Biology subjects

Ludewig, B.

Publications and source records attributed to Ludewig, B..

13 recordsLinked to original sources

bioRxiv2026

The Notch signaling pathway is a master regulator of CD8+ T cell exhaustion and differentiation during chronic infection

During chronic infection, the persistence of antigen and inflammation leads to the differentiation of CD8+ T cells into an exhausted state characterised by expression of inhibitory receptors (IRs) and the progressive loss of T cell functions. Among the different subsets of exhausted CD8+ T (Tex) cells, Tex progenitors expressing SLAMF6 and the transcription factor TCF-1 (TCF-1+) give rise to Tex effector-like cells expressing CX3CR1 and Tex terminal cells expressing CD101. PD-1/PD-L1 blockade acts on TCF-1+ Tex progenitor cells and promotes their differentiation into Tex effector-like cells. The molecular events controlling CD8+ Tex cell differentiation are still poorly defined. As Notch signaling may be sustained during chronic infection by persistent TCR stimulation and inflammation, we tested whether Notch signaling influences CD8+ T cell exhaustion. Using mice lacking (N1N2{triangleup}/{triangleup}) or not (N1N2fl/fl) Notch1/2 expression only in mature CD8+ T cells, we showed that the absence of Notch signal causes severe CD8+ T cell exhaustion during chronic LCMV infection. N1N2{triangleup}/{triangleup} Tex cells express higher levels of IRs and are less functional when compared to their wild-typee counterpart. In the absence of N1N2 receptors, Tex progenitor and Tex terminal cells accumulate and Tex cells cannot be reinvigorated by PD-1/PD-L1 blockade. We further demonstrated that Notch signaling is essential to promote the differentiation of Tex progenitors into Tex effector-like cells. Moreover, Notch signals, provided by stromal cells expressing the ligands Delta-like 1 and 4, are necessary during all stages of the infection to prevent severe exhaustion. Single-nucleus RNA and ATAC multiome profiling identifies Notch signaling as a critical role on effector transcriptional programming in exhausted CD8 T cells. Loss of Notch signaling impairs transcriptional program associated with migration and perception of CD4+ T cell help. Together, these alterations drive the differentiation of Tex progenitor cells toward a terminally exhausted Tex fate.

immunology↗

bioRxiv2025

PMGen: From Peptide-MHC Prediction to Neoantigen Generation

Accurate structural modeling of peptide-MHC (pMHC) complexes is a prerequisite for understanding adaptive immunity and developing data-driven immunotherapies. However, current tools are often limited by narrow class coverage, restricted peptide lengths, or insufficient accuracy for downstream design tasks. Here, we introduce PMGen (Peptide MHC Generator), an integrated framework for structure prediction and structure-guided design of variable-length peptides across MHC class I and II. By introducing Initial Guess and Template Engineering as strategies to enforce anchor constraints in AlphaFold2, PMGen achieves state-of-the-art structural fidelity with median peptide core RMSDs of 0.54 [A] for MHC-I and 0.33 [A] for MHC-II, outperforming five state-of-the-art methods. We further demonstrate that PMGen captures the subtle structural impact of single-point neoantigen mutations and that model confidence (pLDDT) reliably correlates with structural accuracy. We investigated two potential applications of our framework: structure-aware peptide design and generating data for machine learning (ML) models. To this end, we introduced a framework to sample peptides with preserved structures and improved binding affinity. As an example for ML application, we fine-tuned ProteinMPNN on PMGen-modeled structures. This improved sequence recovery from 0.19 to 0.40 compared to the baseline. Ultimately, PMGen bridges the gap between high-fidelity structural prediction and downstream sequence design, offering a scalable solution to generate the large-scale, high-quality structural datasets required to train advanced predictive models in immunology. Available at https://github.com/soedinglab/PMGen.

bioinformatics↗

bioRxiv2024

Sustained innate interferon is an essential inducer of tertiary lymphoid structures

