Healthy Adults Do Not Have A Lot Of B Cell Immunity To SARS-CoV-2 And Its Solution

Summarize

Previous immunity to SARS-CoV-2 may have important implications for our understanding of susceptibility and severity of COVID-19. The presence and clinical relevance of preexisting B cell immunity remain to be fully elucidated. Here, we provide a detailed analysis of B-cell immunity in individuals not exposed to SARS-CoV-2. To this end, we extensively investigated SARS-CoV-2 humoral immunity in 150 adults sampled before the pandemic. Comprehensive screening of donor plasma and purified IgG samples, combined and neutralized in various functional assays, showed no substantial activity against SARS-CoV-2, but widespread reactivity against endemic β-coronavirus. In addition, antibody sequences from 8174 sars-cov-2 reactive B cells were analyzed at the single-cell

level, and 158 monoclonal antibodies were generated and tested. None of these antibodies showed binding or neutralizing activity associated with SARS-CoV-2. Taken together, our results show no evidence of prior effective antibody and B-cell immunity in adults not exposed to SARS-CoV-2.

Keywords

T-cell; B-cell; Cistanche; Verbascoside; Immunity

Introduction

The current SARS Coronavirus 2 pandemic is a global health emergency that challenges health care systems around the world. SARS-CoV-2 infection has a wide range of clinical manifestations, from asymptomatic infection to life-threatening acute respiratory distress syndrome, multiple organ failure, septic shock and death. Although influenced by a variety of factors, the severity of the disease is largely determined by innate and adaptive immune responses. Prior immunity to SARS-CoV-2 may be a key determinant of disease severity and clinical outcome. However, preexisting immunity may also adversely affect the clinical course through antibo-dependent enhancement (ADE) or OAS mechanisms that have previously been described for other viral pathogens.

A pre-existing T cell immune response against SARS-CoV-2 was observed in individuals not exposed to SARS-CoV-2. In these studies, T cell reactivity to spike (S) and nucleocapsid (N) proteins and non-structural proteins NSP7 and NSP13 was measured using antigenic peptide pools. Importantly, significant T cell reactivity to S protein polypeptides was detected, which showed a high homology with local "common cold" human coronavirus (HCoV), including HCOV-OC43, HCOV-HK-1, HCOV-NL63, and HCOV-229e. Therefore, it is hypothesized that preexisting T cell immunity against SARS-CoV-2 stems from prior exposure to endemic hcov.

To determine the presence of associated preexisting SARS-CoV-2 B cell immunity, we studied plasma samples, single B cells, and monoclonal antibodies isolated from 150 individuals who had not been exposed to SARS-CoV-2. We found no evidence of prior effective B-cell immunity that would explain the broad clinical spectrum of SARS-CoV-2 infection. It also provides impetus for the search for new solutions. Studies have shown that litenoside enhances lymphocyte proliferation, and therefore has a certain effect in increasing the number of B cells. This provides a new way to boost people's immunity.

Pic: Faw Cistanche

Results

We previously identified rare heavy chain and light chain variable regions in pre-pandemic naive B cells that were unpaired by NGS analysis in healthy individuals, which closely resembled the sars-cov-2 reactive antibody of the near-lineage. Some of these heavy and light chains from primary B cells can replace the original chains of sars-cov-2 reactive antibodies without changing their function. These findings add to previous observations that some sars-cov-2 reactive antibodies show their activity by species-coding sequence signatures. To investigate how these findings translate into plasma responses and whether sars-cov-2 reactive antibodies are already present in the plasma of unexposed individuals, we investigated pre-pandemic blood samples from 150 donors. Samples were collected between August and November 2019 to study its binding and neutralizing activity against SARS-CoV-2. The donors are 18-66 years old. 49.3 percent of donors were male and 50.7 percent were female. Given the conflicting results previously published, we decided to rigorously search for preexisting sars-cov-2 reactive antibodies using different tests simultaneously. Plasma samples showed little or no binding of IgG, IgM and IgA to soluble SARS-CoV-2 S protein. Significant reactivity was detected in a small number of samples. However, no binding was detected by commercial or internal immunoassay in the plasma samples investigated. To confirm this result, we performed flow cytometry. None of the samples showed any binding activity except one donor who showed weak plasma IgG activity in this study. We attribute the incidental detection of binding activity to the use of unpurified plasma samples, which would have produced non-specific reactivity.

