Sebastian Ehrig

Spatial transcriptomics (ST) methods unlock molecular mechanisms underlying tissue development, homeostasis, or disease. However, there is a need for easy-to-use, high-resolution, cost-efficient, and 3D-scalable methods. Here, we report Open-ST, a sequencing-based, open-source experimental and computational resource to address these challenges and to study the molecular organization of tissues in 2D and 3D. In mouse brain, Open-ST captured transcripts at subcellular resolution and reconstructed cell types. In primary head-and-neck tumors and patient-matched healthy/metastatic lymph nodes, Open-ST captured the diversity of immune, stromal, and tumor populations in space, validated by imaging-based ST. Distinct cell states were organized around cell-cell communication hotspots in the tumor but not the metastasis. Strikingly, the 3D reconstruction and multimodal analysis of the metastatic lymph node revealed spatially contiguous structures not visible in 2D and potential biomarkers precisely at the 3D tumor/lymph node boundary. All protocols and software are available at https://rajewsky-lab.github.io/openst.

Surface curvature both emerges from, and influences the behavior of, living objects at length scales ranging from cell membranes to single cells to tissues and organs. The relevance of surface curvature in biology is supported by numerous experimental and theoretical investigations in recent years. In this review, first, a brief introduction to the key ideas of surface curvature in the context of biological systems is given and the challenges that arise when measuring surface curvature are discussed. Giving an overview of the emergence of curvature in biological systems, its significance at different length scales becomes apparent. On the other hand, summarizing current findings also shows that both single cells and entire cell sheets, tissues or organisms respond to curvature by modulating their shape and their migration behavior. Finally, the interplay between the distribution of morphogens or micro-organisms and the emergence of curvature across length scales is addressed with examples demonstrating these key mechanistic principles of morphogenesis. Overall, this review highlights that curved interfaces are not merely a passive by-product of the chemical, biological, and mechanical processes but that curvature acts also as a signal that co-determines these processes.

The collective self-organization of cells into three-dimensional structures can give rise to emergent physical properties such as fluid behavior. Here, we demonstrate that tissues growing on curved surfaces develop shapes with outer boundaries of constant mean curvature, similar to the energy minimizing forms of liquids wetting a surface. The amount of tissue formed depends on the shape of the substrate, with more tissue being deposited on highly concave surfaces, indicating a mechano-biological feedback mechanism. Inhibiting cell-contractility further revealed that active cellular forces are essential for generating sufficient surface stresses for the liquid-like behavior and growth of the tissue. This suggests that the mechanical signaling between cells and their physical environment, along with the continuous reorganization of cells and matrix is a key principle for the emergence of tissue shape.

The complex arrangement of the extracellular matrix (ECM) produced by cells during tissue growth, healing and remodelling is fundamental to tissue function. In connective tissues, it is still unclear how both cells and the ECM become and remain organized over length scales much larger than the distance between neighbouring cells. While cytoskeletal forces are essential for assembly and organization of the early ECM, how these processes lead to a highly organized ECM in tissues such as osteoid is not clear. To clarify the role of cellular tension for the development of these ordered fibril architectures, we used an in vitro model system, where pre-osteoblastic cells produced ECM-rich tissue inside channels with millimetre-sized triangular cross sections in ceramic scaffolds. Our results suggest a mechanical handshake between actively contracting cells and ECM fibrils: the build-up of a long-range organization of cells and the ECM enables a gradual conversion of cell-generated tension to pre-straining the ECM fibrils, which reduces the work cells have to generate to keep mature tissue under tension.

Abstract Spatial transcriptomics (ST) methods have been developed to unlock molecular mechanisms underlying tissue development, homeostasis, or disease. However, there is a need for easy-to-use, high-resolution, cost-efficient, and 3D-scalable methods. Here, we report Open-ST, a sequencing-based, open-source experimental and computational resource to address these challenges and to study the molecular organization of tissues in 3D. In mouse brain, Open-ST captured transcripts at subcellular resolution and reconstructed cell types. In primary tumor and patient-matched healthy/metastatic lymph nodes, Open-ST captured the diversity of immune, stromal and tumor populations in space. Distinct cell states were organized around cell-cell communication hotspots in the tumor, but not the metastasis. Strikingly, the 3D reconstruction and multimodal analysis of the metastatic lymph node revealed spatially contiguous structures not visible in 2D and potential biomarkers precisely at the 3D tumor/lymph node boundary. We anticipate Open-ST to accelerate the identification of spatial molecular mechanisms in 2D and 3D.