Laminin and hyaluronan supplementation of collagen hydrogels enhances endothelial function and tight junction expression on three-dimensional cylindrical microvessel-on-a-chip.

Vasculature-on-chip (VoC) models have become a prominent tool in the study of microvasculature functions because of their cost-effective and ethical production process. These models typically use a hydrogel in which the three-dimensional (3D) microvascular structure is embedded. Thus, VoCs are directly impacted by the physical and chemical cues of the supporting hydrogel. Endothelial cell (EC) response in VoCs is critical, especially in organ-specific vasculature models, in which ECs exhibit specific traits and behaviors that vary between organs. Many studies customize the stimuli ECs perceive in different ways; however, customizing the hydrogel composition accordingly to the target organ's extracellular matrix (ECM), which we believe has great potential, has been rarely investigated. We explored this approach to organ-specific VoCs by fabricating microvessels (MVs) with either human umbilical vein ECs or human brain microvascular ECs in a 3D cylindrical VoC using a collagen hydrogel alone or one supplemented with laminin and hyaluronan, components found in the brain ECM. We characterized the physical properties of these hydrogels and analyzed the barrier properties of the MVs. Barrier function and tight junction (ZO-1) expression improved with the addition of laminin and hyaluronan in the composite hydrogel.

Phenotypic and transcriptional characterization of oligodendrocyte precursor cells in a 3D culture.

Remyelination of the central nervous system (CNS) is a regenerative response that depends on the development of oligodendrocyte precursor cells (OPCs), which are generated from neural stem cells in developmental stages and exist as tissue stem cells in the adult CNS. Three-dimensional (3D) culture systems that recapitulate the complexity of the in vivo microenvironment are important for understanding the behavior of OPCs in remyelination and for exploring effective therapeutic approaches. In general, functional analysis of OPCs has mainly used two-dimensional (2D) culture systems; however, the differences between the properties of OPCs cultured in 2D and 3D have not been fully elucidated despite cellular functions being affected by the scaffold. In this study, we analyzed the phenotypic and transcriptomic differences in OPCs from 2D and collagen gel-based 3D cultures. In the 3D culture, the OPCs exhibited less than half ratio of proliferation and almost half ratio of differentiation to mature oligodendrocytes, compared to the 2D culture in the same culturing period. RNA-seq data showed robust changes in the expression level of genes associated with oligodendrocyte differentiation, and there were more up-regulated genes than down-regulated genes in 3D cultures compared to 2D cultures. In addition, the OPCs cultured in collagen gel scaffolds at lower collagen fiber densities showed higher proliferation activity compared with those cultured in collagen gel with higher collagen fiber densities. Our findings have identified the effect of culture dimension as well as the complexity of the scaffold on OPC responses at the cellular and molecular levels.

3D Printed lattice-structured metal electrodes for enhanced current production in bioelectrochemical systems.

Bioelectrochemical systems (BESs) are emerging techniques that use biological production of current for versatile activities, including energy recovery and bioremediation. The development of high-performance three-dimensional (3D) electrodes has attracted attention for facilitating current production in BESs. Carbon-based electrodes have been commonly used in BESs, but metal electrodes are not generally employed because of their low biocompatibility with microbes. In this study, 3D stainless-steel electrodes, composed of octahedral lattice, were fabricated using the 3D printing technique. Heat treatment was conducted to form an iron-oxide layer on the electrode surface for increasing biocompatibility. Another crucial parameter that determines current production is the pitch length of a lattice electrode as it affects the surface area and substrate diffusion. The pitch length was optimized by testing the lattice electrodes with pitches ranging from 1.5 mm to 6.0 mm. The highest current, obtained with the 3.0 mm-pitch electrode, was 50% higher than that obtained with common 3D carbon-felt electrodes. These results demonstrate the usefulness of 3D lattice-structured metal electrodes in BESs.

Image-based crosstalk analysis of cell-cell interactions during sprouting angiogenesis using blood-vessel-on-a-chip.

