Outer membrane vesicles harboring modified lipid A moiety augment the efficacy of an influenza vaccine exhibiting reduced endotoxicity in a mouse model

Abstract

Influenza is an acute respiratory disease and a major health problem worldwide. Since mucosal immunity plays a critical role in protection against influenza virus infection, mucosal immunization is considered a promising vaccination route. However, except for live-attenuated vaccines, there are no effective killed or recombinant mucosal influenza vaccines to date. Outer membrane vesicles (OMVs) are nano-sized vesicles produced by gram-negative bacteria, and contain various bacterial components capable of stimulating the immune system of the host. We generated an OMV with low endotoxicity (fmOMV) by modifying the structure of the lipid A moiety of lipopolysaccharide and investigated its effect as an intranasal vaccine adjuvant in an influenza vaccine model. In this model, fmOMV exhibited reduced toll-like receptor 4-stimulating activity and attenuated endotoxicity compared to that of native OMV. Intranasal injection of the vaccine antigen with fmOMV significantly increased systemic antibody and T cell responses, mucosal IgA levels, and the frequency of lung-resident influenza-specific T cells. In addition, the number of antigen-bearing CD103 dendritic cells in the mediastinal lymph nodes was significantly increased after fmOMV co-administration. Notably, the mice co-immunized with fmOMV showed a significantly higher protection rate against challenge with a lethal dose of homologous or heterologous influenza viruses without adverse effects. These results show the potential of fmOMV as an effective mucosal adjuvant for intranasal vaccines.

Bacterial Outer Membrane Vesicles Provide Broad-Spectrum Protection against Influenza Virus Infection via Recruitment and Activation of Macrophages

“…5b, left). Consistent with a previous report [21], the mobility, food intake, and body weight of the mice were normal, and inflammation in the lung tissue was not observed when fmOMV was intranasally injected (data not shown). These data indicate that fmOMV provide broad and potent protection against diverse influenza A viruses (IAVs) without endotoxic inflammation.…”

“…Based on the fact that OMVs contain diverse bacterial components which can stimulate TLRs [23,25,26], we previously generated a modified OMV harboring less endotoxic LPS (fmOMV, Fig. 1a) and investigated its safety and adjuvant effect using an influenza vaccine model [21]. In this study, we tested whether fmOMV initiates TLR 2, 3, 4, 5, and 9 signaling simultaneously, using HEK-Blue cells expressing individual TLRs.…”

“…Previously, we reported that OMV injection with a vaccine antigen increases vaccine-induced antibody responses using ovalbumin and influenza vaccine antigens [21,23]. Since rapid induction of protective immunity in particular is required for vaccines against highly transmissible infectious diseases such as influenza, we first tested whether fmOMV could induce protective efficacy of an influenza vaccine at early time points after the injection.…”

“…Previously, we generated fmOMV, which stands for further-modified OMV, of which lipid A is a dephosphorylated and underacylated species that resulted from both expression of the LpxF 4′-phosphatase and deletion of the msbB/pagP lipid A myristoyl/palmitoyltransferase genes. This OMV showed a significant adjuvant effect on a seasonal influenza vaccine exhibiting much attenuated TLR4-stimulating activity in vitro and endotoxicity in vivo [21,22]. Although the adjuvant effect of OMVs has been reported in various antigen models, the antiviral effects via activation of innate immunity have not yet been addressed.…”

Gram-negative bacterial membrane vesicle release in response to the host-environment: different threats, same trick?

“…So, although de-acylation itself can result in an increased membrane vesicle release, it is possible that these effects are cancelled-out by other lipid A modifications. Moreover, de-acylated lipid A (LpxM deletion mutants) or monophosphorylated penta-acylated mutants (LpxM/LpxF double deletions mutants) did not show or only showed a slightly increased vesiculation as compared to wild-type bacteria ( (Chen et al 2016;Lee et al 2016) and personal communication with Dr. SH. Kim and Prof. M. DeLisa, respectively).…”