Over the last 25 years, my lab has evaluated the role that innate (dendritic cells, monocytes, microglial cells and neutrophils) and adaptive cells (B and T cells) play in immune responses to pathogens, autoantigens and allergens. In one project, we are focusing on how monocyte-derived, inflammation-induced, migratory dendritic cells initiate polarized T cell responses that contribute to pathogen clearance and tissue inflammation. In a second project, we study how NAD metabolism regulates innate cells and adaptive cells in the settings of inflammatory disease and cancer. In a third project, we study the molecular, cellular epigenetic and transcriptional signals that control B cell activation, differentiation and effector function. We specifically assess the antibody dependent and antibody independent roles for B cells in humoral and cellular immunity.
My lab was amongst the first to popularize the notion that B cells, when activated in the presence of different cytokine microenvironments, acquire distinct effector functions that are biologically important. We examine the influence of B cell-derived cytokines and chemokines on the development and function of DCs, CD4 T cells and CD8 T cells. More recently, we focused our efforts to address how cytokines and chemokines present in the microenvironment regulate B cell effector functions and their capacity to assume different differentiation cell fates –including the commitment of mature B cells to differentiate into memory B cells and antibody secreting cells (ASC) that mediate protection during vaccination and infection or damage in autoimmune and allergic diseases. Our human B cell work focuses on elucidating the molecular and functional properties of different human memory B cell subsets that are expanded in SLE patients, elicited after influenza or COVID vaccination, induced in HLA-sensitized individuals or found in different human lymphoid and non-lymphoid tissues, including the lung and gut. We also study the transcriptional and epigenetic signals controlling the development of human and mouse ASCs from naïve and memory B cells.
Recently, we have elucidated how the transcription factor T-bet regulates the molecular programming of primary and secondary humoral immune responses and how anti-viral B cell immunity is established and maintained in the lung. We identified and characterized a potentially pathogenic population of T-bet-expressing pre-ASCs that are expanded in a subset of SLE patients and developed in vitro culture conditions to generate these cells so that we can better understand the molecular signals that are required for their development, maintenance and differentiation potential. Most recently, we have returned our attention to elucidating how anti-viral B cell immunity is established and maintained in the respiratory tract. In the last year, we, with our collaborators developed all of the reagents and tools to examine SARS-CoV-2 specific memory B cell and antibody responses in acutely ill, hospitalized and convalescent COVID-19 patients as well as vaccinated individuals. These include multiple high-throughput modalities that allow rapid immune screening of large numbers of diverse human samples (PBMCs, plasma, serum, BAL, nasal/oral wash, tissues). We are currently analyzing COVID-19 vaccine responses (antibody and memory B cells), in preclinical models (mice/hamsters), in human clinical trials (particularly intranasal mucosal vaccines) and in healthy and immune-compromised individuals vaccinated with EUA COVID-19 vaccines.
In the News
A list of publications can be found on the NIH National Library of Medicine
Funding for the projects described above include
Lund, Frances E (PI)
Control of B cell differentiation by IFNg-induced transcription factors. To determine how IRF1 regulates the development of protective humoral immunity in the setting of viral infection and autoimmune disease.
R01 AI153365, NIAID
Lund, Frances E (PI)
TLR7 and TLR9 directed plasma cell formation: Dissecting the molecular basis for their differential dependence on IFN-induced signals. To determine how cooperation between TLR signals, IFNs and IRF1 support human ASC differentiation and to identify the TLR and IFN regulated ASC signaling nodes.
R01 AI150740, NIH/NIAID
Lund, Frances E (PI)
Identification and characterization of effector memory B cell populations that dominate memory responses to subsequent influenza infection and vaccination. To characterize the vaccine and infection-dependent signals required to establish, maintain and recall lung-resident T-bethi Bmem cells.
Lund, Frances E (Project Leader, Project 2)
Plasma Cells in Health and Disease – Project 2: Promoting the Development of Epigenetically and Functionally Distinct Plasma Cell Population by Modulating the Cytokine Microenvironment. To determine the molecular and epigenetic differences between ASC subsets
U19 AI142737, NIH/NIAID
Lund, Frances E. (PI) and Lund, Frances E. and Randall, Troy D (co-Project Leaders, Project 2)
Cooperative Centers on Human Immunology: Tissue and organ specific human B cell immunity. Project 2: Characterization of virus-specific human B cell subsets in lymphoid and non-lymphoid tissues. To characterize flu-specific B cells in tissues of normal human donors and to generate reagents to identify and characterize HCoV-specific B cells and broadly reactive Abs in vaccinated and infected individuals.
