Published papers

This paper presents a bead-shaped hydrogel biochemical sensor. The hydrogel bead shows volumetric change as a response triggered by DNA-aptamers incorporated in the hydrogel. DNA-aptamers react to specific targets, triggering the structural shift of the aptamer (from linear to hairpin) and causing gel shrinkage. Pre-gel solution containing DNA-aptamer were ejected from capillaries to form large-scale (average diameter: ~2.37 mm) and micro-scale (average diameter: ~457 μm) droplets by outflow pressure and centrifugal force, respectively. The formed droplets were polymerized by UV, while maintaining their spherical shape through cryogenic conditions using liquid nitrogen. The fabricated beads detected the target Silver ion (Ag)ions, resulting in contraction, with a maximum volume decrease of 20.3%. We believe that our DNA-aptamer incorporated hydrogel beads can be easily recognizable by the human eye if combined with color indicating gels in the future, and could be applied to small or wearable devices for environment monitoring and food management.

In this study, we propose an edible sensor to evaluate digestive function by gastric peristalsis. This sensor was fabricated by depositing a patterned edible gold layer to firm split-ring resonators on the surface of a vegetable sheet. Gastric peristalsis is estimated from the change in the strength of electromagnetic response of the edible sensor. In this experiment, we evaluated the wireless function of the edible sensor in the stomach-mimicking environment and the difference in the response strength of the sensor due to changes in the distance between the edible sensor and the antenna. This research is expected to contribute to the early detection of gastrointestinal diseases including stomach diseases.

This paper describes an in vitro parallel culture system driven by batch Bayesian optimization (BBO) algorithm to explore an optimal experimental condition for automatically efficient and quick tissue maturation. By our system, automatically generated new electric stimulation parameters to increase the level of maturation in parallel culture of myocyte tissues. We fabricated the electrical stimulation device and set three independent parameters (voltage, frequency, stimulus date) according to the BBO algorithm. After the cell cultivation under the electrical stimuli, the score of the maturation level was evaluated by image analysis of immunostaining and used for generating the next stimulation parameters by BBO. We believe that our system could discover the optimal condition for tissue maturation quickly and automatically, which is difficult for humans to discover through experiments.

Hydrogels have attracted attention as highly biocompatible supporting materials for wearable devices. An antenna is a key component for wireless transmission of information collected by the devices worn on the human body. Furthermore, integrating antennas into the devices eliminates the need for external wiring, which could obstruct the movement of the human body. In this study, we demonstrated the fabrication of planar conductive structures by laser-induced graphitization of tannic acid-containing alginate hydrogels and the fabrication of microstrip patch antennas (MPAs) using the fabricated structures. In a single laser scan onto the hydrogels, cracks were formed in the fabricated structures. For the fabrication of continuous planar structures, we used a method leveraging the water absorption property of hydrogels. By the method, in which an aqueous solution of tannic acid was added to the structures, followed by rescanning the laser beam to convert tannic acid to graphitic carbon, the number of cracks decreased. Additionally, it is confirmed that the fabricated structures can be applied to the radiation patches of hydrogel-based MPAs. A decrease in the resonant frequency of the fabricated MPAs corresponding to the decrease in the number of cracks is observed.

The patterning of conductive graphitic carbon planar structures on hydrogels was demonstrated by laser irradiation onto the hydrogels. Furthermore, the patterned conductive structure was applied to the conductive material of a microstrip patch antenna.

Mechanical stimuli have been recognized as important for tissue maturation, homeostasis and constructing engineered three-dimensional (3D) tissues. However, we know little about the cellular mechanical response in tissues that could be considerably heterogeneous and spatiotemporally dynamic due to the complex structure of tissues. Here, we report a spatiotemporal single-cell tracking analysis of in vitro 3D tissues under mechanical stretch, to reveal the heterogeneous cellular behavior by using a developed stretch and optical live imaging system. The system could affect the cellular orientation and directly measure the distance of cells in in vitro 3D myoblast tissues (3DMTs) at the single-cell level. Moreover, we observed the spatiotemporal heterogeneous cellular locomotion and shape changes under mechanical stretch in 3DMTs. This single-cell tracking analysis can become a principal method to investigate the heterogeneous cellular response in tissues and provide insights that conventional analyses have not yet offered.

Adeno-associated virus (AAV)-based gene therapy holds promise as a fundamental treatment for genetic disorders. For clinical applications, it is necessary to control AAV release timing to avoid an immune response to AAV. Here we propose an ultrasound (US)-triggered on-demand AAV release system using alginate hydrogel microbeads (AHMs) with a release enhancer. By using a centrifuge-based microdroplet shooting device, the AHMs encapsulating AAV with tungsten microparticles (W-MPs) are fabricated. Since W-MPs work as release enhancers, the AHMs have high sensitivity to the US with localized variation in acoustic impedance for improving the release of AAV. Furthermore, AHMs were coated with poly-l-lysine (PLL) to adjust the release of AAV. By applying US to the AAV encapsulating AHMs with W-MPs, the AAV was released on demand, and gene transfection to cells by AAV was confirmed without loss of AAV activity. This proposed US-triggered AAV release system expands methodological possibilities in gene therapy.

