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Abstract Hydrogels are used extensively as cell-culture scaffolds for both 2D and 3D cell cultures due to their biocompatibility and the ease in which their mechanical and biological properties can be tailored to mimic natural tissue. The challenge when working with hydrogel-based scaffolds is in their handling, as hydrogels that mimic e.g. brain tissue, are both fragile and brittle when prepared as thin (sub-mm) membranes. Here, we describe a method for facile handling of thin hydrogel cell culture scaffolds by molding them onto a polycaprolactone (PCL) mesh support attached to a commonly used Transwell set-up in which the original membrane has been removed. In addition to demonstrating the assembly of this set-up, we also show some applications for this type of biological membrane. A polyethylene glycol (PEG)-gelatin hydrogel supports cell adhesion, and the structures can be used for biological barrier models comprising either one or multiple hydrogel layers. Here, we demonstrate the formation of a tight layer of an epithelial cell model comprising MDCK cells cultured over 9 days by following the build-up of the transepithelial electrical resistances. Second, by integrating a pure PEG hydrogel into the PCL mesh, significant swelling is induced, which leads to the formation of a non-adherent biological scaffold with a large curvature that is useful for spheroid formation. In conclusion, we demonstrate the development of a handling platform for hydrogel cell culture scaffolds for easy integration with conventional measurement techniques and miniaturized organs-on-chip systems.

Measurement(s) nucleus • F-actin • zonula occludens 1 Technology Type(s) confocal microscopy Factor Type(s) line width • medium glucose concentration Sample Characteristic - Organism Mus musculus Machine-accessible metadata file describing the reported data: https://doi.org/10.6084/m9.figshare.17129021

Abstract This work reports on an effort to decipher the alignment of brain microvasculature endothelial cells to physical constrains generated via adhesion control on hydrogel surfaces and explore the corresponding responses upon glucose level variations emulating the hypo- and hyperglycaemic effects in diabetes. We prepared hydrogels of hyaluronic acid a natural biomaterial that does not naturally support endothelial cell adhesion, and specifically functionalised RGD peptides into lines using UV-mediated linkage. The width of the lines was varied from 10 to 100 µm. We evaluated cell alignment by measuring the nuclei, cell, and F-actin orientations, and the nuclei and cell eccentricity via immunofluorescent staining and image analysis. We found that the brain microvascular endothelial cells aligned and elongated to these physical constraints for all line widths. In addition, we also observed that varying the cell medium glucose levels affected the cell alignment along the patterns. We believe our results may provide a platform for further studies on the impact of altered glucose levels in cardiovascular disease.

Abstract The generation of hydrogel droplets using droplet microfluidics has emerged as a powerful tool with many applications in biology and medicine. Here, a microfluidic system to control the position of particles (beads or astrocyte cells) in hydrogel droplets using bulk acoustic standing waves is presented. The chip consisted of a droplet generator and a 380 µm wide acoustic focusing channel. Droplets comprising hydrogel precursor solution (polyethylene glycol tetraacrylate or a combination of polyethylene glycol tetraacrylate and gelatine methacrylate), photoinitiator and particles were generated. The droplets passed along the acoustic focusing channel where a half wavelength acoustic standing wave field was generated, and the particles were focused to the centre line of the droplets (i.e. the pressure nodal line) by the acoustic force. The droplets were cross-linked by exposure to UV-light, freezing the particles in their positions. With the acoustics applied, 89 ± 19% of the particles (polystyrene beads, 10 µm diameter) were positioned in an area ± 10% from the centre line. As proof-of-principle for biological particles, astrocytes were focused in hydrogel droplets using the same principle. The viability of the astrocytes after 7 days in culture was 72 ± 22% when exposed to the acoustic focusing compared with 70 ± 19% for samples not exposed to the acoustic focusing. This technology provides a platform to control the spatial position of bioparticles in hydrogel droplets, and opens up for the generation of more complex biological hydrogel structures.