The Matrix-M™ adjuvant: A critical component of vaccines for the 21st century.

Matrix-M™ adjuvant is a key component of several novel vaccine candidates. The Matrix-M adjuvant consists of two distinct fractions of saponins purified from the Quillaja saponaria Molina tree, combined with cholesterol and phospholipids to form 40-nm open cage-like nanoparticles, achieving potent adjuvanticity with a favorable safety profile. Matrix-M induces early activation of innate immune cells at the injection site and in the draining lymph nodes. This translates into improved magnitude and quality of the antibody response to the antigen, broadened epitope recognition, and the induction of a Th1-dominant immune response. Matrix-M-adjuvanted vaccines have a favorable safety profile and are well tolerated in clinical trials. In this review, we discuss the latest findings on the mechanisms of action, efficacy, and safety of Matrix-M adjuvant and other saponin-based adjuvants, with a focus on the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) subunit vaccine candidate NVX-CoV2373 developed to prevent coronavirus disease 2019 (COVID-19).

Innate Immunity Induces the Accumulation of Lung Mast Cells During Influenza Infection.

Mast cells release disease-causing mediators and accumulate in the lung of asthmatics. The most common cause of exacerbations of asthma is respiratory virus infections such as influenza. Recently, we demonstrated that influenza infection in mice triggers the recruitment of mast cell progenitors to the lung. This process starts early after infection and leads to the accumulation of mast cells. Previous studies showed that an adaptive immune response was required to trigger the recruitment of mast cell progenitors to the lung in a mouse model of allergic lung inflammation. Therefore, we set out to determine whether an adaptive immune response against the virus is needed to cause the influenza-induced recruitment of mast cell progenitors to the lung. We found that influenza-induced recruitment of mast cell progenitors to the lung was intact in Rag2-/- mice and mice depleted of CD4+ cells, implicating the involvement of innate immune signals in this process. Seven weeks after the primary infection, the influenza-exposed mice harbored more lung mast cells than unexposed mice. As innate immunity was implicated in stimulating the recruitment process, several compounds known to trigger innate immune responses were administrated intranasally to test their ability to cause an increase in lung mast cell progenitors. Poly I:C, a synthetic analog of viral dsRNA, induced a TLR3-dependent increase in lung mast cell progenitors. In addition, IL-33 induced an ST2-dependent increase in lung mast cell progenitors. In contrast, the influenza-induced recruitment of mast cell progenitors to the lung occurred independently of either TLR3 or ST2, as demonstrated using Tlr3-/- or Il1rl1-/- mice. Furthermore, neutralization of IL-33 in Tlr3-/- mice could not abrogate the influenza-induced influx of mast cell progenitors to the lung. These results suggest that other innate receptor(s) contribute to mount the influx of mast cell progenitors to the lung upon influenza infection. Our study establishes that mast cell progenitors can be rapidly recruited to the lung by innate immune signals. This indicates that during life various innate stimuli of the respiratory tract trigger increases in the mast cell population within the lung. The expanded mast cell population may contribute to the exacerbations of symptoms which occurs when asthmatics are exposed to respiratory infections.

SLC10A4 regulates IgE-mediated mast cell degranulation in vitro and mast cell-mediated reactions in vivo.

Mast cells act as sensors in innate immunity and as effector cells in adaptive immune reactions. Here we demonstrate that SLC10A4, also referred to as the vesicular aminergic-associated transporter, VAAT, modifies mast cell degranulation. Strikingly, Slc10a4 -/- bone marrow-derived mast cells (BMMCs) had a significant reduction in the release of granule-associated mediators in response to IgE/antigen-mediated activation, whereas the in vitro development of mast cells, the storage of the granule-associated enzyme mouse mast cell protease 6 (mMCP-6), and the release of prostaglandin D2 and IL-6 were normal. Slc10a4-deficient mice had a strongly reduced passive cutaneous anaphylaxis reaction and a less intense itching behaviour in response to the mast cell degranulator 48/80. Live imaging of the IgE/antigen-mediated activation showed decreased degranulation and that ATP was retained to a higher degree in mast cell granules lacking SLC10A4. Furthermore, ATP was reduced by two thirds in Slc10a4 -/- BMMCs supernatants in response to IgE/antigen. We speculate that SLC10A4 affects the amount of granule-associated ATP upon IgE/antigen-induced mast cell activation, which affect the release of granule-associated mast cell mediators. In summary, SLC10A4 acts as a regulator of degranulation in vitro and of mast cell-related reactions in vivo.

Influenza Infection in Mice Induces Accumulation of Lung Mast Cells through the Recruitment and Maturation of Mast Cell Progenitors.

