B cell depletion impairs vaccination-induced CD8+ T cell responses in a type I interferon-dependent manner

Abstract

Objectives: The monoclonal anti-CD20 antibody rituximab is frequently applied in the treatment of lymphoma as well as autoimmune diseases and confers efficient depletion of recirculating B cells. Correspondingly, B cell-depleted patients barely mount de novo antibody responses during infections or vaccinations. Therefore, efficient immune responses of B cell-depleted patients largely depend on protective T cell responses.

Methods: CD8⁺ T cell expansion was studied in rituximab-treated rheumatoid arthritis (RA) patients and B cell-deficient mice on vaccination/infection with different vaccines/pathogens.

Results: Rituximab-treated RA patients vaccinated with Influvac showed reduced expansion of influenza-specific CD8⁺ T cells when compared with healthy controls. Moreover, B cell-deficient JHT mice infected with mouse-adapted Influenza or modified vaccinia virus Ankara showed less vigorous expansion of virus-specific CD8⁺ T cells than wild type mice. Of note, JHT mice do not have an intrinsic impairment of CD8⁺ T cell expansion, since infection with vaccinia virus induced similar T cell expansion in JHT and wild type mice. Direct type I interferon receptor signalling of B cells was necessary to induce several chemokines in B cells and to support T cell help by enhancing the expression of MHC-I.

Conclusions: Depending on the stimulus, B cells can modulate CD8⁺ T cell responses. Thus, B cell depletion causes a deficiency of de novo antibody responses and affects the efficacy of cellular response including cytotoxic T cells. The choice of the appropriate vaccine to vaccinate B cell-depleted patients has to be re-evaluated in order to efficiently induce protective CD8⁺ T cell responses.

Keywords: Arthritis; B-Lymphocytes; Rheumatoid; Rituximab; T-Lymphocyte subsets; Vaccination.

Figure 1

B cell depletion affects CD8⁺ T cell response upon influenza vaccination. Healthy subjects and rituximab-treated RA patients were vaccinated against seasonal influenza. (A) Rituximab treatment efficiently depletes circulating B cells from blood. (B) Influenza-specific CD8⁺ T cells were determined after excluding CD14⁺/CD19⁺/CD56⁺ cells by using one or more personalised MHC-I multimers (left panels). B cell depletion efficiency was monitored using flow cytometry (right panel). (C) The frequency of influenza-specific T cells of CD8⁺ T cells was monitored on day 0 and 7 post vaccination. (D) Fold induction was calculated for each MHC-I multimer measurement (n=10 healthy, n=5 rituximab). Healthy subjects and one rituximab-treated GpA patient were fully vaccinated against SARS-CoV-2. (E) Serum IgG against SARS-CoV-2 S1 was determined (n=4 healthy, n=1 rituximab). Titre was considered positive when >0.8 ratio to calibrator (dotted line). error bars indicate mean±SD; **p≤0.01; one-tailed Mann-Whitney U test. FSC-A, forward scatter-area; RA, rheumatoid arthritis; SSC-A side scatter-area.

Figure 2

B cell deficient mice show reduced virus-specific CD8⁺ T cell response upon influenza and MVA, but not VACV infection. (A) Wild type (WT) and JHT mice were infected with 5×10³ ffu mouse adapted influenza virus for 7 days. (B) Influenza-specific CD8⁺ T cells were determined by using nucleoprotein (NP) or polymerase acidic protein (PAP) specific MHC-I multimers. WT and JHT mice were infected with 10⁵ pfu of (C) VACV or (D) MVA and B8-specific CD8⁺ T cells were determined by using a MHC-I multimer. Data shown are pooled from 2 to 3 experiments with n=3–4. JHT mice were reconstituted with (E) 10⁷ B cells or (F) 300 µL serum of WT mice 1 day prior to MVA infection and B8-specific T cell expansion was monitored. One out of two independent experiments is shown. Error bars indicate mean±SD; *p≤0.05, ***p≤0.001; one-tailed Mann-Whitney U test. MVA, modified vaccinia virus Ankara; ns, not significant; VACV, vaccinia virus.

Figure 3

IFNAR depletion on B cells affects B8-specific CD8⁺ T cell responses upon MVA, but not VACV infection. Wild type (WT) and CD19-Cre^+/-IFNAR^flox/flox (IFNAR-B) mice were infected with 10⁵ pfu (A) VACV or (B) MVA. B8-specific CD8⁺ T cells were determined by using an MHC-I multimer. Data shown are pooled from 3 to 4 experiments with n=3–4. Error bars indicate mean±SD; ***p≤0.001; one-tailed Mann-Whitney U test. IFNAR, type I interferon receptor; MVA, modified vaccinia virus Ankara; ns, not significant; VACV, vaccinia virus.

Figure 4

MVA-induced IFN-I responses directly trigger B cells in vivo. 10⁷ B cells isolated from Mx2-luc reporter mice were adoptively transferred into albino C57BL/6 wild type mice 1 day prior to infection. Upon treatment with phosphate buffered saline (PBS) (first mouse per row) or infection with 10⁵ pfu MVA (mouse 2–4 per row), luciferase reporter expression in adoptively transferred B cells was monitored after luciferin administration by in vivo imaging at different days (d) postinfection (scale=p/sec/cm²/sr). one out of two independent experiments is shown. IFN-I, type I interferon; MVA, modified vaccinia virus Ankara.

Figure 5

MVA-induced IFN-I responses activate B cells, but do not affect CXCR5⁺CD8⁺ T cell responses. Wild type (WT) and IFNAR-B mice were infected with 10⁵ pfu MVA and B cells were isolated 1 day post infection via untouched magnetic cell separation and prepared for mRNA sequencing. Differentially regulated (A) surface molecules and (B) chemokine as well as chemokine receptor expression profiles are shown. n=3 (C) WT and IFNAR-B mice were infected with 10⁵ pfu MVA and B8-specific CD8⁺ T cells were FACS-sorted six days post infection from spleens using a B8-specific MHC-I multimer. RNA sequencing samples were pooled from three different mice and chemokine expression profiles were analysed. IFN-I, type I interferon; IFNAR, IFN-I receptor; MVA, modified vaccinia virus Ankara.

Figure 6

MVA-induced IFN-I responses modulate antigen presentation in B cells. Wild type (WT) and IFNAR-B mice were infected with 10⁵ pfu MVA and splenocytes were isolated 48 hours post infection. Expression of (A) MHC-II, (B) MHC-I, (C) H2-kb, (D) CD86, and (E) CD69 was analysed by flow-cytometry. Data shown are pooled from 2 to 3 experiments with n=2–4. Error bars indicate mean±SD; *p≤0.01; ***p≤0.001; one-tailed Mann-Whitney U test. DPI, days post infection; IFN-I, type I interferon; IFNAR, IFN-I receptor; MVA, modified vaccinia virus Ankara; MFI, mean fluorecscence intensity; NS, not significant.