Thermoresponsive hydrogels, which exhibit changes in their optical properties and volume due to temperature variations, are promising candidates for applications in soft devices. In this study, we demonstrate the modulation of transmittance in a thermoresponsive hydrogel through light stimulation employing gold microstructures fabricated via multiphoton photoreduction. The spatial integration of photoresponsiveness, attributed to high-density gold nanoparticles within the thermoresponsive hydrogel, was accomplished through the high-speed laser scanning of femtosecond laser pulses. The temperature measurement during the fabrication of the gold microstructure revealed that the high-speed and multiple scanning over the same path effectively reduced the temperature in the irradiated area of femtosecond laser pulses. The present approach enabled the mitigation of thermal effects during the fabrication, resulting in minimizing distortion in the fine lines of the structures. Upon exposure to stimulus light, a rapid change in the transmittance of the region where the structures were fabricated was prominently observed. The present method unveils a promising avenue for the advancement of light-responsive soft devices.

Biological muscles mainly consist of motor proteins that play essential roles and have a hierarchical structure covered with collagen that can transmit the tension to bones via tendons. The artificial reconstruction of motor proteins is highly expected from the viewpoint of engineering and biology. Although artificial muscles have been developed via synthetic and biological molecular motors, there is a considerable gap between the components of artificial and biological muscles, resulting in a significant obstacle to further scale-up and force transfer through macroscale structures. To overcome these obstacles via bioderived biomolecules, this study proposes a collagen-actomyosin hybrid soft actuator inspired by the structure of biological muscles. Collagen gel entrapping actomyosin can play roles in maintaining the actuator's shape and transmitting the tensile force. The generated forces reach the micro-newton range, enabling the actuation of millimeter-scale mechanical components. These properties may be helpful for the fabrication of soft robotic systems with advanced functionalities.

Light-driven control of the flow velocity inside a hydrogel microchannel is demonstrated by fabricating metal microstructures by multiphoton photoreduction. A technique to evaluate the local strain and compression of the hydrogel will also be described.

Light-driven control of the flow velocity inside a hydrogel microchannel is demonstrated by fabricating metal microstructures by multiphoton photoreduction. A technique to evaluate the local strain and compression of the hydrogel will also be described.

Chitosan has various tissue regeneration effects. This study was designed to investigate the nerve regeneration effect of Schwann cell (SC)-encapsulated chitosan-collagen hydrogel nerve conduit (CCN) transplanted into a rat model of sciatic nerve defect. We prepared a CCN consisting of an outer layer of chitosan hydrogel and an inner layer of collagen hydrogel to encapsulate the intended cells. Rats with a 10-mm sciatic nerve defect were treated with SCs encapsulated in CCN (CCN+), CCN without SCs (CCN−), SC-encapsulated silicone tube (silicone+), and autologous nerve transplanting (auto). Behavioral and histological analyses indicated that motor functional recovery, axonal regrowth, and myelination of the CCN+ group were superior to those of the CCN− and silicone+ groups. Meanwhile, the CCN− and silicone+ groups showed no significant differences in the recovery of motor function and nerve histological restoration. In conclusion, SC-encapsulated CCN has a synergistic effect on peripheral nerve regeneration, especially axonal regrowth and remyelination of host SCs. In the early phase after transplantation, SC-encapsulated CCNs have a positive effect on recovery. Therefore, using SC-encapsulated CCNs may be a promising approach for massive peripheral nerve defects.

To advance the development of flexible materials for soft robotics applications, it is crucial to enhance the elastic modulus and breaking the stress of soft materials, such as hydrogels. Double network hydrogels (DN gels) have displayed remarkable mechanical strength owing to their unique network structure composed of two types of polymer networks. However, current fabrication methods for DN gels entail cross-linking two distinct hydrogel polymers throughout the entire hydrogel matrix. In this study, we focused on employing multi-photon polymerization (MPP) with femtosecond laser pulses as a cross-linking method for hydrogels to achieve spatially selective formation of DN gel structures at the micrometer scale, along with the consequent improvement in local mechanical strength. We assessed the mechanical properties of the fabricated structures and confirmed that the mechanical strength varied within the regions where DN gel was locally formed.

