Simian immunodeficiency virus confounds T follicular helper T cells and the germinal centre

In this study, Yamamoto and colleagues (1) examine the association of T follicular helper (T_FH) cell characteristics with the production of broadly neutralising antibodies (bNAbs) in rhesus macaques (RM) infected with simian immunodeficiency virus (SIV) AD8 as a model of human immunodeficiency virus (HIV) infection. T_FH cells are specialised CD4+ T cells essential to the germinal centre (GC) reaction, promoting affinity maturation and class switching of their cognate GC B cell receptors (2). This process is essential to the generation of long-lived high-quality antibody responses which form the basis of vaccination. The HIV-1 envelope glycoprotein spike that mediates viral entry is the primary epitope of naturally occurring neutralising antibodies against the virus and therefore a central target for HIV vaccine studies (3). However, genes encoding the HIV-1 envelope glycoproteins are also highly mutagenic with subsequent amino acid substitutions, insertions/deletions, and glycan shifting all occurring in the most easily accessible regions (4). The consequence of this is that development of bNAbs in HIV infection typically occurs late in chronic infection.

Yamamoto et al. infected eight RM with SIV-AD8 and monitored them for up to 120 weeks with periodic peripheral blood lymphocyte, lymph node and plasma sampling. The total strength and diversity of anti-HIV antibody responses from each RM were summarily scored based on the 50% inhibitory dilution titres of their plasma against 19 simian-human immunodeficiency virus (SHIV) and HIV clade viral isolates. The authors reproduce the observation that higher SIV set point viral load correlate with eventual CD4+ T cell depletion (5), late accumulation of T_FH and development of neutralising antibodies (6). T_FH accumulation was observed from 20 weeks post-infection, it is not clear if this represents a relative or absolute increase as identified previously (7). However, it did not appear to have a temporal relationship with generation of neutralising antibody which peaked at around 40 weeks, suggesting elevated T_FH numbers were not necessarily responsible for eventual antibody responses. At 44 weeks after infection Env-specific T_FH, defined as T_FH producing CD154 (CD40L), IL-4 or IFNγ in response to stimulation with HIV Env protein, were found in greater frequency in RM with higher scoring bNAbs. In particular the Env-responsive T_FH expressing IL-4 and CD154 correlated with both the number of IgG+ Env-specific GC B cells and stronger antibody responses. These findings are perhaps unsurprising as IL-4 and CD154 are crucial to GC interactions and their absence profoundly impairs GC and antibody responses (8,9), whereas the contribution of IFNγ is less clear. Whilst excess IFNγ can enhance GC responses and contribute to autoimmunity, its deficiency does not substantially impair the GC responses (10) and it is also unclear whether IFNγ has a similar function in primates as reported in mice.

The authors examined the transcriptional profile of these Env-responsive T_FH. Although absence of conventional normalisation makes interpretation difficult, several intriguing observations were made. Whilst conventional T_FH genes such as BCL6, MYB and IL-21 were expressed as expected, surprisingly the transcriptional repressor PRDM1 was found at equivalent levels between Env-responsive CD4+CD44+CXCR5+PD-1+ T_FH and non-T_FH CD4+ cells. BCL6, as the master transcriptional regulator essential to T_FH differentiation, is crucial to both the differentiation and promotion of many important T_FH functions (11). BCL6 and PRDM1 are mutually repressive in CD4+ T cells for the purpose of directing Th1/Th2 versus T_FH CD4+ effector lineage commitment (12). Thus it is perplexing that PRDM1 expression was equivalent between non-T_FH and T_FH effector CD4+ T cells; it would be expected that T_FH have BCL6-mediated repression of PRDM1. Curiously, the transcription factors TBET (T_H1), GATA3 (T_H2), and FOXP3 (T_REG) were also observed at equivalent levels between the non-T_FH and T_FH effector CD4+ T cells. Although T_FH may potentially arise from other CD4+ effectors, non-BCL6 transcriptional regulators such as TBET are normally only found in subsets of T_FH and at lower levels compared with non-T_FH effector CD4+ populations (13). Given abnormally high levels of transcriptional regulators and cytokines typically associated with other CD4+ subsets within the Env-responsive T_FH raises the possibility of either aberrant CXCR5/PD1 expression amongst effector CD4 sets, contamination of T_FH sorted pools, or issues with gene expression data.

Finally, although sequencing of single-cell sorted Env-specific IgG+ B cells heavy chain variable genes (VH4) correlated the degree of mutation with the calibre of late antibody response, the overall rate of mutation was very low suggesting hypofunctional GC responses. Unfortunately, it was not clear whether infected RM developed the hypergammaglobulinemia associated with HIV and SIV infection and which accompanies T_FH abnormalities. Thus it is uncertain whether the increase in bNAbs represented a smaller pool containing more higher-quality antibody, or simply a larger pool of antibody with bNAbs representing overwhelming hypergammaglobulinemia, a sign of the quality of the GC and T_FH response.

