D. Channe Gowda

Malaria infection causes multiple organ-specific lethal pathologies, including cerebral malaria, and severe liver and lung pathologies by inducing strong inflammatory responses. Gene polymorphism studies suggest that TLR4 and TLR2 contribute to severe malaria, but the roles of these signaling molecules in malaria pathogenesis remain incompletely understood. We hypothesize that danger-associated molecular patterns (DAMPs) produced in response to malaria activate TLR2 and TLR4 signaling and contribute to liver and lung pathologies. By using a mouse model of Plasmodium berghei NK65 infection, we show that the combined TLR2 and TLR4 signaling contributes to malaria liver and lung pathologies, and mortality. Macrophages, neutrophils, NK cells, and T cells infiltrate to the livers and lungs of infected wild type (WT) mice more than TLR2,4^-/- mice. Additionally, endothelial barrier disruption, tissue necrosis, and hemorrhage were higher in the livers and lungs of infected WT mice than in those of TLR2,4^-/- mice. Consistent with these results, the levels of chemokine production, chemokine receptor expression, and liver and lung pathological markers were higher in infected WT mice than in TLR2,4^-/- mice. In addition, the levels of HMGB1, a potent TLR2- and TLR4-activating DAMP, were higher in livers and lungs of WT mice than TLR2,4^-/- mice. Treatment with glycyrrhizin, an immunomodulatory agent known to inhibit HMGB1 activity, markedly reduced mortality in WT mice. These results suggest that TLR2 and TLR4 activation by HMGB1 and possibly other endogenously produced DAMPs contribute to malaria liver and lung injury via signaling mechanisms distinct from those involved in cerebral malaria pathogenesis.

This chapter provides an overview of structures and functions of complex carbohydrates (commonly called glycans) that are covalently linked to proteins or lipids to form glycoconjugates known as glycoproteins, glycolipids, and proteoglycans. To understand the complexity of the glycan structures, the nature of their monosaccharide building blocks, how the monomeric units are covalently linked to each other, and how the resulting glycans are attached to proteins or lipids are discussed. Then, the classification, nomenclature, structural features, and functions of the glycan moieties of animal glycoconjugates are briefly described. All three classes of glycoconjugates are constituents of plasma membranes of all animal cells, including those of the nervous system. Glycoproteins and proteoglycans are also found abundantly as constituents of tissue matrices. Additionally, glycan-rich mucin glycoproteins are the major constituents of mucus secretions of epithelia of various organs. Furthermore, the chapter draws attention to the incredible structural complexity and diversity of the glycan moieties of cell surface and extracellular glycoconjugates. Finally, the involvement of glycans as informational molecules in a wide range of essential functions in almost all known biological processes, which are crucial for development, differentiation, and normal functioning of animals, is discussed. Keywords Complex carbohydrates N-Glycans O-Glycans Glycosaminoglycans Glycoconjugates Glycoproteins Glycolipids GPI anchors Proteoglycans Structure and functions of glycoconjugates

Cytokine responses to malaria play important roles in both protective immunity development and pathogenesis. Although the roles of cytokines such as TNF-α, IL-12, IFN-γ, and IL-10 in immunity and pathogenesis to the blood stage malaria are largely known, the role of IL-4 remains less understood. IL-4 targets many cell types and induces multiple effects, including cell proliferation, gene expression, protection from apoptosis, and immune regulation. Accordingly, IL-4 has been exploited as a therapeutic for several inflammatory diseases. Malaria caused by Plasmodium falciparum manifests in many organ-specific fatal pathologies, including cerebral malaria (CM), driven by a high parasite load, leading to parasite sequestration in organs and consequent excessive inflammatory responses and endothelial damage. We investigated the therapeutic potential of IL-4 against fatal malaria in Plasmodium berghei ANKA–infected C57BL/6J mice, an experimental CM model. IL-4 treatment significantly reduced parasitemia, CM pathology, and mortality. The therapeutic effect of IL-4 is mediated through multiple mechanisms, including enhanced parasite clearance mediated by upregulation of phagocytic receptors and increased IgM production, and decreased brain inflammatory responses, including reduced chemokine (CXCL10) production, reduced chemokine receptor (CXCR3) and adhesion molecule (LFA-1) expression by T cells, and downregulation of cytotoxic T cell lytic potential. IL-4 treatment markedly reduced the infiltration of CD8⁺ T cells and brain pathology. STAT6, PI3K–Akt–NF-κB, and Src signaling mediated the cellular and molecular events that contributed to the IL-4–dependent decrease in parasitemia. Overall, our results provide mechanistic insights into how IL-4 treatment mitigates experimental CM and have implications in developing treatment strategies for organ-specific fatal malaria. Key Points Exogenous IL-4 reduces parasitemia and prevents experimental cerebral malaria. IL-4 increases phagocytic receptor expression, B1 cell expansion, and IgM production. IL-4 reduces cytotoxicity and infiltration of CD8⁺ T cells in cerebral malaria model.

