T follicular helper 17 (Tfh17) cells are superior for immunological memory maintenance

  1. Xin Gao
  2. Kaiming Luo
  3. Diya Wang
  4. Yunbo Wei
  5. Yin Yao
  6. Jun Deng
  7. Yang Yang
  8. Qunxiong Zeng
  9. Xiaoru Dong
  10. Le Xiong
  11. Dongcheng Gong
  12. Lin Lin
  13. Kai Pohl
  14. Shaoling Liu
  15. Yu Liu
  16. Lu Liu
  17. Thi HO Nguyen
  18. Lilith F Allen
  19. Katherine Kedzierska
  20. Yanliang Jin
  21. Mei-Rong Du
  22. Wanping Chen
  23. Liangjing Lu
  24. Nan Shen
  25. Zheng Liu
  26. Ian A Cockburn
  27. Wenjing Luo
  28. Di Yu

  • Curated by eLife

    eLife logo

    eLife assessment

    The enrichment of Tfh17 cells in Tfh cell central memory compartment and the dominance of Tfh17 cell population and the Tfh17 transcriptional signature in circulating Tfh cells at the memory phase are nicely demonstrated, and may well be helpful for understanding the heterogeneity of memory Tfh cells and potentially providing clues for vaccine design. The in vitro differentiation system for mouse Tfh cells also provides a strategy for others to build upon in dissection of Tfh cell development and function.

Reviewed by

Read the full article See related articles

Listed in

Log in to save this article

Abstract

A defining feature of successful vaccination is the ability to induce long-lived antigen-specific memory cells. T follicular helper (Tfh) cells specialize in providing help to B cells in mounting protective humoral immunity in infection and after vaccination. Memory Tfh cells that retain the CXCR5 expression can confer protection through enhancing humoral response upon antigen re-exposure but how they are maintained is poorly understood. CXCR5 + memory Tfh cells in human blood are divided into Tfh1, Tfh2, and Tfh17 cells by the expression of chemokine receptors CXCR3 and CCR6 associated with Th1 and Th17, respectively. Here, we developed a new method to induce Tfh1, Tfh2, and Tfh17-like (iTfh1, iTfh2, and iTfh17) mouse cells in vitro. Although all three iTfh subsets efficiently support antibody responses in recipient mice with immediate immunization, iTfh17 cells are superior to iTfh1 and iTfh2 cells in supporting antibody response to a later immunization after extended resting in vivo to mimic memory maintenance. Notably, the counterpart human Tfh17 cells are selectively enriched in CCR7 + central memory Tfh cells with survival and proliferative advantages. Furthermore, the analysis of multiple human cohorts that received different vaccines for HBV, influenza virus, tetanus toxin or measles revealed that vaccine-specific Tfh17 cells outcompete Tfh1 or Tfh2 cells for the persistence in memory phase. Therefore, the complementary mouse and human results showing the advantage of Tfh17 cells in maintenance and memory function supports the notion that Tfh17-induced immunization might be preferable in vaccine development to confer long-term protection.

Article activity feed

  1. Version published to 10.7554/elife.82217 on eLife
  2. eLife

    Author Response

    Reviewer #2 (Public Review):

    This paper investigates the maintenance and function of memory follicular helper T (Tfh) cell subsets using in vitro approaches, murine immunization models and vaccine-challenged humans. Murine Tfh cell subsets (Tfh1, Tfh2, Tfh17) were generated using in vitro polarization (iTfh1, iTfh2, iTfh17), and then tested for support of humoral response following adoptive transfer or adoptive transfer with resting in vivo for 35 days. iTfh17 cells were statistically better than iTfh1 and iTfh2 cells in promoting GC B cell and plasma cell maturation after resting in vivo, although all 3 populations were capable of B cell help. Tfh17 cells were comparatively enriched among blood borne Tfh central memory cells in humans, and were enriched at the memory phase of vaccination with hepatitis B and influenza vaccines, compared to effector phase, suggesting the possibility they are comparatively superior in Tfh cell memory formation, with greater persistence in aged individuals.

    Significance

    The enrichment of Tfh17 cells in Tfh cell central memory compartment and the dominance of Tfh17 cell population and the Tfh17 transcriptional signature in circulating Tfh cells at the memory phase are nicely demonstrated, and may well be helpful for understanding the heterogeneity of memory Tfh cells and potentially providing clues for vaccine design. The in vitro differentiation system for mouse Tfh cells also provides a strategy for others to build upon in dissection of Tfh cell development and function.

    Points to consider

    1. Even though Tfh17 cells are more likely to persist at memory timepoints in mice and in humans, or produce more GC B cells or plasma cells following transfer, all subsets can do this. Is GC output otherwise distinguishable following transfer of the individual subsets, or is their effect (cytokine related perhaps) pre-GC with differential CSR? It is also not clear if the individual subsets populate the GC and assuming they do so, if their respective phenotypes persist when they become GC Tfh cells.