Tertiary lymphoid structures (TLS) resemble follicles of secondary lymphoid organs and develop in non-lymphoid tissues during inflammation and cancer. Which cell types and signals drive the development of TLS is largely unknown. To investigate early events of TLS development in the lungs, we repeatedly instilled p(I:C) plus ovalbumin (Ova) intranasally. This induced TLS ranging from lymphocytic aggregates to organized and functional structures containing germinal centers. We found that TLS development is independent of FAP+ fibroblasts, alveolar macrophages or CCL19 but crucially depends on type I (IFN-I)-but not type III interferon (IFN-III)-signaling. Mechanistically, IFN-I initiates two synergistic pathways that culminate in the development of TLS. On the one hand, IFN-I induces lymphotoxin (LT) in lymphoid cells, which stimulate stromal cells to produce the B-cell-attracting chemokine CXCL13 through LT{beta}R-signaling. On the other hand, IFN-I is sensed by stromal cells that produce the T-cell-attracting chemokines CXCL9, CXCL10 as well as CCL19 and CCL21 independently of LT{beta}R. Consequently, B-cell aggregates develop within a week, whereas follicular dendritic cells and germinal centers appear after 3 weeks. Thus, sustained production of IFN-I together with an antigen is essential for the induction of functional TLS in the lungs. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=152 SRC="FIGDIR/small/591846v1_ufig1.gif" ALT="Figure 1"> View larger version (18K): org.highwire.dtl.DTLVardef@a2d940org.highwire.dtl.DTLVardef@165294aorg.highwire.dtl.DTLVardef@79c43forg.highwire.dtl.DTLVardef@4316ce_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2024

Distinct circulating monocytes up-regulate CD52 and sustain innate immune function in patients with cirrhosis unless acute decompensation emerges

Background & AimsInfectious complications determine the prognosis of cirrhosis patients. Their infection susceptibility relates to the development of immuneparesis, a complex interplay of different immunosuppressive cells and soluble factors. Mechanisms underlying the dynamics of immuneparesis of innate immunity remain inconclusive. We aimed to dissect the heterogeneity of circulating monocyte states in different cirrhosis stages, and pursued the function of selected differentially expressed (DE) genes. MethodsWe systematically investigated circulating monocytes in health, compensated and not-acutely decompensated (NAD) cirrhosis using single cell RNA sequencing. Selective genes were confirmed by flow cytometry and diverse functional assays on monocytes ex vivo. ResultsWe identified seven monocyte clusters. Their abundances varied between cirrhosis stages, confirming previously reported changes i.e. reduction in CD14lowCD16++ and emergence of M-MDSC in advanced stages. DE genes between health and disease and among stages were detected, including for the first time CD52. CD52-expression on monocytes significantly enhanced throughout compensated and NAD cirrhosis. Heretofore the biological significance of CD52-expression on monocytes remained unknown. CD52highCD14+CD16highHLA-DRhigh monocytes in patients with cirrhosis revealed a functional phenotype of active phagocytes with enhanced migratory potential, increased cytokine production, but poor T cell activation. Following acute decompensation (AD), CD52 was cleaved by elevated phospholipase C (PLC), and soluble CD52 (sCD52) was detected in the circulation. Inhibition and cleavage of CD52 significantly suppressed monocyte functions ex vivo and in vitro, and the predominance of immunosuppressive CD52low circulating monocytes in patients with AD was associated with infection and low transplant-free survival. ConclusionCD52 may represent a biologically relevant target for future immunotherapy. Stabilising CD52 may enhance monocyte functions and infection control in the context of cirrhosis, guided by sCD52/PLC as biomarkers indicating immuneparesis. Lay summaryRecurrent infections are a major cause of death in patients with liver cirrhosis. A fundamental understanding of the mechanisms that suppress immune responses in patients with cirrhosis is lacking, but required for the development of strategies to restore innate immunity in cirrhosis patients and prevent infection. The current study identified a novel marker for deficient immune responses and a potential target for such a future immune-based therapy. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=134 SRC="FIGDIR/small/587894v1_ufig1.gif" ALT="Figure 1"> View larger version (40K): org.highwire.dtl.DTLVardef@1a42b1borg.highwire.dtl.DTLVardef@10a0df2org.highwire.dtl.DTLVardef@1199087org.highwire.dtl.DTLVardef@15fb074_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical AbstractC_FLOATNO scRNA-seq identified seven circulating monocyte states, changing in cirrhosis patients at different stages of disease. Circulating monocytes overexpress CD52 in cirrhosis, but are absent in AD/ACLF due to PLC. CD52-expressing monocytes show high capability for phagocytosis, cytokine production, adhesion and migration potential and T cell suppression. Created with BioRender.com C_FIG

immunology↗

bioRxiv2023

B cell-stromal cell cross talk drives mesenteric lymph node eosinophilia during intestinal helminth infection