This study reproduces several unusual aspects of the GC biology in HIV/SIV infection. Despite declines in CD4+ T cells, phenotypically bona fide T_FH (ICOS+Bcl6+CD40L+PD1+) are known to accumulate in SIV infected RM (14). These T_FH are correlated with increased GC B and plasma cells and a reduction in memory B cells as observed by Yamamoto et al. This data supports the observation that it is not total numbers of T_FH, but rather the quality which may impair their function. It remains elusive as to why T_FH do not function normally in HIV infection. The GC is an established reservoir for HIV, particularly follicular dendritic cells (15), and T_FH are known to be infected and form sites of viral replication (14,16). Due to PD1-PDL1 interactions between T_FH and GC B cells repressing IL-21, T_FH in HIV-infected individuals may be unable to induce normal antigen-specific immune responses (17). This impairment in T_FH function is also observed with impaired influenza vaccination responses in HIV infected patients (18). The expression of PRDM1 and other transcription regulators in T_FH in this study by Yamamoto et al. suggests abnormalities in T_FH function, highlighting the need to better understand the effect of HIV/SIV infection on T_FH biology. bNAbs may represent high-quality antibody production arising from T_FH cell with typical transcriptional responses and GC characteristics, and optimising T_FH response may represent novel target for HIV vaccine development.

Acknowledgements

SH Jiang has received support from NHMRC, RACP and Jacquot Foundation grants.

Footnote

Conflicts of Interest: The authors have no conflicts of interest to declare.

References

1. Yamamoto T, Lynch RM, Gautam R, et al. Quality and quantity of TFH cells are critical for broad antibody development in SHIVAD8 infection. Sci Transl Med 2015;7:298ra120. [Crossref] [PubMed]
2. Vinuesa CG, Tangye SG, Moser B, et al. Follicular B helper T cells in antibody responses and autoimmunity. Nat Rev Immunol 2005;5:853-65. [Crossref] [PubMed]
3. Sattentau QJ. Envelope Glycoprotein Trimers as HIV-1 Vaccine Immunogens. Vaccines (Basel) 2013;1:497-512. [Crossref] [PubMed]
4. Wei X, Decker JM, Wang S, et al. Antibody neutralization and escape by HIV-1. Nature 2003;422:307-12. [Crossref] [PubMed]
5. Deeks SG, Kitchen CM, Liu L, et al. Immune activation set point during early HIV infection predicts subsequent CD4+ T-cell changes independent of viral load. Blood 2004;104:942-7. [Crossref] [PubMed]
6. Lindqvist M, van Lunzen J, Soghoian DZ, et al. Expansion of HIV-specific T follicular helper cells in chronic HIV infection. J Clin Invest 2012;122:3271-80. [Crossref] [PubMed]
7. Hong JJ, Amancha PK, Rogers K, et al. Spatial alterations between CD4(+) T follicular helper, B, and CD8(+) T cells during simian immunodeficiency virus infection: T/B cell homeostasis, activation, and potential mechanism for viral escape. J Immunol 2012;188:3247-56. [Crossref] [PubMed]
8. Kawabe T, Naka T, Yoshida K, et al. The immune responses in CD40-deficient mice: impaired immunoglobulin class switching and germinal center formation. Immunity 1994;1:167-78. [Crossref] [PubMed]
9. Yusuf I, Kageyama R, Monticelli L, et al. Germinal center T follicular helper cell IL-4 production is dependent on signaling lymphocytic activation molecule receptor (CD150). J Immunol 2010;185:190-202. [Crossref] [PubMed]
10. Lee SK, Silva DG, Martin JL, et al. Interferon-γ excess leads to pathogenic accumulation of follicular helper T cells and germinal centers. Immunity 2012;37:880-92. [Crossref] [PubMed]
11. Nurieva RI, Chung Y, Martinez GJ, et al. Bcl6 mediates the development of T follicular helper cells. Science 2009;325:1001-5. [Crossref] [PubMed]
12. Johnston RJ, Poholek AC, DiToro D, et al. Bcl6 and Blimp-1 are reciprocal and antagonistic regulators of T follicular helper cell differentiation. Science 2009;325:1006-10. [Crossref] [PubMed]
13. Schmitt N, Liu Y, Bentebibel SE, et al. The cytokine TGF-β co-opts signaling via STAT3-STAT4 to promote the differentiation of human TFH cells. Nat Immunol 2014;15:856-65. [Crossref] [PubMed]
14. Petrovas C, Yamamoto T, Gerner MY, et al. CD4 T follicular helper cell dynamics during SIV infection. J Clin Invest 2012;122:3281-94. [Crossref] [PubMed]
15. Ringler DJ, Wyand MS, Walsh DG, et al. Cellular localization of simian immunodeficiency virus in lymphoid tissues. I. Immunohistochemistry and electron microscopy. Am J Pathol 1989;134:373-83. [PubMed]
16. Perreau M, Savoye AL, De Crignis E, et al. Follicular helper T cells serve as the major CD4 T cell compartment for HIV-1 infection, replication, and production. J Exp Med 2013;210:143-56. [Crossref] [PubMed]
17. Cubas RA, Mudd JC, Savoye AL, et al. Inadequate T follicular cell help impairs B cell immunity during HIV infection. Nat Med 2013;19:494-9. [Crossref] [PubMed]
18. Pallikkuth S, Pilakka Kanthikeel S, Silva SY, et al. Upregulation of IL-21 receptor on B cells and IL-21 secretion distinguishes novel 2009 H1N1 vaccine responders from nonresponders among HIV-infected persons on combination antiretroviral therapy. J Immunol 2011;186:6173-81. [Crossref] [PubMed]