    We have conducted new experiments and showed that iTfh17 preferentially generate more GC-Tfh cells in the delay immunization (after 35 day’s resting in vivo). Furthermore, different iTfh subsets maintained polarized cytokine profiles after antigen re-exposure and prompt specific CSR as their Th1 or Th2 counterparts. Please refer to the response (2) to Essential Revisions for details.

    1. iTfh17 cells induce more GC B cells and antibodies after resting and antigen challenge (Figures 1, 2). However, it's not clear whether this effect is a consequence of comparatively enhanced iTfh17 survival during resting (as suggested by latter figures), or better expansion or differential skewing to Tfh differentiation during challenge (as suggested by Figure 1 J,K). The total number of remaining adoptively-transferred cells right before challenge and 7 days post challenge will be helpful to understand that.

    We have conducted new experiments and our results suggested that the superior immunological memory maintenance of iTfh17 cells was attributed to their better survival capacity and better maintenance of the potential to differentiate into GC-Tfh cells. Please refer to the response (2) to Essential Revisions for details.

    1. The authors tried to address whether Tfh17 cells have better ability to survive till memory phase or Tfh17 cells with memory potential are generated at higher frequency at the effector phase of vaccination (Figure 5); however, the experiment is not conclusive. The cTfh population 7 days post vaccination is a mixed population with effector Tph cells and Tfh memory precursors. The increased frequency of Th17 cells at day 28 compared to day 7 could be a consequence of superior survival ability, or Tfh memory precursors with Tfh17 signature are better generated.

    As indicated in our gating strategy and the widely accepted definition of cTfh cells - CD4+ CD45RA- CXCR5+ (line 69), we respectively disagree with the reviewer’s comment ‘The cTfh population 7 days post vaccination is a mixed population with effector Tph cells and Tfh memory precursors’. The effector Tph population is defined as PD-1hiCXCR5-CD4+ T cells (Rao DA et al. Pathologically expanded peripheral T helper cell subset drives B cells in rheumatoid arthritis, Nature 2017)

    1. Experiments to confirm expansion ability of the human subsets or their B cell helper ability were not performed.

    In our new experiments, we demonstrated that iTfh1/2/17 cells showed comparable expansion ability.

    Human cTfh1/2/17 cells’ expansion ability and B helper ability were reported previously by Morita et al. (Human blood CXCR5(+)CD4(+) T cells are counterparts of T follicular cells and contain specific subsets that differentially support antibody secretion, Immunity 2011, Figure 4C-D). Human cTfh1/2/17 cells showed comparable expansion ability when co-culturing with SEB-pulsed naive B cells, and cTfh17 cells had superior B cell helper function over cTfh1 but not cTfh2 cells in promoting the B cell expansion and plasma cell formation.

  3. eLife

    eLife assessment

    The enrichment of Tfh17 cells in Tfh cell central memory compartment and the dominance of Tfh17 cell population and the Tfh17 transcriptional signature in circulating Tfh cells at the memory phase are nicely demonstrated, and may well be helpful for understanding the heterogeneity of memory Tfh cells and potentially providing clues for vaccine design. The in vitro differentiation system for mouse Tfh cells also provides a strategy for others to build upon in dissection of Tfh cell development and function.

  4. eLife

    Reviewer #1 (Public Review):

    Xin Gao et al. have performed mouse and human studies on the role of Tfh17 cells in the maintenance and function of central memory (Tcm). The authors conclude that antigen-specific Tfh17 cells outcompete Tfh1 or Tfh2 cells for persistence in the memory phase. Overall, the manuscript is well written and addresses an important issue in the field of Tfh biology. However, further investigation is warranted to understand how the CCR6 expressing Tfhcm contributes to the recall of humoral responses.

    The strength of this manuscript is the experimental system in the mouse model, indicating that the adoptive transfer of the in vitro induced Tfh17-like cells induced higher antibody responses and more GC responses than those received Tfh1 or Tfh2 cells. Another strength is the analysis of multiple human cohorts indicating that cTfh17 cells are superior in memory maintenance for HBV, influenza virus, and tetanus toxin vaccines.

    The weakness of this manuscript is not clear enough about how the CCR6 expressing Tfhcm contributes to the humoral responses. CCR6 controls mainly the localization of T cells into the inflammatory site but not into the GC site. Therefore, I could not understand the advantage of cTfh17 cells for memory maintenance in vaccination.

