The cornea serves as a transparent protective barrier for the eye, with epithelial homeostasis and renewal critically dependent on limbal epithelial stem cells residing in a specialized niche characterized by stromal invaginations that form limbal crypts. Elegant advanced in vitro models of the limbal niche have been described, but most are technologically demanding, requiring multi-step fabrication that limits scalability. We developed a simple and rapid strategy to fabricate 3D scaffolds with undulating topography using a shape-morphing hydrogel concept. A photocrosslinkable bioink composed of methacrylated collagen, hyaluronic acid, and silk fibroin was patterned using grayscale UV projection. Digital Light Processing technology enabled spatially controlled heterogeneous crosslinking densities in a single-step process. The undulating topography was then formed through differential shrinking dynamics of the hydrogel at 37 °C; specifically, the volume of areas exposed to lower UV doses decreased while areas exposed to higher UV doses remained stable. The addition of silk fibroin proved essential for both temperature-dependent shape morphing and sustained corneal epithelial cell adhesion and growth. The undulating scaffolds supported epithelial stratification for at least three weeks in culture. Physiologically relevant corneal mechanotransduction and apicobasal organization was achieved, with nuclear YAP localization in soft areas, P63-positive progenitor cells enriched in basal layers and PAX6-positive differentiated cells in apical layers. Additionally, progressive epithelial maturation was demonstrated by increased CK3 expression, establishment of tight junctions, and the deposition of a basement membrane. STATEMENT OF SIGNIFICANCE: Paschalidis et al. This work offers dual significance. First, we introduce an innovative, simple and single-step bioprinting approach to create 3D scaffolds with complex topography. Using Digital Light Processing technology, a silk fibroin-containing photocurable bioink is exposed to spatially heterogeneous UV doses. This enables differential crosslinking densities in the printed scaffold, which undergoes programmable shape-morphing. Second, we applied this methodology to engineer advanced corneal models that recapitulate the structural features of the epithelial stem cell niche, an undulating topography. This addresses critical needs in human disease modeling and preclinical testing while reducing animal use. The resulting model sustains three-week cultures and faithfully mimics physiological epithelial organization, with progenitor markers expressed in basal layers and differentiation markers indicating barrier function in apical layers.

Tissue engineering holds great promise for regenerative medicine, drug discovery, and as an alternative to animal models. However, as soon as the dimensions of engineered tissue exceed the diffusion limit of oxygen and nutriments, a necrotic core forms leading to irreversible damage. To overcome this constraint, the establishment of a functional perfusion network is essential. In this work, digital light processing bioprinting is used to encapsulate endothelial progenitor cells (EPCs) in 3D light-cured hydrogel scaffolds to guide them toward vascular network formation. In these scaffolds, EPCs proliferate and self-organize within a few days into branched tubular structures with predefined geometry, forming capillary-like vascular tubes or trees of diameters in the range of 10 to 100 µm. Presenting a confluent monolayer wall of cells strongly connect by tight junctions around a central lumen-like space, these structures can be microinjected with a fluorescent dye and are stable for several weeks in vitro. These endothelial structures can be recovered and manipulated in an alginate patch without altering their shape or viability. This approach opens new opportunities for future applications, such as stacking with other cell sheets or multicellular constructs to yield bioengineered tissue with higher complexity and functionality.

Many tissue engineering approaches are being explored to offer solutions for diseased esophageal tissue and its repair. However, classical techniques in tissue engineering are not capable of recapitulating the structural and functional parameters of native esophagi. Esophageal 3D bioprinting is yet an emerging field but holds great promise in meeting this challenge. Herein, the use of extrusion‐based 3D printing is examined to generate esophageal substitutes from a polymer blend‐based formulation. The fabricated 3D esophageal structures are printed at a very high speed without collapse and without the use of supporting material. In vitro analysis reveals that the printed material enables cell adhesion, proliferation, and migration into the deeper zones. Moreover, the designed construct is suturable to the native esophagus and exhibits no leakage while offering mechanical properties similar to that of the native tissue. Overall, these biochemical and biomechanical features make the reported artificial esophagus a promising solution for the repair of damaged esophagi.