Sponsored Research Agreement Gates Foundation
Lund, Frances E (PI, subcontract Emory)
Understanding Long-lived Plasma Cell and Memory B cell Programs. To define the best culture conditions to induce the formation of large numbers of plasma cells that can be genetically modified.
Sponsored Research Agreement Altimmune
Lund, Frances E (PI)
Preclinical Modeling of Altimmune COVID-19 vaccine platform in mice. To provide necessary pre-clinical validation data for Altimmune Ad5-vectored COVID-19 vaccines.
Selected recent abstracts from the lab
Naïve and Memory B Cells Differentiate into Plasma Cells with Distinct Survival Outcomes
Plasma cells (PCs) are responsible for the production and secretion of protective antibodies for pathogen clearance but they also mediate pathogenic auto/alloimmune responses and produce immunoregulatory cytokines. Following B cell receptor (BCR) ligation, a subset of B cells forms short-lived PCs (SL-PCs) in extrafollicular reactions while another subset of B cells is directed to proliferate in germinal center reactions (GC). The GC response promotes the generation of memory B cells (BMem) and long-lived plasma cells (LL-PCs). The LL-PCs traffic and reside into specialized bone marrow (BM) microenvironments, or niches, where they receive signals from multiple cells for their retention and survival. Many alternative signals will give rise to distinct PCs with diverse survival, secretory and functions profiles. However, we still do not know whether different B cell subsets are endowed with pre-established programs that will commit them to differentiate into specific PC populations. To answer this question, we developed an in vitro culture method to support the activation, proliferation, differentiation and survival of PCs derived from both naïve (BN) and memory (BMem) human B cells isolated from tonsil tissue. BN and BMem cells were activated with anti-Ig, IL-2 and the TLR7/8 ligand (R848) for 3 days (Activation Step). The cells were then exposed to R848 and IL21 for 3 additional days or, alternatively, the cells were co-cultured with CD40L feeder cells and IL-21 for 9 additional days (Proliferation and Differentiation Step). In vitro generated PCs were sorted and maintained in conditioned media (Survival Step). While PCs generated from BN die within a week in culture, PCs generated from BMem survived longer. Interestingly, when IFNγ was included in the cultures during the activation step (days 0-3), PC recovery was significantly increased in the BN but not in the BMEM cultures. However, IFNγ had no effect on the long-term survival of the PCs from either culture. We conclude that BN and BMem cells, when stimulated in the same manner, give rise PCs with different survival properties. Now, we are evaluating whether PCs generated from specific memory B cell subsets isolated from different human tissues are endowed with distinct functional properties
IFN-independent IRF1 induction promotes marginal zone B cell commitment by enhancement of Notch2 pathway
Marginal zone (MZ) B cells are innate-like B cells that are positioned and poised to rapidly differentiate into antibody secreting cells (ASCs) in response to blood-borne pathogens. We show that in the absence of an interferon (IFN)-inducible transcription factor, IRF1, there is loss of the MZ B cell compartment and a significant reduction of serum antibody in response to T-independent antigens. Interestingly, IFN was dispensable for MZ B cell development.IRF1 was alternatively induced by TLR and BCR signals in transitional B cells, a MZ B cell precursor, in an IFN-independent manner. While IRF1 was not required for the positioning of B cells within the marginal zone, IRF1 enhanced the expression of ADAM10, which initiates Notch2 pathway in transitional B cells. Collectively, these data show that IFN-independent IRF1 induction promotes the development of MZ B cells by enhancing Notch2 pathway in transitional B cells.
T-bet expression marks a transcriptionally and functionally distinct population of memory B cells
Christopher A. Risley
Memory B cells (Bmem) rapidly differentiate and mount antibody (Ab) responses to previously encountered antigen (Ag). Mouse Bmem can be subdivided using BCR isotype or expression of CD73, CD80, and PD-L2. Many of these Bmem populations were discovered following vaccination with alum-adjuvanted Ag, which drives a distinct immune response from those generated after infection with IFN-inducing viruses. We previously demonstrated that the IFNγ-inducible transcription factor T-bet is required for Bmem recall responses to influenza but not nematode infections, suggesting that T-bet might regulate anti-viral Bmem. Therefore, we hypothesized that the anti-viral Bmem compartment likely contained cells that were distinct from the established subpopulations. Here, we show that T-bet expression within the flu-specific Bmem is heterogenous and not restricted to any of the previously described Bmem subsets. Furthermore, B cell-intrinsic T-bet ablation caused the loss of one of these Bmem subpopulations, and inducible deletion of T-bet in Bmem caused the loss of the same subpopulation. To determine how T-bet regulates the Bmem compartment, we performed single cell RNAseq and CITEseq on flu infection-induced Bmem. We identified numerous transcriptionally distinct clusters of Bmem, several of which express T-bet as well as genes associated with plasma cell function. Consistent with this, in vitro assays show that T-bet+ Bmem differentiate more robustly than T-betneg Bmem. These data show that the anti-viral Bmem compartment is heterogeneous and includes a T-bet expressing subpopulation that appears poised for rapid Ab responses. Thus, T-bet appears to be important for the maintenance and function of some, but not all, Bmem.