  5. eLife

    Reviewer #2 (Public Review):

    This paper investigates the maintenance and function of memory follicular helper T (Tfh) cell subsets using in vitro approaches, murine immunization models and vaccine-challenged humans. Murine Tfh cell subsets (Tfh1, Tfh2, Tfh17) were generated using in vitro polarization (iTfh1, iTfh2, iTfh17), and then tested for support of humoral response following adoptive transfer or adoptive transfer with resting in vivo for 35 days. iTfh17 cells were statistically better than iTfh1 and iTfh2 cells in promoting GC B cell and plasma cell maturation after resting in vivo, although all 3 populations were capable of B cell help. Tfh17 cells were comparatively enriched among blood borne Tfh central memory cells in humans, and were enriched at the memory phase of vaccination with hepatitis B and influenza vaccines, compared to effector phase, suggesting the possibility they are comparatively superior in Tfh cell memory formation, with greater persistence in aged individuals.

    Significance
    The enrichment of Tfh17 cells in Tfh cell central memory compartment and the dominance of Tfh17 cell population and the Tfh17 transcriptional signature in circulating Tfh cells at the memory phase are nicely demonstrated, and may well be helpful for understanding the heterogeneity of memory Tfh cells and potentially providing clues for vaccine design. The in vitro differentiation system for mouse Tfh cells also provides a strategy for others to build upon in dissection of Tfh cell development and function.

    Points to consider
    1. Even though Tfh17 cells are more likely to persist at memory timepoints in mice and in humans, or produce more GC B cells or plasma cells following transfer, all subsets can do this. Is GC output otherwise distinguishable following transfer of the individual subsets, or is their effect (cytokine related perhaps) pre-GC with differential CSR? It is also not clear if the individual subsets populate the GC and assuming they do so, if their respective phenotypes persist when they become GC Tfh cells.

    2. iTfh17 cells induce more GC B cells and antibodies after resting and antigen challenge (Figures 1, 2). However, it's not clear whether this effect is a consequence of comparatively enhanced iTfh17 survival during resting (as suggested by latter figures), or better expansion or differential skewing to Tfh differentiation during challenge (as suggested by Figure 1 J,K). The total number of remaining adoptively-transferred cells right before challenge and 7 days post challenge will be helpful to understand that.

    3. The authors tried to address whether Tfh17 cells have better ability to survive till memory phase or Tfh17 cells with memory potential are generated at higher frequency at the effector phase of vaccination (Figure 5); however, the experiment is not conclusive. The cTfh population 7 days post vaccination is a mixed population with effector Tph cells and Tfh memory precursors. The increased frequency of Th17 cells at day 28 compared to day 7 could be a consequence of superior survival ability, or Tfh memory precursors with Tfh17 signature are better generated.

    4. Experiments to confirm expansion ability of the human subsets or their B cell helper ability were not performed.

  6. eLife

    Reviewer #3 (Public Review):

    In order to study memory Tfh cell subsets the authors develop an in vitro assay to generate Ovalbumin (OVA) specific Tfh1, Tfh2 and Tfh17 cells. In vitro, these subsets express the expected hallmarks of successful differentiation. These subsets are able to mostly maintain their phenotype upon adoptive transfer and reactivation (by immunisation) in vivo providing an experimental system to test their function. The transferred cells can support germinal centres and antibody production, with iTfh17 having a larger effect after a long in vivo rest period, proposed to be due to enhanced expression of CCR7.

    The authors then focus on human CXCR5+CD45RA-CD4+ cells that they call circulating Tfh-like (cTfh) cells, and divide these into CCR7+PD-1- TfhCM and PD-1+CCR7low TfhEM. RNAseq shows that there are different pathways enriched in these groups, with TfhCM having superior survival and proliferation in vitro as compared to TfhEM. The authors then further subdivide TfhCM and TfhEM into Tfh1/2/17 and show that there are differences in the ratios of these subgroups, and that the TfhEM have more pronounced effector characteristics that are typically associated with Th1/2/17 cells. In an HBV vaccination cohort, antigen specific cTfh17 cells were expanded in people who produced an early antibody response to HBV, but not in those who responded later. The authors then used a publicly available dataset of scRNAseq of HA-specific CD4+ T cells to identify an enrichment of T cells Tfh17 signature prior to vaccination and with a Tfh1 signature 12 days after vaccination, the latter finding is consistent with previous reports. Finally, the authors examine long term immunity by focusing on antigen-specific cells that likely were generated during childhood vaccination. cTfh17 cells were the most abundant cTfh subset recalled. Further these appear to accumulate with increasing age, indicating that these cells are likely retained as memory. Together, this body of work makes the case that CCR6+CXCR3-CXCR5+CD45RA-CD4+ cells (cTfh17 cells) are memory cells that are recalled upon challenge.

  7. Version published to 10.1101/2022.07.31.502219v1 on bioRxiv