemia vera, anemias such as sickle cell anemia and thalassemia, and infectious processes including Plasmodium infection. Infection-induced eryptosis is believed to contribute to damage caused by Plasmodium, but it's still a topic of debate in the literature. In this review, we provided an overview of eryptosis mechanisms and its possible pathogenic role in malaria.

Immune Responses in Leishmaniasis: An Overview.

Leishmaniasis is a parasitic, widespread, and neglected disease that affects more than 90 countries in the world. More than 20 Leishmania species cause different forms of leishmaniasis that range in severity from cutaneous lesions to systemic infection. The diversity of leishmaniasis forms is due to the species of parasite, vector, environmental and social factors, genetic background, nutritional status, as well as immunocompetence of the host. Here, we discuss the role of the immune system, its molecules, and responses in the establishment, development, and outcome of Leishmaniasis, focusing on innate immune cells and Leishmania major interactions.

Bone marrow granulocytes downregulate IL-1ß and TNF production and the microbicidal activity of inflammatory macrophages.

Macrophages play critical roles in inflammation and defense against pathogens, as well as in the return to tissue homeostasis. Macrophage subpopulations displaying antagonistic phenotypes are generally classified as proinflammatory M1, implicated in antipathogen and antitumoral activities, or as anti-inflammatory M2, associated with tissue repair. Granulocytic and monocytic myeloid-derived suppressor cells recruited from the bone marrow to tissues and phagocytosis of apoptotic neutrophils can attenuate macrophage microbicidal activity. Here, we showed that bone marrow neutrophils, but not thioglycollate-recruited neutrophils, directly suppress the responses of macrophages that were previously committed to an inflammatory phenotype. Cocultures of inflammatory macrophages with bone marrow CD11b+Ly6Ghi granulocytes led to reduced release of IL-1ß, TNF-α, and IL-6 by macrophages after lipopolysaccharide stimulation. The suppressive activity was unrelated to granulocyte apoptosis or to secreted factors and required cell-to-cell contact. The suppressive effect was paralleled by reduction in the nuclear levels of the NF-κB p65 subunit, but not of the p50 subunit. Furthermore, bone marrow granulocytes decreased the phagocytic activity of macrophages and their capacity to kill intracellular Escherichia coli. Taken together, these results show that bone marrow granulocytes can function as suppressors of the proinflammatory activity and microbial-killing responses of macrophages.

Trypanosoma brucei rhodesiense Inhibitor of Cysteine Peptidase (ICP) Is Required for Virulence in Mice and to Attenuate the Inflammatory Response.

The protozoan Trypanosoma brucei rhodesiense causes Human African Trypanosomiasis, also known as sleeping sickness, and penetrates the central nervous system, leading to meningoencephalitis. The Cathepsin L-like cysteine peptidase of T. b. rhodesiense has been implicated in parasite penetration of the blood-brain barrier and its activity is modulated by the chagasin-family endogenous inhibitor of cysteine peptidases (ICP). To investigate the role of ICP in T. b. rhodesiense bloodstream form, ICP-null (Δicp) mutants were generated, and lines re-expressing ICP (Δicp:ICP). Lysates of Δicp displayed increased E-64-sensitive cysteine peptidase activity and the mutant parasites traversed human brain microvascular endothelial cell (HBMEC) monolayers in vitro more efficiently. Δicp induced E-selectin in HBMECs, leading to the adherence of higher numbers of human neutrophils. In C57BL/6 mice, no Δicp parasites could be detected in the blood after 6 days, while mice infected with wild-type (WT) or Δicp:ICP displayed high parasitemia, peaking at day 12. In mice infected with Δicp, there was increased recruitment of monocytes to the site of inoculation and higher levels of IFN-γ in the spleen. At day 14, mice infected with Δicp exhibited higher preservation of the CD4+, CD8+, and CD19+ populations in the spleen, accompanied by sustained high IFN-γ, while NK1.1+ populations receded nearly to the levels of uninfected controls. We propose that ICP helps to downregulate inflammatory responses that contribute to the control of infection.