Eosinophils are involved in host protection against multicellular organisms including helminths and often participate in regulating long-lasting humoral responses. However, their recruitment to the gut-draining mesenteric lymph node (mLN), where they support the development of the adaptive immune response is still elusive. Here, we demonstrate the mechanism underlying the recruitment of eosinophils to the murine mLN post gastrointestinal helminth infection. We found that mLN eosinophils accumulated at immune interactive sites such as the interfollicular and paracortical regions in an IL-4R-dependent manner and was directly associated with the reduced availability of stromal derived eosinophil chemoattractants. Using multiplex imaging we confirmed that eosinophils associate within a stromal niche containing Lyve1+ lymphatic vessels, ER-TR7+Pdpn+ FRCs, and extrafollicular CD138+ plasma cells. Experiments utilising complete and mixed bone marrow chimeras demonstrated that mice lacking IL-4R expression or LT{beta} expression selectively on B cells had diminished eosinophilia and reduced extrafollicular plasma cell numbers within the mLN. When co-cultured with LT{beta}R activated FRCs, eosinophils gained an active phenotype with enhanced Il1rl1 (ST2) receptor expression. LT{beta}R ligation on FRCs resulted in enhanced IL-33 expression along with enrichment of distinct reactomes. Additionally, deletion of LT{beta}R in FRCs reduced the homing capability of eosinophils to the mLN, confirming the significance of lymphotoxin signalling in granulocyte recruitment. Overall, these results highlight the previously unknown role of B cell-stromal cell crosstalk in driving mLN eosinophilia and their potential role in regulating the quality and magnitude of the humoral immune response generated within the mLN.

immunology↗

bioRxiv2023

A novel cryopreservation and biobanking strategy to study lymphoid tissue stromal cells in human disease

Non-hematopoietic lymph node stromal cells (LNSCs) regulate lymphocyte trafficking, survival, and function for key roles in host defense, autoimmunity, alloimmunity, and lymphoproliferative disorders. However, study of LNSCs in human diseases is complicated by a dependence on viable lymphoid tissues, which are most often excised prior to establishment of a specific diagnosis. Here, we demonstrate that cryopreservation can be used to bank lymphoid tissue for the study of LNSCs in human disease. Using human tonsils, lymphoid tissue fragments were cryopreserved for subsequent enzymatic digestion and recovery of viable non-hematopoietic cells. Flow cytometry and single-cell transcriptomics identified comparable proportions of LNSC cell types in fresh and cryopreserved tissue. Moreover, cryopreservation had little effect on transcriptional profiles, which showed significant overlap between tonsils and lymph nodes. The presence and spatial distribution of transcriptionally defined cell types was confirmed by in situ analyses. Our broadly applicable approach promises to greatly enable research into the roles of LNSC in human disease.

immunology↗

bioRxiv2022

Long-term retention of antigens in germinal centres is controlled by the spatial organisation of the follicular dendritic cell network

Germinal centers (GCs) require sustained availability of antigens to promote antibody affinity maturation against pathogens and vaccines. A key source of antigens for GC B cells are immune complexes (ICs) displayed on follicular dendritic cells (FDCs). Here we show that FDC spatial organization regulates antigen dynamics in the GC. We show the existence of two light zone (LZ) FDC populations, which differ in the duration of antigen retention. While the entire light zone (LZ) FDC network captures ICs initially, only the central cells of the network function as a long-term antigen reservoir, where different antigens arriving from subsequent immunizations co-localize. Mechanistically, central FDCs constitutively express subtly higher CR2 membrane densities than peripheral FDCs, which strongly increases the IC retention half-life. Even though repeated immunizations gradually saturate central FDCs, B cell responses remain efficient because new antigens partially displace old ones. These results reveal the principles shaping antigen display on FDCs during the GC reaction.

immunology↗

bioRxiv2022

Rapid differentiation of regulatory CD4+ T cells in the infarcted myocardium blunts in situ inflammation