Merozoite surface protein-1 (MSP-1) of malaria parasites has been extensively studied as a malaria vaccine candidate and the antibody response to this protein is an important indicator of protective immunity to malaria. Mangaluru city and its surrounding areas in southwestern India are endemic to malaria with Plasmodium vivax being the most widespread and prevalent species although P. falciparum also frequently infects. However, no information is available on the level of protective immunity in this population. In this regard, a prospective hospital-based study was performed in malarial patients to assess antibody responses against the 19-kDa C-terminal portion of P. vivax and P. falciparum MSP-1 (MSP-1₁₉). Serum samples from 51 healthy endemic controls and 267 infected individuals were collected and anti-MSP-1₁₉ antibody levels were analyzed by ELISA. The possible association between the antibody responses and morbidity parameters such as malarial anemia and thrombocytopenia was investigated. Among the 267 infected cases, 144 had P. vivax and 123 had P. falciparum infections. Significant levels of anti-MSP-1₁₉ antibody were observed both in P. vivax (123/144; 85.4%) and P. falciparum (108/123; 87.9%) infected individuals. In both type of infections, the major antibody isotypes were IgG1 and IgG3. The IgG levels were found to be increased in patients with severe anemia and thrombocytopenia. The antibody levels were also higher in infected individuals who had several previous infections, although antibodies produced during previous infections were short lived. The predominance of cytophilic anti-MSP-1₁₉ IgG1 and IgG3 antibodies suggests the possibility of a dual role of Pv MSP-1₁₉ and Pf MSP-1₁₉ during malarial immunity and pathogenesis.

Significance: Malaria kills ∼0.4 million people a year. Malaria parasite infection triggers complex immune responses that may control the infection or result in severe disease. The molecular mechanisms underlying host–parasite interaction and immune regulation are not completely understood. We perform a genome-wide genetic screen during earlyPlasmodium yoeliiinfection in mice and identify a large number of host and parasite genes/loci for future studies of the complex host–parasite interactions. We next investigate the functions of one of these host genes that encodes an E3 ubiquitin ligase (MARCH1). We show that MARCH1 regulates type I interferon signaling, T cell activation, and IFN-γ production during malaria infections. MARCH1 is a key molecule in immune responses and a potential target for immunotherapies.

VAR2CSA is the placental-malaria specific member of the antigenically variant Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) family. It is expressed on the surface of Plasmodium falciparum infected host red blood cells and binds to specific chondroitin-4-sulfate (CSA) chains of the placental proteoglycan receptor. The functional ~310 kDa ectodomain of VAR2CSA is a multi-domain protein that requires a minimum 12-mer CSA molecule for specific, high affinity receptor binding. However, how these domains interact to create the receptor binding surface is not known, limiting efforts to exploit its potential as an effective vaccine or drug target. Using small angle X-ray scattering and single particle reconstruction from negative stained electron micrographs of the ectodomain and multidomain constructs, we have determined the structural architecture of VAR2CSA. The relative location of the domains creates two distinct pores that can each accommodate the 12-mer of CSA, suggesting a model for receptor binding. This model has important implications for understanding cytoadherence of IRBCs and potentially provides a starting point for developing novel strategies to prevent and/or treat placental malaria.