4 Discussion
In this study, we described new findings on intratracheal administration of mucosal adjuvant SF-10 derived from PS for COVID-19 vaccines. The main findings include i) S1-SF-10-IT vaccine induced S1-specific IgG in serum to levels similar to those found with S1-AS03-IM, together with induction of higher S-IgA and IgG levels in the respiratory mucosa than S1-AS03-IM, which plays an important role in protection against SARS-CoV-2 infection. ii) S1-SF-10-IT induced S1-specific IgG and IgA ASCs and S1-responsive IFN-γ, IL-4 and IL-17A CSCs in both the spleen and lungs.
Respiratory mucosal immunity plays an important front-line role against respiratory viral infections. Especially, S-IgA in the respiratory mucosa protects against invasion of viruses into mucosal cells, resulting in prevention of infection and transmission of pathogens to other individuals.12, 25 Our results demonstrated that S1-SF-10-IT vaccine induced S1-specific IgG and IgA ASCs in the spleen and lungs (Figure 3) and S1-specific antibodies in the serum and BALF (Figure 1, 2A-D), which efficiently inhibited S1/ACE2 binding (Figure 2E-H). The S1-AS03-IM induced S1-specific IgG in the serum and BALF (Figure 2A, B), but not S1-specific IgA in serum and BALF (Figure 2C, D), and S1-specific IgA ASCs in the spleen and lungs (Figure 3B, D). The antibodies-related inhibitory effects of S1/ACE2 binding in BALF induced by S1-AS03-IM were also weaker than intratracheal application of S1-SF-10-IT (Figure 2F, H).
In addition, the role of mucosal IgG induced by S1-SF-10-IT cannot be ignored; S1-SF-10-IT induced significantly higher BALF IgG levels compared with S1-AS03-IM regardless of the time of administration (Figure 2B). Based on the above results, our findings suggest that vaccination with S1-SF-10-IT provides efficient protection against respiratory infection.
In addition to protective humoral immunity, T cell-mediated immunity is important for protection of SARS-CoV-2 infection. It was reported recently that SARS-CoV-2 antigen responsive IFN-γ CSCs in blood plays an important role in the early phase of SARS-CoV-2 infection and acceleration of virus clearance.26 Also, IFN-γ producing lung tissue-resident memory T cells (TRMs) conduct immune surveillance for pathogens that could invade the tissues and robustly protect against site-specific infection by viruses, including SARS-CoV-2 in the respiratory tract, thus preventing infection.27S1-AS03-IM induced S1-responsive IFN-γ CSCs in the spleen, but not in the lungs. In contrast, S1-SF-10-IT induced S1-responsive IFN-γ CSCs in not only the spleen but also in the lungs. Additionally, S1-SF-10-IT vaccine induced S1-responsive granzyme β expression in splenic CD8+ T cells, a marker of cellular immunity (Supplementary Figure 1). Based on this likely scenario, we suggest that the use of the intratracheal route for S1-SF-10 vaccine delivery offers the advantage of preventing respiratory infection, such as SARS-CoV-2, better than intramuscularly delivered vaccine.
Although S1-SF-10-IT induced significantly higher splenic S1-responsive IL-17A CSCs relative to S1-AS03-IM, splenic S1-responsive IL-4 CSCs induced by S1-SF-10-IT were lower than those by S1-AS03-IM (Figure 4E). These results indicate that S1-SF-10-IT tends to induce mild Th2 type immunity and marked Th17 type immunity, compared with intramuscularly injected vaccine. Since IL-17A is secreted by Th17 and involved in production and migration to mucosal sites of IgA,28, 29 these results imply that IL-17A induced by S1-SF-10-IT vaccine is mediated at least in part by marked IgA induction in serum and respiratory tract mucosa. Although the functional properties of vaccine-induced Th17 TRM in the lungs remain to be examined in detail, Th17 TRM could mediate the induction of respiratory IgA following vaccination with S1-SF-10-IT.28, 29 On the other hand, others indicated that Th17 induced by SARS-CoV-2 infection can induce a cytokine storm.30, 31 Since there are two types of Th17; the first is non-pathogenic Th17, which produces IL-10, has anti-inflammatory activity, while the second is the pathogenic Th17, which does not induce IL-10,31, 32 further studies are needed to determine whether lung Th17 induced by S1-SF-10-IT immunization are non-pathogenic type capable of inducing IL-17A and IL-10 to protect against SARS-CoV-2 infection.
Our study has certain limitations. We investigated protective immunity against SARS-CoV-2 infection using by S1/ACE2 binding inhibition assayin vitro . To confirm the protective immunity induced by mucosal administration of S1-SF-10 against mortality, severity and transmission by SARS-CoV-2 infection in vivo , we need to conduct protection experiments in SARS-CoV-2-infected animals.
In conclusion, we have demonstrated in the present study that SF-10, which mimics human PS, is a suitable and effective mucosal adjuvant for COVID-19 vaccines and can be used for intratracheal delivery of vaccines to initiate rapid and potent local respiratory mucosal and systemic immunity. Intratracheal administration of COVID-19 vaccines using SF-10 are expected to provide effective protection against respiratory infection of SARS-CoV-2.