BackgroundMyocardial infarction (MI) is a sterile inflammatory condition associated with tissue injury that results in the activation of T helper cell targeting cardiac antigens. However, the differentiation trajectories and in situ activity of heart-specific CD4+T cells activated in the MI context remain poorly understood. MethodsHerein, we combined T-cell receptor transgenic models targeting myocardial protein, single-cell transcriptomics, and functional phenotyping to elucidate how the myosin-specific CD4+ T cells (TCR-M) differentiate in the murine infarcted myocardium and ultimately influence tissue repair. Furthermore, we adoptively transferred heart-specific T-cells that were pre-differentiated in vitro towards pro-inflammatory versus regulatory phenotypic states to dissect how they differentially regulate post-myocardial infarction (MI) inflammation. ResultsFlow cytometry and single-cell transcriptomics findings reveled that transferred TCR-M cells rapidly acquired an induced regulatory phenotype (iTreg) in the infarcted myocardium and blunt local inflammation. Myocardial TCR-M cells differentiated into two main lineages enriched with cell activation and pro-fibrotic transcripts (e.g. Tgfb1) or with suppressor immune checkpoints (e.g. Pdcd1), which we also found in human myocardial tissue. These cells produced high levels of latency-associated peptide (LAP) and inhibited interleukine-17 (IL-17) responses. Notably, TCR-M cells that were pre-differentiated in vitro towards a regulatory phenotype maintained a stable in vivo FOXP3 expression and anti-inflammatory activity when adoptively transferred prior to MI induction. In contrast, TCR-M cells that were pre-differentiated in vitro towards a pro-inflammatory TH17 phenotype were partially converted towards a regulatory phenotype in the injured myocardium and blunted myocardial inflammation. ConclusionsThese findings reveal that the myocardial milieu provides a suitable environment for iTreg differentiation and reveals novels mechanisms by which the healing myocardium shapes local immunological processes.

immunology↗

bioRxiv2022

TYPE I INTERFERON-DEPENDENT CELLULAR REMODELING DURING ACUTE PULMONARY MHV INFECTION

Impaired type I interferon (IFN) responses are predictive of severe disease during pulmonary coronavirus infection. In the periphery, dampened IFN-responsiveness is associated with viremia and hypercytokinemia, however the resolution of IFN-dependent innate immune responses in the lungs remains limited. Here, we aimed to elucidate the early dynamics of antiviral immunity and define the IFN-dependent mechanisms limiting viral spread during pulmonary infection with the murine hepatitis virus (MHV), a beta-coronavirus. While several innate immune cell types infiltrated the lungs concomitant with viral replication, the influx of type I IFN-responsive myeloid cells was essential for viral containment and prevention of fatal disease. Combining high-resolution transcriptomic analysis and genetic attenuation of interferon signaling, we delineated IFN-dependent cell-intrinsic and population-based transcriptional changes that determined viral replication and inflammatory maturation, respectively. While monocyte-derived macrophages exhibited the strongest pro-inflammatory transcriptional reprogramming during pulmonary infection, these maturation programs were impaired in the absence of IFN-signalling. Instead, IFN-deficient monocyte-derived macrophages expressed genes encoding for neutrophil attractants and IL1{beta}, resulting in enhanced lung injury. Our results reveal the requirement for type I IFN-induced antiviral states and inflammation-induced maturation programs to secure pulmonary viral containment.

immunology↗

bioRxiv2021

Fibroblastic reticular cells provide a supportive niche for lymph node-resident macrophages

The lymph node (LN) is home to resident macrophage populations that are essential for immune function and homeostasis. The T cell paracortical zone is a major site of macrophage efferocytosis of apoptotic cells, but key factors controlling this niche are undefined. Here we show that fibroblastic reticular cells (FRCs) are an essential component of the LN macrophage niche. Macrophages co-localised with FRCs in human LNs, and murine single-cell RNA-sequencing revealed that most reticular cells expressed master macrophage regulator CSF1. Functional assays showed that CSF1R signalling was sufficient to support macrophage development. In the presence of LPS, FRCs underwent a mechanistic switch and maintained support through CSF1R-independent mechanisms. These effects were conserved between mouse and human systems. Rapid loss of macrophages and monocytes from LNs was observed upon genetic ablation of FRCs. These data reveal a critically important role for FRCs in the creation of the parenchymal macrophage niche within LNs.

immunology↗

bioRxiv2021

Multi-tier mechanics control stromal adaptations in swelling lymph nodes

Lymph nodes (LNs) comprise two main structural elements: Fibroblastic reticular cells (FRCs) that form dedicated niches for immune cell interaction and capsular fibroblasts that build a shell around the organ. While LNs are fairly stable in size during homeostatic conditions, immunological challenge causes more than 10-fold increase in size within only a few days. How a solid organ can accommodate such extreme volumetric changes is poorly understood. Here, we characterize the biomechanics of LN swelling on the cellular and organ scale. We identify lymphocyte trapping by influx and proliferation as drivers of an outward pressure force, causing FRCs and their associated conduits to stretch. After an initial phase of relaxation, FRCs sense the resulting strain via cell matrix adhesions, which coordinates local growth and remodeling of the stromal network. While the expanded FRC network adopts its typical configuration, a massive fibrotic reaction of the organ capsule sets in and counters further organ expansion. Thus, different fibroblast populations mechanically control LN swelling in a multi-tier fashion.

