Age-associated Changes in the Germinal Centre Response Upon Vaccination

SUMMARY Title: Age-associated Changes in the Germinal Centre Response upon Vaccination Name: Alyssa Silva Cayetano Vaccination efficacy is reduced in older individuals due to diminished immune system function with advancing age. The germinal centre (GC) response plays a crucial role in generating protective immunity upon vaccination through the production of memory B cells and long-lived antibody-secreting plasma cells. However, the GC response deteriorates with age contributing to poor humoral immunity after vaccination and therefore to increased susceptibility to infections in older individuals. I aimed to assess the age-dependent cellular changes of the GC response using a combination of in vivo and in silico tools to determine the key mechanisms driving the GC deterioration with age. After immunisation, aged (24-month old) BALB/c and C57BL/6 mice had a decrease in frequency of GC B cells and an accumulation of T follicular helper (Tfh) cells compared to adult (2-month old) mice. This indicated a reduction in the magnitude of the GC response with age. The quality of the GC was also impaired as aged mice had lower serum titres of high-affinity antigen-specific antibodies. Confocal imaging revealed that the structure of GCs is altered with age. The GC is segregated into two specialised compartments: the dark zone (DZ) where B cells proliferate and the light zone (LZ) which contains the follicular dendritic cell (FDC) network and Tfh cells that facilitate the selection of high affinity GC B cells. I found that the size of the FDC network is significantly reduced and that Tfh cells are overrepresented in the DZ of the GC in aged mice. I hypothesised a B cell intrinsic defect was responsible for the decrease in GC magnitude in aged mice. However, adoptive transfer of SWHEL B cells from 24-month old mice refuted this hypothesis. To understand which cellular factor(s) may be causing the poor GC response in aged mice I used in silico modelling of the GC. This revealed that a combination of a reduced FDC network and an aberrant localisation of Tfh cells in the DZ is responsible for the poor GC response in aged mice. This altered localisation of Tfh cells corresponded with a higher expression of CXCR4 on Tfh cells from aged mice and increased sensitivity to CXCL12. I investigated the role of CXCR4 on Tfh function by adoptive transfer of CXCR4 deficient CD4+ T cells. Upon immunisation, these cells were capable of differentiating into phenotypically intact Tfh cells and were restricted to the LZ area of the GC. This finding suggests CXCR4 expression by Tfh cells is required for DZ localisation and overexpression of CXCR4 in aged Tfh cells may have deleterious effects on the GC response by driving Tfh cell localisation towards the DZ. Together, I have shown that the age-dependent GC deterioration is a multifactorial process, likely driven by a reduction in FDC network size and aberrant Tfh cell localisation, and results in poor protective immunity after vaccination.European Union's Horizon 2020 research and innovation programme under the Marie Sklodowska-Curie grant agreement No.:67539

Regulation of the Germinal Center Response

The germinal center (GC) is a specialized microstructure that forms in secondary lymphoid tissues, producing long-lived antibody secreting plasma cells and memory B cells, which can provide protection against reinfection. Within the GC, B cells undergo somatic mutation of the genes encoding their B cell receptors which, following successful selection, can lead to the emergence of B cell clones that bind antigen with high affinity. However, this mutation process can also be dangerous, as it can create autoreactive clones that can cause autoimmunity. Because of this, regulation of GC reactions is critical to ensure high affinity antibody production and to enforce self-tolerance by avoiding emergence of autoreactive B cell clones. A productive GC response requires the collaboration of multiple cell types. The stromal cell network orchestrates GC cell dynamics by controlling antigen delivery and cell trafficking. T follicular helper (Tfh) cells provide specialized help to GC B cells through cognate T-B cell interactions while Foxp3+ T follicular regulatory (Tfr) cells are key mediators of GC regulation. However, regulation of GC responses is not a simple outcome of Tfh/Tfr balance, but also involves the contribution of other cell types to modulate the GC microenvironment and to avoid autoimmunity. Thus, the regulation of the GC is complex, and occurs at multiple levels. In this review we outline recent developments in the biology of cell subsets involved in the regulation of GC reactions, in both secondary lymphoid tissues, and Peyer's patches (PPs). We discuss the mechanisms which enable the generation of potent protective humoral immunity whilst GC-derived autoimmunity is avoided

A booster dose enhances immunogenicity of the COVID-19 vaccine candidate ChAdOx1 nCoV-19 in aged mice.

BACKGROUND: The spread of SARS-CoV-2 has caused a worldwide pandemic that has affected almost every aspect of human life. The development of an effective COVID-19 vaccine could limit the morbidity and mortality caused by infection and may enable the relaxation of social-distancing measures. Age is one of the most significant risk factors for poor health outcomes after SARS-CoV-2 infection; therefore, it is desirable that any new vaccine candidates elicit a robust immune response in older adults. METHODS: Here, we use in-depth immunophenotyping to characterize the innate and adaptive immune response induced upon intramuscular administration of the adenoviral vectored ChAdOx1 nCoV-19 (AZD-1222) COVID-19 vaccine candidate in mice. FINDINGS: A single vaccination generates spike-specific Th1 cells, Th1-like Foxp3+ regulatory T cells, polyfunctional spike-specific CD8+ T cells. and granzyme-B-producing CD8 effectors. Spike-specific IgG and IgM are generated from both the early extrafollicular antibody response and the T follicular helper cell-supported germinal center reaction, which is associated with the production of virus-neutralizing antibodies. A single dose of this vaccine generated a similar type of immune response in aged mice but of a reduced magnitude than in younger mice. We report that a second dose enhances the immune response to this vaccine in aged mice. CONCLUSIONS: This study shows that ChAdOx1 nCoV-19 induces both cellular and humoral immunity in adult and aged mice and suggests a prime-boost strategy is a rational approach to enhance immunogenicity in older persons. FUNDING: This study was supported by BBSRC, Lister institute of Preventative Medicine, EPSRC VaxHub, and Innovate UK

Rejuvenating conventional dendritic cells and T follicular helper cell formation after vaccination.

