Co-adjuvanting Nod2-stimulating bacteria with a TLR7 agonist elicits potent protective immunity against respiratory virus infection.

The microbiota plays a crucial role in inducing immune responses. Our previous studies have shown that symbiotic bacterial sensing by the nucleotide-binding oligomerization-domain-containing protein 2 (Nod2) receptor is involved in the mucosal adjuvanticity of cholera toxin. However, cholera toxin's potential toxicity limits human use. Here, screening other less toxic adjuvants showed that toll-like receptor (TLR) 4 and 7 agonists synergized with the microbiota in inducing adaptive immune responses upon nasal immunization. Particularly, Imiquimod, a TLR7 agonist, exhibited synergistic effects with bacterial component MDP, a Nod2 ligand, in inducing immune responses, such as IL-12p40 and IL-6 productions in bone marrow-derived dendritic cells (BMDCs) and follicular helper T (TFH) cell differentiation and high-affinity antibody production in immunized mice. The Imiquimod-MDP combination notably elicited immune protection against influenza and SARS-CoV-2 infections. Furthermore, we isolated some bacteria from the nasal cavity of healthy donors, and their Nod2-stimulating activities were measured using a reporter cell line. Staphylococcus aureus, with notable Nod2-stimulating activity, showed higher synergy with Imiquimod than Staphylococcus epidermidis, while the synergistic effects by Imiquimod-bacteria combination disappeared in Nod2-knockout mice. Moreover, the pretreatment with S. aureus enhanced the protective effect of Imiquimod-mediated vaccination against influenza virus compared to S. epidermidis. These results imply that the Imiquimod-MDP and the Imiquimod-bacteria combinations could be novel and promising complex adjuvants in developing intranasal vaccines.

Enhancing immune protection against MERS-CoV: the synergistic effect of proteolytic cleavage sites and the fusion peptide and RBD domain targeting VLP immunization.

Introduction: The Middle East Respiratory Syndrome Coronavirus (MERS-CoV) is a zoonotic infectious virus that has caused significant outbreaks in the Middle East and beyond. Due to a highly mortality rate, easy transmission, and rapid spread of the MERS-CoV, it remains as a significant public health treat. There is currently no licensed vaccine available to protect against MERS-CoV. Methods: In this study, we investigated whether the proteolytic cleavage sites and fusion peptide domain of the MERS-CoV spike (S) protein could be a vaccine target to elicit the MERS-CoV S protein-specific antibody responses and confer immune protection against MERS-CoV infection. Our results demonstrate that immunization of the proteolytic cleavage sites and the fusion peptide domain using virus-like particle (VLP) induced the MERS-CoV S protein-specific IgG antibodies with capacity to neutralize pseudotyped MERS-CoV infection in vitro. Moreover, proteolytic cleavage sites and the fusion peptide VLP immunization showed a synergistic effect on the immune protection against MERS-CoV infection elicited by immunization with VLP expressing the receptor binding domain (RBD) of the S protein. Additionally, immune evasion of MERS-CoV RBD variants from anti-RBD sera was significantly controlled by anti-proteolytic cleavage sites and the fusion peptide sera. Conclusion and discussion: Our study demonstrates the potential of VLP immunization targeting the proteolytic cleavage sites and the fusion peptide and RBD domains of the MERS-CoV S protein for the development of effective treatments and vaccines against MERS-CoV and related variants.

The transcription factor Mef2d regulates B:T synapse-dependent GC-TFH differentiation and IL-21-mediated humoral immunity.

Communication between CD4 T cells and cognate B cells is key for the former to fully mature into germinal center-T follicular helper (GC-TFH) cells and for the latter to mount a CD4 T cell-dependent humoral immune response. Although this interaction occurs in a B:T synapse-dependent manner, how CD4 T cells transcriptionally regulate B:T synapse formation remains largely unknown. Here, we report that Mef2d, an isoform of the myocyte enhancer factor 2 (Mef2) transcription factor family, is a critical regulator of this process. In CD4 T cells, Mef2d negatively regulates expression of Sh2d1a, which encodes SLAM-associated protein (SAP), a critical regulator of B:T synapses. We found that Mef2d regulates Sh2d1a expression via DNA binding-dependent transcriptional repression, inhibiting SAP-dependent B:T synapse formation and preventing antigen-specific CD4 T cells from differentiating into GC-TFH cells. Mef2d also impeded IL-21 production by CD4 T cells, an important B cell help signaling molecule, via direct repression of the Il21 gene. In contrast, CD4 T cell-specific disruption of Mef2d led to a substantial increase in GC-TFH differentiation in response to protein immunization, concurrent with enhanced SAP expression. MEF2D mRNA expression inversely correlates with human systemic lupus erythematosus (SLE) patient autoimmune parameters, including circulating TFH-like cell frequencies, autoantibodies, and SLEDAI scores. These findings highlight Mef2d as a pivotal rheostat in CD4 T cells for controlling GC formation and antibody production by B cells.

Virus-like Particle (VLP) Mediated Antigen Delivery as a Sensitization Tool of Experimental Allergy Mouse Models.

Antigen delivery systems play critical roles in determining the quality and quantity of Ab responses in vivo. Induction of protective antibodies by B cells is essential in the development of vaccines against infectious pathogens, whereas production of IgE antibodies is prerequisite for investigation of allergic responses, or type 1 hypersensitivity reactions. Virus-like particles (VLPs) are efficient platforms for expression of proteins of interest in highly repetitive manners, which grants strong Ab responses to target antigens. Here, we report that delivery of hen egg lysozyme (HEL), a model allergen, through VLP could provoke strong HEL specific IgE Ab responses in mice. Moreover, acute allergic responses were robustly induced in the mice sensitized with VLPs that express HEL, when challenged with recombinant HEL protein. Our data show that antigen delivery in the context of VLPs could function as a platform for sensitization of mice and for subsequent examination of allergic reactions to molecules of interest.