immunology↗

bioRxiv2019

Perinatal Programming of Mucosal Stromal Cell Identity by the Lymphotoxin Pathway Regulates Mucosal Immune Responses in the Adult

Redundant mechanisms support IgA responses to intestinal antigens. These include multiple priming sites (mesenteric lymph nodes (MLN), Peyers patches and isolated lymphoid follicles) and various cytokines that promote class switch to IgA, even in the absence of T cells. In spite of these back-up mechanisms, vaccination against enteric pathogens such as Rotavirus has limited success in some populations. Genetic and environmental signals experienced during early life are known to influence mucosal immunity, yet the mechanisms for how these exposures operate remain unclear. Here we used Rotavirus infection to follow antigen-specific IgA responses through time and in different gut compartments. Using genetic and pharmacological approaches, we tested the role of a pathway known to support IgA responses (Lymphotoxin - LT) at different developmental stages. We found that LT-beta receptor (LT{beta}R) signalling in utero programs intestinal IgA responses in adulthood by affecting antibody class switch recombination to IgA and subsequent generation of IgA antibody-secreting cells within an intact MLN. In addition, in utero LT{beta}R signalling dictates the phenotype and function of MLN stromal cells in order to support IgA responses in the adult. Collectively, our studies uncover new mechanistic insights into how in utero LT{beta}R signalling impacts mucosal immune responses during adulthood. One Sentence SummaryEarly life LT{beta}R signalling is critical for programming the mesenteric lymph node stromal cell environment, impacting both antibody isotype switching to IgA and the differentiation of IgA+ antibody secreting cells. Graphic Abstract O_FIG O_LINKSMALLFIG WIDTH=190 HEIGHT=200 SRC="FIGDIR/small/557652v1_ufig1.gif" ALT="Figure 1"> View larger version (37K): org.highwire.dtl.DTLVardef@4780dborg.highwire.dtl.DTLVardef@5cbe85org.highwire.dtl.DTLVardef@e0465eorg.highwire.dtl.DTLVardef@2bcae6_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2018

Time-resolved systems analysis reveals a critical role of XCR1+ 1 dendritic cells in the maintenance of effector T cells during chronic viral infection

Upon a viral infection, the host immune system attempts to eradicate the virus. However, once the infection threat seems overwhelming, the infected host actively shuts down effector responses to reduce immunopathology. The price to pay for this is the establishment of a chronic infection that is only partially controlled by a lower level immune response. The genetic networks underlying this infection fate decision and the immune adaptation to the lower level response are not well understood. Here we used an integrated approach of gene coexpression network analysis of time-resolved splenic transcriptomes and immunological analysis to characterize the host response to acute and chronic lymphocytic choriomeningitis virus (LCMV) infections. We found first, an early attenuation of inflammatory monocyte/macrophage prior to the onset of T cell exhaustion and second, a critical role of the XCL1-XCR1 communication axis during the functional adaptation of the T cell response to the chronic infection state. These findings not only reveal an important feedback mechanism that couples T cell exhaustion with the maintenance of a lower level of effector T cell response but also suggest therapy options to better control virus levels during the chronic infection phase.\n\nAuthor SummaryThe outcomes of viral infections are the result of dynamic interplays between infecting viruses and induced host responses. They can be categorized as either acute or chronic depending on temporal virus-host relationships. Chronic infections are associated with immune exhaustion, a partial shut-down of effector responses. The processes underlying infection fate decisions are incompletely understood. Here we analyzed, on a systems level, infection-fate-specific gene signatures and the resulting adaptive processes of the host. We used the well-established lymphocytic choriomeningitis virus infection mouse model which has been instrumental to detect many fundamental processes in the virus-immune system crosstalk that are also relevant in human infections. We show an early attenuation of macrophage-mediated inflammation and an involvement of cross-presenting dendritic cells in the maintenance of an antiviral cytotoxic T cell response and virus control in the chronic infection phase. Together our data demonstrate a delicate adaptation process towards a chronic virus infection with both immunosuppressive and immunostimulatory processes. We fill a knowledge gap regarding the mechanisms of effector T cell maintenance and provide a new rational for targeted therapeutic vaccination.

immunology↗