Germinal centres (GCs) are T follicular helper cell (Tfh)-dependent structures that form in response to vaccination, producing long-lived antibody secreting plasma cells and memory B cells that protect against subsequent infection. With advancing age the GC and Tfh cell response declines, resulting in impaired humoral immunity. We sought to discover what underpins the poor Tfh cell response in ageing and whether it is possible to correct it. Here, we demonstrate that older people and aged mice have impaired Tfh cell differentiation upon vaccination. This deficit is preceded by poor activation of conventional dendritic cells type 2 (cDC2) due to reduced type 1 interferon signalling. Importantly, the Tfh and cDC2 cell response can be boosted in aged mice by treatment with a TLR7 agonist. This demonstrates that age-associated defects in the cDC2 and Tfh cell response are not irreversible and can be enhanced to improve vaccine responses in older individuals

Human Foxp3-negative follicular regulatory T cells control IgE responses

Antibody responses to most infectious and food protein antigens depend on help to B cells from specialised T follicular helper (Tfh) cells. A subset of Foxp3+ regulatory T cells (Tregs) has been described in mice, with a prominent role in repressing germinal center reactions that are critical for memory B cell formation and long-lived antibody responses. These specialised Tregs co-opt the Bcl-6-dependent Tfh differentiation pathway in order to access the B cell-rich follicles and have therefore been designated as T follicular regulatory (Tfr) cells

Spatial dysregulation of T follicular helper cells impairs vaccine responses in aging.

Acknowledgements: We thank C. Vinuesa and A. Liston for critical feedback on this paper. We thank the Babraham Institute Biological Support Unit staff, who performed in vivo treatments of our animals and took care of animal husbandry. We thank the staff of the Babraham Flow Cytometry and Imaging Facilities for their technical support. The National Institute for Health Research Cambridge Biomedical Research Center is a partnership between Cambridge University Hospitals NHS Foundation Trust and the University of Cambridge, funded by the National Institute for Health Research. We thank the National Institute for Health Research Cambridge Biomedical Research Center volunteers for their participation and thank staff for their contribution in coordinating the vaccinations and venipuncture. This study was supported by funding from the Biotechnology and Biological Sciences Research Council (grant nos. BB/W001578/1, BBS/E/B/000C0407, BBS/E/B/000C0427 to M.A.L.; grant no. BBSRC BB/N011740/1 to A.E.D; and the Campus Capability Core Grant to the Babraham Institute), the European Union’s Horizon 2020 research and innovation program ‘ENLIGHT-TEN’ under the Marie Sklodowska-Curie grant agreement no. 675395 to M.A.L., a grant from IdEx Université de Paris (grant no. ANR-18-IDEX-0001 to M.E.) and by an ANR PRC grant (grant no. ANR-17-CE14-0019 to K.B.). M.A.L. is an EMBO Young Investigator and a Lister Institute Prize Fellow. D.B. was supported by the Deutsche Forschungsgemeinschaft (DFG, German Research Foundation) under Emmy Noether Programs BA 5132/1-1 and BA 5132/1-2 (grant no. 252623821 to D.B.), SFB 1054 Project B12 (grant no. 210592381 to D.B.) and Germany’s Excellence Strategy EXC2151 (grant no. 390873048 to D.B.). P.A.R. was supported by the Human Frontier Science Program (grant no. RGP0033/2015 to P.A.R.) and a PhD fellowship granted by École Normale Supérieure de Lyon. J.L.L. is supported by a National Science Scholarship (PhD) by the Agency for Science, Technology and Research, Singapore. D.L.H. received a National Health and Medical Research Council Australia Early-Career Fellowship (grant no. APP1139911). A.R.B. received a Sir Henry Wellcome Postdoctoral Fellowship (grant no. 222793/Z/21/Z). J.P.L. was a recipient of the People Program (Marie Curie Actions) of the European Union’s Seventh Framework Program (FP7/2007-2013) under REA grant agreement no. PCOFUND-GA-2013-609102.The magnitude and quality of the germinal center (GC) response decline with age, resulting in poor vaccine-induced immunity in older individuals. A functional GC requires the co-ordination of multiple cell types across time and space, in particular across its two functionally distinct compartments: the light and dark zones. In aged mice, there is CXCR4-mediated mislocalization of T follicular helper (TFH) cells to the dark zone and a compressed network of follicular dendritic cells (FDCs) in the light zone. Here we show that TFH cell localization is critical for the quality of the antibody response and for the expansion of the FDC network upon immunization. The smaller GC and compressed FDC network in aged mice were corrected by provision of TFH cells that colocalize with FDCs using CXCR5. This demonstrates that the age-dependent defects in the GC response are reversible and shows that TFH cells support stromal cell responses to vaccines