Pic: Cistanche Extract

In summary, none of the pre-pandemic samples obtained from 150 adults showed reactivity to SARS-CoV-2 in the various assays applied. Therefore, based on the parallel application of various testing methods, we found no convincing evidence of sars-cov-2 reactive antibodies in blood samples tested before the pandemic. Experiments in rats showed that rats treated with cistanche extract showed higher levels of lymphocytes, which can differentiate into B and T cells, than those that did not. Therefore, it can be proved that cistanche extract has the function of enhancing human immunity.

Pic: Effects of cistanche improve immunity

Discussion

An adaptive immune response to a pathogen is formed by the original immune pool and leaves its mark on previous encounters with the same pathogen or related variants. The investigation of existing immunity to SARS-CoV-2 can advance our understanding of protective immunity, susceptibility to infection, disease severity, and guide the development of vaccination strategies. Although various studies have provided evidence of preexisting T cell immunity against SARS-CoV-2, preexisting B cell immunity remains to be elucidated.

We have recently isolated sars-cov-2 reactive antibodies from convalescent individuals and identified highly similar heavy and/or light chain sequences in the naive B cell receptor library from pre-pandemic samples. Here we show that some of these chains can become components of sars-cov-2 reactive antibodies without the need for further affinity maturation. This finding supports previous observations that antibody responses that are easily detected in some individuals may use existing antibody heavy or light chains characteristic of different CDR3 recombination patterns or germ line coding sequences. In line with this, SARSCoV-2 neutralizing antibodies were successfully isolated from naive B cell compartals using antigen-specific single B cell classification or phage display. Feldman et al. isolated single SARS-CoV-2 and Sarbecovire-reactive naive B cells by successively using different SARS-CoV-2 spike protein subdomains as antigen probes. Using this approach, they reported a median frequency of 0.0025% for rbm reactive naive B cells. BCR sequence analysis showed that heavy and light chains used a variety of polyclonal genes, but heavy chain v gene IGHV3-9 increased the average retention frequency by 20%. Finally, monoclonal antibodies were isolated from primitive B cell precursors that exhibit binding and neutralizing activity against the popular SARS-CoV-2 variant and bat-derived coronavirus-virus. Bertoglio et al. used phage display technology to isolate the candidate antibody STE73-2E9 from the original B cell bank. The antibody targeted the ACE2-RBD interface, did not cross-react with other coronavirus, and neutralized the real SARS-CoV-2 wild-type virus, with an IC50 of 0.43 nM. Both cistanche polysaccharide and creinoside can increase the enzyme activity of heart and brain tissue, enhance the phagocytosis function of abdominal cavity cells, and enhance the proliferative response of lymphocytes.

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Conclusion

To investigate potential preexisting immunity at the molecular level, we applied antigen-specific single-cell classification to 40 donors and isolated a total of 8,174 B cells assumed to be sars-cov-2 reactive. Consistent with our binding and neutralizing screening of plasma samples and polyclonal IgG, the frequency of SARS-CoV-2-reactive B cells in pre-pandemic samples was very low, especially compared to COVID-19 convalescent samples. The selection of suitable decoy proteins for single cell sorting is crucial to the isolation of antigen-specific B cells. The selected decoy protein does not show immunogenic structure and is excluded from isolation. Therefore, to ensure full isolation of antigen-specific B cells, we chose to apply native full trimer SARS-CoV-2 S protein and adjust the sorting gate. With regard to the low frequency of sars-cov-2 reactive B cells in our study, it can be argued that the application of alternative decoy proteins, such as the S2 subunit, would be advantageous. Antibody sequence analysis of isolated sars-cov-2 reactive B cells showed that the heavy chain V gene was used differently, the length of CDRH3 was normally distributed, and the phylogenetic identity of the VH gene was similar to the naive BCR receptor library of healthy individuals sampled before the pandemic. In conclusion, healthy adults do not have a large number of B cell immunity to SARS-CoV-2, but a certain period of administration of cistanche extract can improve human immune function and increase the content of B cells and T cells. To some extent, this has provided a new path for the treatment and prevention of COVID-19.

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