Mast cells (MCs) are powerful immune cells that mature in the peripheral tissues from bone marrow (BM)-derived mast cell progenitors (MCp). Accumulation of MCs in lung compartments where they are normally absent is thought to enhance symptoms in asthma. The enrichment of lung MCs is also observed in mice subjected to models of allergic airway inflammation. However, whether other types of lung inflammation trigger increased number of MCp, which give rise to MCs, is unknown. Here, mouse-adapted H1N1 influenza A was used as a model of respiratory virus infection. Intranasal administration of the virus induced expression of VCAM-1 on the lung vascular endothelium and an extensive increase in integrin ß7hi lung MCp. Experiments were performed to distinguish whether the influenza-induced increase in the number of lung MCp was triggered mainly by recruitment or in situ cell proliferation. A similar proportion of lung MCp from influenza-infected and PBS control mice were found to be in a proliferative state. Furthermore, BM chimeric mice were used in which the possibility of influenza-induced in situ cell proliferation of host MCp was prevented. Influenza infection in the chimeric mice induced a similar number of lung MCp as in normal mice. These experiments demonstrated that recruitment of MCp to the lung is the major mechanism behind the influenza-induced increase in lung MCp. Fifteen days post-infection, the influenza infection had elicited an immature MC population expressing intermediate levels of integrin ß7, which was absent in controls. At the same time point, an increased number of toluidine blue+ MCs was detected in the upper central airways. When the inflammation was resolved, the MCs that accumulated in the lung upon influenza infection were gradually lost. In summary, our study reveals that influenza infection induces a transient accumulation of lung MCs through the recruitment and maturation of MCp. We speculate that temporary augmented numbers of lung MCs are a cause behind virus-induced exacerbations of MC-related lung diseases such as asthma.

Mefloquine, an anti-malaria agent, causes reactive oxygen species-dependent cell death in mast cells via a secretory granule-mediated pathway.

Mast cells are known to have a detrimental impact on a variety of pathological conditions. There is therefore an urgent need of developing strategies that limit their harmful effects. The aim of this study was to accomplish this by developing a means of inducing mast cell apoptosis. The strategy was to identify novel compounds that induce mast cell apoptosis by permeabilization of their secretory lysosomes (granules). As a candidate, we assessed mefloquine, an anti-malarial drug that has been proposed to have lysosome-permeabilizing activity. Mefloquine was added to mast cells and administered in vivo, followed by assessment of the extent and mechanisms of mast cell death. Mefloquine was cytotoxic to murine and human mast cells. Mefloquine induced apoptotic cell death of wild-type mast cells whereas cells lacking the granule compounds serglycin proteoglycan or tryptase were shown to undergo necrotic cell death, the latter finding indicating a role of the mast cell granules in mefloquine-induced cell death. In support of this, mefloquine was shown to cause compromised granule integrity and to induce leakage of granule components into the cytosol. Mefloquine-induced cell death was refractory to caspase inhibitors but was completely abrogated by reactive oxygen species inhibition. These findings identify mefloquine as a novel anti-mast cell agent, which induces mast cell death through a granule-mediated pathway. Mefloquine may thus become useful in therapy aiming at limiting harmful effects of mast cells.

Genomic analysis and mRNA expression of equine type I interferon genes.

This study aimed at identifying all of the type I interferon (IFN) genes of the horse and at monitoring their expression in equine cells on in vitro induction. We identified 32 putative type I IFN loci on horse chromosome 23 and an unplaced genomic scaffold. A phylogentic analysis characterized these into 8 different type I IFN classes, that is, putative functional genes for 6 IFN-α, 4 IFN-ß, 8 IFN-ω (plus 4 pseudogenes), 3 IFN-δ (plus 1 pseudogene), 1 IFN-κ and 1 IFN-ε, plus 1 IFN-ν pseudogene, and 3 loci belonging to what has previously been called IFN-αω. Our analyses indicate that the IFN-αω genes are quite distinct from both IFN-α and IFN-ω, and we refer to this type I IFN as IFN-µ. Results from cell cultures showed that leukocytes readily expressed IFN-α, IFN-ß, IFN-δ, IFN-µ, and IFN-ω mRNA on induction with, for example, live virus; while fibroblasts only expressed IFN-ß mRNA on stimulation. IFN-κ or IFN-ε expression was not consistently induced in these cell cultures. Thus, the equine type I IFN family comprised 8 classes, 7 of which had putative functional genes, and mRNA expression of 5 was induced in vitro. Moreover, a relatively low number of IFN-α subtypes was found in the horse compared with other eutherian mammals.

Phylogenetic analysis of bovine respiratory syncytial viruses from recent outbreaks in feedlot and dairy cattle herds.

Bovine respiratory syncytial virus (BRSV) is one of the major causes of bovine respiratory disease worldwide. In order to study the molecular epidemiology of the virus, samples from 30 BRSV outbreaks in cattle herds located in different parts of Sweden were collected from 2007 to 2011. The samples were analyzed by PCR, and the glycoprotein (G) gene was sequenced. BRSV was detected in outbreaks of respiratory disease in both dairy and feedlot herds most often during the winter period but also during the summer months (May to August). This indicates that circulation of the virus between herds occurs throughout the year. Comparative sequence analysis revealed a high degree (more than 94.5%) of sequence identity among the collected strains. Phylogenetic analysis showed that 29 out of the 30 strains formed a unique clade. Identical sequences found in herds sampled within a few months' time suggested that these herds were part of a common transmission chain. One strain from a single outbreak in a herd in southern Sweden clustered with Danish strains and showed a distant relationship to the rest of the Swedish strains. Further studies are highly warranted to clarify the inter-herd transmission routes of BRSV. Such knowledge is essential for the control of the spread of this virus between herds, regions and even countries.