Molecular Signature of Monocytes Shaped by the Shigella sonnei 1790-Generalized Modules for Membrane Antigens Vaccine.

Shigellosis, an acute gastroenteritis infection caused by Shigella species, remains a public health burden in developing countries. Recently, many outbreaks due to Shigella sonnei multidrug-resistant strains have been reported in high-income countries, and the lack of an effective vaccine represents a major hurdle to counteract this bacterial pathogen. Vaccine candidates against Shigella sonnei are under clinical development, including a Generalized Modules for Membrane Antigens (GMMA)-based vaccine. The mechanisms by which GMMA-based vaccines interact and activate human immune cells remain elusive. Our previous study provided the first evidence that both adaptive and innate immune cells are targeted and functionally shaped by the GMMA-based vaccine. Here, flow cytometry and confocal microscopy analysis allowed us to identify monocytes as the main target population interacting with the S. sonnei 1790-GMMA vaccine on human peripheral blood. In addition, transcriptomic analysis of this cell population revealed a molecular signature induced by 1790-GMMA mostly correlated with the inflammatory response and cytokine-induced processes. This also impacts the expression of genes associated with macrophages' differentiation and T cell regulation, suggesting a dual function for this vaccine platform both as an antigen carrier and as a regulator of immune cell activation and differentiation.

Straight to the point: targeted mRNA-delivery to immune cells for improved vaccine design.

With the deepening of our understanding of adaptive immunity at the cellular and molecular level, targeting antigens directly to immune cells has proven to be a successful strategy to develop innovative and potent vaccines. Indeed, it offers the potential to increase vaccine potency and/or modulate immune response quality while reducing off-target effects. With mRNA-vaccines establishing themselves as a versatile technology for future applications, in the last years several approaches have been explored to target nanoparticles-enabled mRNA-delivery systems to immune cells, with a focus on dendritic cells. Dendritic cells (DCs) are the most potent antigen presenting cells and key mediators of B- and T-cell immunity, and therefore considered as an ideal target for cell-specific antigen delivery. Indeed, improved potency of DC-targeted vaccines has been proved in vitro and in vivo. This review discusses the potential specific targets for immune system-directed mRNA delivery, as well as the different targeting ligand classes and delivery systems used for this purpose.

Dissecting in Vitro the Activation of Human Immune Response Induced by Shigella sonnei GMMA.

Generalized Modules for Membrane Antigens (GMMA) are outer membrane exosomes purified from Gram-negative bacteria genetically mutated to increase blebbing and reduce risk of reactogenicity. This is commonly achieved through modification of the lipid A portion of lipopolysaccharide. GMMA faithfully resemble the bacterial outer membrane surface, and therefore represent a powerful and flexible platform for vaccine development. Although GMMA-based vaccines have been demonstrated to induce a strong and functional antibody response in animals and humans maintaining an acceptable reactogenicity profile, the overall impact on immune cells and their mode of action are still poorly understood. To characterize the GMMA-induced immune response, we stimulated human peripheral blood mononuclear cells (hPBMCs) with GMMA from Shigella sonnei. We studied GMMA both with wild-type hexa-acylated lipid A and with the corresponding less reactogenic penta-acylated form. Using multicolor flow cytometry, we assessed the activation of immune cell subsets and we profiled intracellular cytokine production after GMMA stimulation. Moreover, we measured the secretion of thirty cytokines/chemokines in the cell culture supernatants. Our data indicated activation of monocytes, dendritic, NK, B, and γδ T cells. Comparison of the cytokine responses showed that, although the two GMMA have qualitatively similar profiles, GMMA with modified penta-acylated lipid A induced a lower production of pro-inflammatory cytokines/chemokines compared to GMMA with wild-type lipid A. Intracellular cytokine staining indicated monocytes and dendritic cells as the main source of the cytokines produced. Overall, these data provide new insights into the activation of key immune cells potentially targeted by GMMA-based vaccines.

Staphylococcus aureus-Specific Tissue-Resident Memory CD4+ T Cells Are Abundant in Healthy Human Skin.

The skin is an immunocompetent tissue that harbors several kinds of immune cells and a plethora of commensal microbes constituting the skin microbiome. Staphylococcus aureus is a prominent skin pathogen that colonizes a large proportion of the human population. We currently have an incomplete understanding of the correlates of protection against S. aureus infection, however genetic and experimental evidence has shown that CD4+ T cells play a key role in orchestrating a protective anti-S. aureus immune response. A high S. aureus-specific memory CD4+ T cell response has been reported in the blood of healthy subjects. Since T cells are more abundant in the skin than in blood, we hypothesized that S. aureus-specific CD4+ T cells could be present in the skin of healthy individuals. Indeed, we observed proliferation of tissue-resident memory CD4+ T cells and production of IL-17A, IL-22, IFN-γ and TNF-ß by cells isolated from abdominal skin explants in response to heat-killed S. aureus. Remarkably, these cytokines were produced also during an ex vivo epicutaneous S. aureus infection of human skin explants. These findings highlight the importance of tissue-resident memory CD4+ T cells present at barrier sites such as the skin, a primary entry site for S. aureus. Further phenotypical and functional characterization of these cells will ultimately aid in the development of novel vaccine strategies against this elusive pathogen.

Dissecting the Human Response to Staphylococcus aureus Systemic Infections.

Staphylococcus aureus is a common human commensal and the leading cause of diverse infections. To identify distinctive parameters associated with infection and colonization, we compared the immune and inflammatory responses of patients with a diagnosis of invasive S. aureus disease to healthy donors. We analyzed the inflammatory responses founding a pattern of distinctive cytokines significantly higher in the patients with invasive disease. The measure of antibody levels revealed a wide antibody responsiveness from all subjects to most of the antigens, with significantly higher response for some antigens in the invasive patients compared to control. Moreover, functional antibodies against toxins distinctively associated with the invasive disease. Finally, we examined the genomic variability of isolates, showing no major differences in genetic distribution compared to a panel of representative strains. Overall, our study shows specific signatures of cytokines and functional antibodies in patients with different primary invasive diseases caused by S. aureus. These data provide insight into human responses towards invasive staphylococcal infections and are important for guiding the identification of novel preventive and therapeutic interventions against S. aureus.