Primary hepatocytes are essential cellular resources for drug screening and medical transplantation. While culture systems have already succeeded in reconstituting the biomimetic microenvironment of primary hepatocytes, acquiring additional capabilities to handle them easily as well as to expand them remains unmet needs. This paper describes a culture system for primary rat hepatocytes, based on cell fiber technology, that brings scalability and handleability. Cell fibers are cell-laden core–shell hydrogel microfibers; in the core regions, cells are embedded in extracellular matrix proteins, cultured three-dimensionally, and exposed to soluble growth factors in the culture medium via the hydrogel shells. By encapsulating primary rat hepatocytes within cell fibers, we first demonstrated their proliferation while maintaining their viability and their hepatic specific functions for up to thirty days of subsequent culture. We then demonstrated the efficiency of proliferating primary rat hepatocytes in cell fibers not only as cell-based sensors to detect drugs that damage hepatic functions and hepatocellular processes but also as transplants to improve the plasma albumin concentrations of congenital analbuminemia. Our culture system could therefore be included in innovative strategies and promising developments in applying primary hepatocytes to both pharmaceutical and medical fields.

Cholangiocytes, biliary epithelial cells, are known to spontaneously self-organize into spherical cysts with a central lumen. In this work, we explore a promising biocompatible stereolithographic approach to encapsulate cholangiocytes into geometrically controlled 3D hydrogel structures to guide them towards the formation of branched tubular networks. We demonstrate that within the appropriate mix of hydrogels, normal rat cholangiocytes can proliferate, migrate, and organize into branched tubular structures with walls consisting of a cell monolayer, transport fluorescent dyes into the luminal space, and show markers of epithelial maturation such as primary cilia and continuous tight junctions. The resulting structures have dimensions typically found in the intralobular and intrahepatic biliary tree and are stable for weeks, without any requirement of bulk supporting material, thereby offering total access to the external side of these biliary epithelial constructs.

In esophageal pathologies, such as esophageal atresia, cancers, caustic burns, or post-operative stenosis, esophageal replacement is performed by using parts of the gastrointestinal tract to restore nutritional autonomy. However, this surgical procedure most often does not lead to complete functional recovery and is instead associated with many complications resulting in a decrease in the quality of life and survival rate. Esophageal tissue engineering (ETE) aims at repairing the defective esophagus and is considered as a promising therapeutic alternative. Noteworthy progress has recently been made in the ETE research area but strong challenges remain to replicate the structural and functional integrity of the esophagus with the approaches currently being developed. Within this context, 3D bioprinting is emerging as a new technology to facilitate the patterning of both cellular and acellular bioinks into well-organized 3D functional structures. Here, we present a comprehensive overview of the recent advances in tissue engineering for esophageal reconstruction with a specific focus on 3D bioprinting approaches in ETE. Current biofabrication techniques and bioink features are highlighted, and these are discussed in view of the complexity of the native esophagus that the designed substitute needs to replace. Finally, perspectives on recent strategies for fabricating other tubular organ substitutes via 3D bioprinting are discussed briefly for their potential in ETE applications.

This review summarizes recent advances in micro- and nanopore technologies with a focus on the functionalization of pores using a promising method named contactless electro-functionalization (CLEF). CLEF enables the localized grafting of electroactive entities onto the inner wall of a micro- or nano-sized pore in a solid-state silicon/silicon oxide membrane. A voltage or electrical current applied across the pore induces the surface functionalization by electroactive entities exclusively on the inside pore wall, which is a significant improvement over existing methods. CLEF's mechanism is based on the polarization of a sandwich-like silicon/silicon oxide membrane, creating electronic pathways between the core silicon and the electrolyte. Correlation between numerical simulations and experiments have validated this hypothesis. CLEF-induced micro- and nanopores functionalized with antibodies or oligonucleotides were successfully used for the detection and identification of cells and are promising sensitive biosensors. This technology could soon be successfully applied to planar configurations of pores, such as restrictions in microfluidic channels.

