Staff member

Continuous monitoring of cell-to-cell interactions and multicellular network dynamics is essential for in situ biosensing, drug screening and cell-based assays. However, conventional live-cell imaging relies on benchtop optical microscopes placed outside the incubator, which limits throughput and makes long-term, minimally perturbative monitoring difficult. Here, we present a chip-sized, lensless digital holographic microscope (CSM) designed for compact, scalable, low-cost and in-incubator imaging of live-cell assays. The CSM combines a CMOS sensor and a micro-LED array in an inline holographic configuration, enabling wide-field, label-free imaging directly inside a standard CO2 incubator. The CSM was validated using a Vascular Endothelial Growth Factor (VEGF)-induced tube formation assay with primary human umbilical vein endothelial cells (HUVECs). Quantitative network metrics (total tube length, number of junctions and meshes, and mean mesh area) are extracted from numerically reconstructed images and are shown to capture dose-dependent effects of VEGF. At 24 h, the CSM resolved significant increases in tube length and junction count at 20-40 ng/mL VEGF-165, followed by a marked regression at 50 ng/mL, and achieved a morphological detection limit of 1383 mu m2 in mesh area with a corresponding morphological dynamic range of 1643. Time-lapse imaging every 30 min further resolves the full temporal evolution of tube formation and regression, revealing kinetic differences that are not accessible from endpoint measurements alone. These results establish the CSM as a versatile platform for quantitative, in-incubator monitoring of angiogenesis assays, bridging continuous imaging and endpoint read-outs.

JTD

Microscopy is a fundamental tool in biological research. However, conventional microscopes require manual operation and depend on user and equipment availability, limiting their suitability for continuous observation. Moreover, their size and complexity make them impractical for in situ experimentation. In this work, we present a novel, compact, affordable, and portable microscope that enables continuous in situ monitoring by being placed directly on biological samples. This chip-sized lensless holographic microscope (CLHM) is specifically designed to overcome the limitations of traditional microscopy. The device consists solely of an ultra-compact, state-of-the-art micro-LED display and a CMOS sensor, all enclosed within a 3D-printed housing. This unique light source enables a size that is markedly smaller than any comparable technology, allowing a resolution of 2.19 mu m within a 7 mm distance between the light source and the camera. This paper demonstrates the CLHM's versatility by monitoring in vitro models and performing whole-organism morphological analyses of small specimens. These experiments underscore its potential as an on-platform sensing device for continuous, in situ biological monitoring across diverse models.

JTD Keywords: Angiogenesis, Compact microscope, Fermentation, Holography, Lab-on-a-chip, Lensless, Real-time monitoring, Zebrafish

Intestinal organoids capture essential features of the intestinal epithelium such as crypt folding, cellular compartmentalization and collective movements. Each of these processes and their coordination require patterned forces that are at present unknown. Here we map three-dimensional cellular forces in mouse intestinal organoids grown on soft hydrogels. We show that these organoids exhibit a non-monotonic stress distribution that defines mechanical and functional compartments. The stem cell compartment pushes the extracellular matrix and folds through apical constriction, whereas the transit amplifying zone pulls the extracellular matrix and elongates through basal constriction. The size of the stem cell compartment depends on the extracellular-matrix stiffness and endogenous cellular forces. Computational modelling reveals that crypt shape and force distribution rely on cell surface tensions following cortical actomyosin density. Finally, cells are pulled out of the crypt along a gradient of increasing tension. Our study unveils how patterned forces enable compartmentalization, folding and collective migration in the intestinal epithelium. Perez-Gonzalez et al. explore the mechanical properties of intestinal organoids, and report the existence of distinct mechanical domains and that cells are pulled out of the central crypt along a gradient of increasing tension.

JTD Keywords: Forces, Growth, Gut, Monolayers, Morphogenesis, Reveal, Stem-cells, Tension

Different mechanisms are triggered when tissue is exposed to a biomaterial. The success of the biomaterial targeted process, like the release of chemicals, promoted angiogenesis, tissue regeneration, etc. depends on its integration in the tissue [1]. Studying this interaction in vivo requires the ability to image simultaneously deep immersed proteins and biomaterials with high resolution and low damage. Several methods offer solutions but only multiphoton microscopy (MM) has the ability to image with high resolution deep inside the sample. Why is not MM more extensively applied as a platform for investigating biomaterial integration in vivo? The high cost of the typical source for multiphoton microscopy is a clear limitation. Furthermore, imaging several channels simultaneously becomes out of reach for most of the labs.

JTD

Cell function depends on tissue rigidity, which cells probe by applying and transmitting forces to their extracellular matrix, and then transducing them into biochemical signals. Here we show that in response to matrix rigidity and density, force transmission and transduction are explained by the mechanical properties of the actin-talin-integrin-fibronectin clutch. We demonstrate that force transmission is regulated by a dynamic clutch mechanism, which unveils its fundamental biphasic force/rigidity relationship on talin depletion. Force transduction is triggered by talin unfolding above a stiffness threshold. Below this threshold, integrins unbind and release force before talin can unfold. Above the threshold, talin unfolds and binds to vinculin, leading to adhesion growth and YAP nuclear translocation. Matrix density, myosin contractility, integrin ligation and talin mechanical stability differently and nonlinearly regulate both force transmission and the transduction threshold. In all cases, coupling of talin unfolding dynamics to a theoretical clutch model quantitatively predicts cell response.

JTD