Session 3 - Imaging and Spatial Transcriptomics
Chairs: Anja Hauser, Raluca Niesner, Berlin
Speaker: Adrien Hallou, Kennedey Institut, Oxford, UK
Over the past decade, the technologies and analysis to study cells in tissue context have evolved significantly. They now allow to comprehensively understand molecular mechanisms in health and disease, providing information about the effect of environmental and microenvironmental cues in the tissue. This includes effects of mechanical stimuli on cells. In this session, Adrien Hallou will give us an overview about the novel field of mechano-transcriptomics, which investigates how mechanical cues shape transcriptional profiles within the tissue context.
Adrien Hallou
Kennedey Institut, Oxford, UK
Mechanical Control of Macrophage Immunometabolism: Piezo1–Calcium Axis in Fracture Healing
Deniz Tasgin 1,2, Alex Fiedler 3, Ruth Leben 3, Robert Günther 2, Luise Asmussen 1,2, Sandy Kroh 1,2, Johanna Ehl 1,2, Ralf Uecker 2, Raluca Niesner 3, Anja E. Hauser 1,2
1 Charité – Universitätsmedizin Berlin, Department of Rheumatology and Clinical Immunology, Berlin, Germany.
2 Immune Dynamics, Deutsches Rheuma-Forschungszentrum Berlin, Germany.
3 Biophysical Analytics, Deutsches Rheuma-Forschungszentrum Berlin, Germany.
Mechanical cues are integral features of tissue microenvironments and become profoundly altered during chronic inflammation, fibrosis, and bone injury. While mechanotransduction is well characterized in neurons and stromal cells, emerging evidence suggests that immune cells also convert mechanical inputs into biochemical programs that shape inflammation and repair. Tissue-resident macrophages are prime candidates for such regulation because they adapt their polarization, cytokine output, and metabolism to local tissue demands. Mechanosensing may provide an additional regulatory layer, fine-tuning these cells to heterogeneous tissue contexts. Piezo1, a highly expressed mechanosensitive cation channel in macrophages, has been implicated in inflammatory signaling, yet how Piezo1-dependent calcium influx rewires macrophage metabolism in vivo, and how this impacts bone healing, remains unclear.
Our project will define Piezo1-mediated mechanotransduction in bone marrow macrophages and establish its relevance for fracture healing. Using intravital two-photon microscopy coupled to fluorescence lifetime imaging (FLIM), we will quantify macrophage metabolic states in their native niche via NAD(P)H lifetime signatures and link these readouts to Piezo1-associated calcium dynamics measured with genetically encoded calcium reporters such as GCaMP or TN-XXL in selected immune populations. In parallel, controlled in vitro perturbations of mechanical context and pharmacological Piezo1 modulation will causally map mechanical inputs to calcium signaling, metabolic switching, and polarization outputs. To test in vivo relevance, fracture healing after osteotomy will be evaluated in myeloid-specific Piezo1-deficient mice. Longitudinal outcomes will be assessed by micro-CT, histology, and multiepitope-ligand cartography to relate macrophage states to vascular, stromal, and inflammatory zones within the healing callus. By integrating intravital metabolic imaging, reporter mouse models, and spatial histology, this project will define how mechanical cues shape macrophage metabolism and test whether Piezo1 functions as a mechanometabolic regulator of bone repair.
Spatial and Functional Insights into Plasma Cell Longevity and Immune Memory Maintenance
Leonard Fiebig1, Florian Kralik, Heike Hirseland1, Axel R. Schulz1, Sandy Kroh1, Ralf Uecker1, Lisa-Marie Diekmann1, Antonia Niedobitek1, Simon Reinke2, Sebastian Hardt3, Schayan Yousefian4, Simon Haas4, Anja Hauser1, Henrik E. Mei1, Hyun-Dong Chang1
1 German Rheumatology Research Center (DRFZ) Berlin, a Leibniz Institute, Berlin, Germany,
2 Cell Harvesting Core, Berlin Institute of Health, Berlin, Germany
3 Center for Musculoskeletal Surgery, Charité Universitätsmedizin Berlin, Berlin, Germany
4 Max Delbrück Center – Berlin Institute for Medical Systems Biology
Mature antibody-secreting plasma cells (PCs) in human the bone marrow (BM) are critical for long-term humoral immunity and immunological memory, yet regulatory cues and tissue niches that support their persistence remain poorly understood. Here, we investigate the spatial, phenotypic, and functional heterogeneity of mature human BM PCs using an integrated approach combining spatial proteomics with primary BM culture systems.
We established a platform using fresh human femur heads that enables tissue sectioning and highly multiplexed spatial proteomic imaging of up to 55 markers without decalcification, which preserves sensitive epitopes that are often difficult to detect in conventional FFPE bone sections. In parallel, flow cytometric and secretome profiling of matched primary whole BM cultures from the same donors offers a complementary functional systeme to define survival requirements across the major BM memory cell compartments
Using this integrated workflow, we aim to localize plasma cell subsets within the BM microenvironment, characterize their interactions with stromal and immune cell populations associated with PC longevity, and functionally assess their dependence on key survival pathways.
Together, these analyses provide new insight into the cellular organization of the human BM niche and the mechanisms that sustain durable antibody and immune memory. Ultimately, we aim to discover new targets for therapies that selectively disrupt pathogenic plasma cells or immune memory compartments while sparing protective immune memory in autoimmune disease.