Technical progress in materials science and bioprinting has for the past few decades fostered considerable advances in medicine. More recently, the understanding of the processes of self-organization of cells into three-dimensional multicellular structures and the study of organoids have opened new perspectives for tissue engineering. Here, we review microengineering approaches for building functional tissues, and discuss recent progress in the understanding of morphogenetic processes and in the ability to steer them in vitro. On the basis of biological and technical considerations, we emphasize the achievements and remaining challenges of bringing together microengineering and morphogenesis. Our viewpoint underlines the importance of cellular self-organization for the success of tissue engineering in therapeutic applications. We reason that directed self-organization, at the convergence of microengineering and cellular self-organization, is a promising direction for the manufacturing of complex functional tissues.

Since 2014, the Multi Unit Spectroscopic Explorer (MUSE) instrument generates hyperspectral datacubes (300 by 300 pixels by 3600 wavelength in the visible range) of the deep Universe. One of the main purposes of the wide field spectrograph MUSE is to analyse galaxies and their surroundings by the study of their spectra. Galaxy spectra are composed of a continuum emission and of sparse emission (or absorption) peaks. On the contrary surrounding gas only contains peak such as the Lyman alpha emission line. Several methods are developed here to detect the gas signature as far as possible in the galaxy surroundings. These methods combined clustering approaches and several pre-processing steps.

This paper presents an overview of criteria and methods in multiple testing, with an emphasis on the false discovery rate control. The popular Benjamini and Hochberg procedure is described. The rationale for this approach is explained through a simple Bayesian interpretation. Some state-of-the-art variations and extensions are also presented.

This paper proposes a multi-branch model to deal with Remaining Useful Life (RUL) estimation problem in the case where several deterioration modes co-exist within a single component. By basing on Hidden semi-Markov Models (HsMM), the component is supposed to pass through some discrete and unobservable health states until it fails. The rate and the manner of these state transitions, however, depend on the mode of deterioration that is actually active. We show that by taking into account the co-existence of different deterioration modes, the multi-branch model can help to improve prognosis results, which is essential for the implementation of a predictive maintenance. A practical case study is investigated to evaluate the advantages of the proposed model.

Video microscopy offers outstanding capabilities to investigate the dynamics of biological and pathological mechanisms in optimal culture conditions. Contact imaging is one of the simplest imaging architectures to digitally record images of cells due to the absence of any objective between the sample and the image sensor. However, in the framework of in-line holography, other optical components, e.g., an optical filter or a pinhole, are placed underneath the light source in order to illuminate the cells with a coherent or quasi-coherent incident light. In this study, we demonstrate that contact imaging with an incident light of both limited temporal and spatial coherences can be achieved with sufficiently high quality for most applications in cell biology, including monitoring of cell sedimentation, rolling, adhesion, spreading, proliferation, motility, death and detachment. Patterns of cells were recorded at various distances between 0 and 1000 μm from the pixel array of the image sensors. Cells in suspension, just deposited or at mitosis focalise light into photonic nanojets which can be visualised by contact imaging. Light refraction by cells significantly varies during the adhesion process, the cell cycle and among the cell population in connection with every modification in the tridimensional morphology of a cell.

La nature stochastique de l’expression des genes au niveau cellulaire a ete demontree par un grand nombre d’etudes. La plupart de ces etudes ont ete menees sur des organismes unicellulaires procaryotes (notamment chez Escherichia coli) ou eucaryotes (chez la levure Saccharomyces cerevisiae). Chez les eucaryotes superieurs, les travaux pionniers effectues par Levsky conduisent a des conclusions similaires....