Quantifying hidden sensitization: HLA-reactive memory B cells in the spleens of sensitized women
John T. Killian
Purpose: Measurement of circulating anti-HLA antibody (anti-HLA-Ab) in transplant candidates and recipients is required for successful organ transplantation. However, serum Ab measurement alone likely underestimates the full spectrum of HLA reactivity in sensitized individuals, as memory B cells (Bmems) which target HLA specificities distinct from anti-HLA-Abs may also be generated during a sensitization event. While this “hidden” sensitization (HS) may predispose patients to an early anamnestic response posttransplant, the prevalence of HS is unknown as studies of HLA-reactive Bmems have relied on peripheral blood samples with a limited number of HLA-reactive Bmems. We aimed to improve our understanding of the prevalence and specificity of HS by analyzing Bmems in human spleens.
Methods: IgG+ Bmems were purified from the spleens of nine sensitized female organ donors. Cells, which were activated with IL-2, IL-21, and R848, secreted Abs into the supernatants (SNs) that reflected the Bmems’ specificities. Serum and SN anti-HLA-Abs were identified using a Luminex assay (OneLambda). Panel Reactive Antibody (PRA) was calculated for Class I and Class II specificities. The number of HLA specificities was compared between serum and SN for each donor. Statistical analyses were performed using Graphpad Prism. Student’s t test or Chi-square test was used to determine p values.
Results: HS was detected in 10 of 12 females, and detection markedly improved in samples with greater than 200,000 Bmems. Among samples with fewer than 200,000 activated Bmems (Group A), SNs contained an average of 17% of the serum specificities, while samples with greater than 200,000 activated Bmems (Group B) produced antibodies that contained an average of 84% of the serum specificities (p=0.02). HS was detected in 100% of Group B samples and only 33% of Group A samples (p=0.02).
Conclusions: Using serum anti-HLA-Abs alone underestimates the breadth of sensitization. By analyzing the spleen, we were able to increase the number of Bmems sampled by over 100-fold relative to peripheral blood-based studies. Compared to those studies, we detected a higher percentage of serum specificities contained in SNs. HS was also detected more frequently. This approach may improve the sensitivity for detecting HLA-reactive Bmems and for developing a more comprehensive picture of HLA-reactive B cell memory.
Transcriptional Features of TH1, TH2, and TH17 priming migratory dendritic cells – a common role for CXCR5 and Chi3l1
Miranda L Curtiss
Rationale: In a mouse infection model with the nematode Heligmosomoides polygurus (Hp), TH2 responses require CXCR5+ DC. The significance of CXCR5+ DCs in TH1 or TH17 infections is unknown. We probed the transcriptomes of migratory DCs in response to Hp (TH2), Citrobacter rodentii (Cr) (TH17), and Leishmania major (Lm) infection (BALB/c mice: TH2, C57BL/6 mice: TH1). We further parsed the transcriptomics of draining lymph node DCs to identify gene signatures associated with CXCR5+ DC.
Methods: Wildtype mice were infected with Hp larvae or Cr by gavage. Irradiated mice reconstituted with a 1:1 ratio of wildtype (CD45.1) and CXCR5 deficient (CD45.2) bone marrow were infected with either Hp larvae or intradermal Lm. Migratory dendritic cells were sorted from mesenteric lymph nodes (msLN) of Hp or Cr infected mice and popliteal lymph nodes of Lm infected mice into CD103+ (cDC1) and CD11b+ (cDC2) subsets, then performed RNA seq. In chimeric mice DC were also sorted by the congenic marker CD45.
Results: Thousands of differentially expressed genes (DEG) were found in cDC1 and cDC2 from Hp, Cr and Lm infected mice. Cxcr5 and Chi3l1 were expressed by cDC2 in Hp, Cr and Lm. In contrast, few DEG were identified between CXCR5 + and CXCR5 – cDC2 during Hp or Lm infection. The most differentially expressed gene, Chi3l1, was restricted to CXCR5+ cDC2.
Conclusions: cDC1 and cDC2 DEG differ in TH1 (B6 Lm), TH2 (Hp and BALB/c Lm) and TH17 (Cr) infection, but in all settings Cxcr5 and Chi3l1 were expressed by cDC2. CXCR5 deficient cDC2 fail to express Chi3l1.