BACKGROUND: Sprouting angiogenesis is an important mechanism for morphogenetic phenomena, including organ development, wound healing, and tissue regeneration. In regenerative medicine, therapeutic angiogenesis is a clinical solution for recovery from ischemic diseases. Mesenchymal stem cells (MSCs) have been clinically used given their pro-angiogenic effects. MSCs are reported to promote angiogenesis by differentiating into pericytes or other vascular cells or through cell-cell communication using multiple protein-protein interactions. However, how MSCs physically contact and move around ECs to keep the sprouting angiogenesis active remains unknown. METHODS: We proposed a novel framework of EC-MSC crosstalk analysis using human umbilical vein endothelial cells (HUVECs) and MSCs obtained from mice subcutaneous adipose tissue on a 3D in vitro model, microvessel-on-a-chip, which allows cell-to-tissue level study. The microvessels were fabricated and cultured for 10 days in a collagen matrix where MSCs were embedded. RESULTS: Immunofluorescence imaging using a confocal laser microscope showed that MSCs smoothed the surface of the microvessel and elongated the angiogenic sprouts by binding to the microvessel's specific microstructures. Additionally, three-dimensional modeling of HUVEC-MSC intersections revealed that MSCs were selectively located around protrusions or roots of angiogenic sprouts, whose surface curvature was excessively low or high, respectively. CONCLUSIONS: The combination of our microvessel-on-a-chip system for 3D co-culture and image-based crosstalk analysis demonstrated that MSCs are selectively localized to concave-convex surfaces on scaffold structures and that they are responsible for the activation and stabilization of capillary vessels.

Nailfold capillary patterns correlate with age, gender, lifestyle habits, and fingertip temperature.

Nailfold capillaroscopy is a simple and noninvasive imaging tool to visualize the pattern of capillaries. Microvascular abnormalities have been previously observed in autoimmune disease such as systemic sclerosis and diabetes. Thus, early detection of microvascular dysfunction or changes has promising way for the one of the disease preventions. In this study, for routine health checkups, we evaluated the relationship between the structure of nailfold capillaries and lifestyle habits in healthy participants. First, we analyzed the correlation of structural parameters of nailfold capillaries with values of responses to questions on their lifestyle habits in 224 participants. The results suggested that an unhealthy lifestyle, including poor sleeping habits, smoking, intense exercise, and drinking alcohol, causes a change in the pattern of nailfold capillaries. We then investigated whether the pattern of nailfold capillaries changed after a conscious improvement in lifestyle habits. One to two weeks after the self-improvement of lifestyle habits, the hairpin loops sharpened or straightened. In conclusion, this study is the first report indicating a correlation between the structure of nailfold capillaries and lifestyle habits in a non-clinical population. The simple, inexpensive, and noninvasive method using nailfold microscopy can be employed for routine health checkups everywhere even at a bedside.

A 3D tissue model-on-a-chip for studying the effects of human senescent fibroblasts on blood vessels.

All human tissues experience aging that eventually causes organ dysfunction and disease. Cellular senescence was discovered in fibroblasts cultured in vitro. In adults, it is a primary defense mechanism against cancer, but also a major contributor to lifespan limits and disorders associated with aging. To assess how human blood vessels change in an aged environment, we developed an elementary tissue model-on-a-chip that comprises an in vitro three-dimensional model of a blood vessel embedded in a collagen gel with young or senescent skin fibroblasts. We found that senescent fibroblasts mechanically altered the surrounding extracellular matrix by exerting excessive traction stress. We then found that senescent fibroblasts induced sprouting angiogenesis of a microvessel via their senescence-associated secretory phenotype (SASP). Finally, we gathered evidence that the mechanical changes of the microenvironment play a role in sustaining SASP-induced angiogenesis. The model proved useful in monitoring morphological changes in blood vessels induced by senescent fibroblasts while controlling the proportion of senescent cells, and enabled the study of SASP inhibitors, a class of drugs useful in aging and cancer research.