Planar patch-clamp is a two-dimensional variation of traditional patch-clamp. By contrast to classical glass micropipette, the seal quality of silicon patch-clamp chips (i.e. seal resistance and seal success rate) have remained poor due to the planar geometry and the nature of the substrate and thus partially obliterate the advantages related to planar patch-clamp. The characterization of physical parameters involved in seal formation is thus of major interest. In this paper, we demonstrate that the physical characterization of surfaces by a set of techniques (Atomic Force Microscopy (AFM), Scanning Electron Microscopy (SEM), X-ray Photoelectron Spectroscopy (XPS), surface energy (polar and dispersive contributions), drop angles, impedance spectroscopy, combined with a statistical design of experiments (DOE)) allowed us discriminating chips that provide relevant performances for planar patch-clamp analysis. Analyses of seal quality demonstrate that dispersive interactions and micropore size are the most crucial physical parameters of chip surfaces, by contrast to surface roughness and dielectric membrane thickness. This multi-scale study combined with electrophysiological validation of chips on a diverse set of cell-types expressing various ion channels (IRK1, hERG and hNa(v)1.5 channels) unveiled a suitable patch-clamp chip candidate. This original approach may inspire novel strategies for selecting appropriate surface parameters dedicated to biochips.

To date, most HCA (High Content Analysis) studies are carried out with adherent cell lines grown on a homogenous substrate in tissue-culture treated micro-plates. Under these conditions, cells spread and divide in all directions resulting in an inherent variability in cell shape, morphology and behavior. The high cell-to-cell variance of the overall population impedes the success of HCA, especially for drug development. The ability of micropatterns to normalize the shape and internal polarity of every individual cell provides a tremendous opportunity for solving this critical bottleneck 1-2. To facilitate access and use of the micropatterning technology, CYTOO has developed a range of ready to use micropatterns, available in coverslip and microwell formats. In this video article, we provide detailed protocols of all the procedures from cell seeding on CYTOOchip micropatterns, drug treatment, fixation and staining to automated acquisition, automated image processing and final data analysis. With this example, we illustrate how micropatterns can facilitate cell-based assays. Alterations of the cell cytoskeleton are difficult to quantify in cells cultured on homogenous substrates, but culturing cells on micropatterns results in a reproducible organization of the actin meshwork due to systematic positioning of the cell adhesion contacts in every cell. Such normalization of the intracellular architecture allows quantification of even small effects on the actin cytoskeleton as demonstrated in these set of protocols using blebbistatin, an inhibitor of the actin-myosin interaction.

The cover picture illustrates an innovative functionalization technique, referred to as “contactless electrofunctionalization” (CLEF). The inner surface of a pore etched in a dielectric membrane can be easily coated in a single step and in physiologically compatible conditions. Localized deposition of polypyrrole copolymers bearing oligonucleotides, as well as metallic oxides, are experimentally demonstrated. Both are selectively deposited on the pore walls with no additional deposition onto the dielectric membrane. CLEF thus constitutes a significant improvement over classically used surface-modification techniques. Its high efficiency makes it a promising technique, particularly for biosensor applications. For more information, please read the Full Paper “Contactless Electrofunctionalization of a Single Pore” by A. Bouchet, V. Haguet, et al., beginning on page 2297.

Die Bildung von Regentropfen auf einer Blattoberfläche spiegelt eine wesentliche Eigenschaft von Wasser wider: die hohe Oberflächenspannung. Eine künstliche Anordnung sehr einheitlicher Tropfen lässt sich auf einer lithographisch bemusterten Glasoberfläche mit einer differenziellen Oberflächenenergie erzeugen. M. Y. Balakirev et al. schildern in der Zuschrift auf S. 7775 ff. die Verwendung dieser Tropfen für die Synthese kleiner Moleküle in Lösung und die anschließende quantitative Hochdurchsatzanalyse der Enzymkinetik.