Viral fusion proteins mediate cell entry by undergoing a series of conformational changes that result in virion-target cell membrane fusion. Class I viral fusion proteins, such as those encoded by influenza virus and human immunodeficiency virus (HIV), contain two prominent alpha helices. Peptides that mimic portions of these alpha helices inhibit structural rearrangements of the fusion proteins and prevent viral infection. The envelope glycoprotein (E) of flaviviruses, such as West Nile virus (WNV) and dengue virus (DENV), are class II viral fusion proteins comprised predominantly of beta sheets. We used a physio-chemical algorithm, the Wimley-White interfacial hydrophobicity scale (WWIHS) [1] in combination with known structural data to identify potential peptide inhibitors of WNV and DENV infectivity that target the viral E protein. Viral inhibition assays confirm that several of these peptides specifically interfere with target virus entry with 50% inhibitory concentration (IC50) in the 10 μM range. Inhibitory peptides similar in sequence to domains with a significant WWIHS scores, including domain II (IIb), and the stem domain, were detected. DN59, a peptide corresponding to the stem domain of DENV, inhibited infection by DENV (>99% inhibition of plaque formation at a concentrations of 99% inhibition at <25 μM) was also demonstrated with DN59. However, a potent WNV inhibitory peptide, WN83, which corresponds to WNV E domain IIb, did not inhibit infectivity by DENV. Additional results suggest that these inhibitory peptides are noncytotoxic and act in a sequence specific manner. The inhibitory peptides identified here can serve as lead compounds for the development of peptide drugs for flavivirus infection.

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Peptide inhibitors of dengue virus and West Nile virus infectivity

Dengue fever is an infectious tropical disease that is transmitted by mosquitoes carrying dengue virus. Almost half of the world’s population lives in dengue-plagued regions and it is estimated that between 50 and 100 million people are infected annually. However, there is no effective vaccine and researchers have only a limited understanding of the mechanisms behind the disease. While most infected individuals experience fever, headache, muscle and joint pain, and a skin rash, a small percentage go on to develop a life-threatening condition known as dengue hemorrhagic fever (DHF). This is marked by internal bleeding, and by the leakage of water and salts from blood vessels (vascular leakage). At present, it is difficult to tell which patients will develop this complication, making it hard to tailor treatment appropriately. Here, St John et al. reveal that the vascular leakage that occurs in DHF is triggered by mast cells, which line blood vessels and regulate their permeability through the release of molecules such as histamines and leukotrienes. They also found that mice deficient in mast cells did not show dengue-induced vascular leakage, and that wild-type animals treated with drugs that block the actions of proteins produced by these cells, showed less vascular leakage than controls. Moreover, levels of an enzyme called chymase, another mast cell product, are higher in human patients with DHF than in those with dengue fever. Since chymase release occurs early in infection, tests for the presence of this enzyme could be used to predict which patients are likely to develop DHF. The work of St John et al. suggests new lines of inquiry into the mechanisms that lead some individuals infected with dengue fever to develop DHF, and indicates that drugs that target mast cells could offer an effective treatment.

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Contributions of mast cells and vasoactive products, leukotrienes and chymase, to dengue virus-induced vascular leakage

Human mast cell activation with virus-associated stimuli leads to the selective chemotaxis of natural killer cells by a CXCL8-dependent mechanism.

Dengue virus (DENV) is the most significant human arboviral pathogen and causes ∼400 million infections in humans each year. In previous work, we observed that mast cells (MC) mediate vascular leakage during DENV infection in mice and that levels of MC activation are correlated with disease severity in human DENV patients (St John et al., 2013b). A major risk factor for developing severe dengue is secondary infection with a heterologous serotype. The dominant theory explaining increased severity during secondary DENV infection is that cross-reactive but non-neutralizing antibodies promote uptake of virus and allow enhanced replication. Here, we define another mechanism, dependent on FcγR-mediated enhanced degranulation responses by MCs. Antibody-dependent mast cell activation constitutes a novel mechanism to explain enhanced vascular leakage during secondary DENV infection.

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Dengue vascular leakage is augmented by mast cell degranulation mediated by immunoglobulin Fcγ receptors

Influenza A virus (IAV) is a widespread infectious agent commonly found in mammalian and avian species. In humans, IAV is a respiratory pathogen that causes seasonal infections associated with significant morbidity in young and elderly populations, and has a large economic impact. Moreover, IAV has the potential to cause both zoonotic spillover infection and global pandemics, which have significantly greater morbidity and mortality across all ages. The pathology associated with these pandemic and spillover infections appear to be the result of an excessive inflammatory response leading to severe lung damage, which likely predisposes the lungs for secondary bacterial infections. The lung is protected from pathogens by alveolar epithelial cells, endothelial cells, tissue resident alveolar macrophages, dendritic cells, and mast cells. The importance of mast cells during bacterial and parasitic infections has been extensively studied; yet, the role of these hematopoietic cells during viral infections is only beginning to emerge. Recently, it has been shown that mast cells can be directly activated in response to IAV, releasing mediators such histamine, proteases, leukotrienes, inflammatory cytokines, and antiviral chemokines, which participate in the excessive inflammatory and pathological response observed during IAV infections. In this review, we will examine the relationship between mast cells and IAV, and discuss the role of mast cells as a potential drug target during highly pathological IAV infections. Finally, we proposed an emerging role for mast cells in other viral infections associated with significant host pathology.

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Mast cells and influenza a virus: association with allergic responses and beyond.

Mice were immunized with dengue type 2 virus (DEN 2) under a schedule favoring the production of IgE antibody The antibody obtained could sensitize peritoneal resident mast cells both in vitro and in vivo so that the sensitized cells were degranulated and released histamine on challenge with the DEN 2 antigen It was also demonstrated that the antibody was cytophilic and heat-labile The above observations suggest that the present experimental system can be used to detect anti-DEN 2 IgE antibody in mice

Degranulation and Histamine Release from Murine Mast Cells Sensitized with Dengue Virus-Immune Sera

Dengue virus multiplication in cultures of a murine myelomonocytic cell line (WEHI-3) as well as mouse peritoneal macrophages was enhanced by treatment of the cells with lipophilic derivatives of muramyl peptides for 2 or 3 days before virus inoculation, but not for 2 hr before virus inoculation or during the adsorption period. The infection-enhancing activity of the materials was dependent on their chemical structure, correlating with their immunoadjuvanticity. The infection enhancement in WEHI-3 cells was due primarily to an increase in the number of virus-infected cells which was accompanied by an increased cellular capacity to bind latex particles to their cell surfaces.

Luis Fernando Sanchez

Papers

Degranulation and Histamine Release from Murine Mast Cells Sensitized with Dengue Virus-Immune Sera

Enhancement of dengue virus infection in cultured mouse macrophages by lipophilic derivatives of muramyl peptides.