The light-stimulated control of flow velocity in the hydrogel microchannel was demonstrated by utilizing a metal microstructure fabricated by multiphoton photoreduction. The metal microstructure was fabricated adjacent to the microchannel in a poly-N- isopropylacrylamide (PNIPAm) hydrogel. Owing to the photothermal conversion at the metal microstructure, local deformation of the microchannel and change in flow velocity was induced as a result of the volume phase transition of the supporting hydrogel around the metal microstructure. Wavelength-selective change in the flow velocity was realized by utilizing dissimilar metal microstructures, that exhibit different optical resonances. The results indicate that multiphoton photoreduction is a promising method for fabricating novel hydrogel devices having flow-controllable switches by arranging light-absorbing structures with high spatial selectivity.

To generate three-dimensional tissue in vitro, promoting vasculogenesis in cell aggregates is an important factor. Here, we found that ultrasound promoted vasculogenesis of human umbilical vein endothelial cells (HUVECs). Promotion of HUVEC network formation and lumen formation were observed using our method. In addition to morphological evaluations, protein expression was quantified by western blot assays. As a result, expression of proteins related to vasculogenesis and the response to mechanical stress on cells was enhanced by exposure to ultrasound. Although several previous studies have shown that ultrasound may promote vasculogenesis, the effect of ultrasound was unclear because of unregulated ultrasound, the complex culture environment, or two-dimensional-cultured HUVECs that cannot form a lumen structure. In this study, regulated ultrasound was propagated on three-dimensional-monocultured HUVECs, which clarified the effect of ultrasound on vasculogenesis. We believe this finding may be an innovation in the tissue engineering field.

Animal pathology models play an important role in elucidating pathology, evaluating drug efficacy, and studying therapeutic. This paper presents the creation of rats’ renal failure model by embolizing the renal artery using radiopaque hydrogel microfibers. By using a dual coaxial laminar flow microfluidic device, barium alginate gel microfibers containing zirconia particles were fabricated. Since the zirconia particles are radiopaque, the microfiber can be delivered and embolized while confirming the position of the microfiber in real-time under X-ray imaging. The delivery of microfibers through a catheter into the renal artery of rats to create a renal failure model was tested. The radiopaque microfibers were visualized by X-ray imaging when delivered by a catheter. Furthermore, the microfibers remained stable for 2 weeks after delivery. In addition, by adjusting the number of microfibers, two different rat models with severe and mild renal failure conditions were created. The proposed delivery of radiopaque hydrogel microfibers to create an embolization model is expected to be an effective approach to control the degree of symptoms and elucidate the pathology.

This paper describes up-scalable microfiber-shaped tissues for macroscale tendon tissue reconstruction in vitro. C3H10T1/2 cells were encapsulated in a calcium alginate hydrogel microfiber that was fabricated via a double coaxial microfluidic device. The C3H10T1/2 cells gradually merged to construct the microfiber-shaped tendon-like tissue. Our microfiber-shaped tendon-like tissues were alive and maintained their microfiber-shaped morphology over 600 days. Immunostaining and real-time quantitative polymerase chain reaction analyses showed that our fabricated microfiber-shaped tendon-like tissue properly expressed tenomodulin and the orientation of the filaments of actin, which are one of the characteristics of tendon tissue in vivo. Furthermore, a macroscale tendon tissue assembly with ∼1 cm in length and ∼200 µm in thickness was successfully constructed by bundling the microfiber-shaped tendon-like tissues together. This feature enabled us to fabricate a macroscale tendon tissue with uniform cell distribution. We believe that our fabricated microfiber-shaped tendon-like tissue would be a suitable strategy to reconstruct tendon tissue in vitro for the treatments of tendon-related injuries.

Since the biochemical reaction of blood vessels plays an essential role in immune response or various diseases, in vitro vascular models have high demand from medical fields. However, vascular models often tend to be difficult to mimic the biomedical reaction faithfully because of the lack of implementation of the tissue deformation. Here, an inflammatory mediator-induced deformation reaction of a blood vessel on a flexibly deformable collagen hydrogel tube device is reproduced. A self-standing collagen tube enables the tissue to deform flexibly in biochemical reaction and achieves contraction both at tissue and cell level. The contraction of tissue relieves the stress between cells under reaction to maintain cellular junctions even tight junctions are broken. Also, the drug perfusion test with antihistamine chemical is easily performed due to the connector part and properly inhibits the inflammatory reaction. Moreover, the traction force on endothelial cells is analyzed as about 0.9 µN on two types of scaffolds with different stiffness. It is believed that the potential of the flexible tissue model to reproduce biochemical reactions can contribute to the fabrication of vascular tissue models mimicking in vivo in high similarity as a platform for biomedical researches and pharmacokinetic testing.

Programmable materials have artificially designed physical shapes responding to external stimuli, as well as high design capability and high flexibility. Here, we propose a microfiber-shaped programmable material with an axial pattern of stimuli-responsive (SR) and nonresponsive hydrogels. The SR pre-gel solution was mixed to sodium alginate pre-gel solution for instantaneous gelation with ionic crosslinking and solidified on a nonresponsive hydrogel microfiber with a valve-controlled microfluidic system. A design of microfiber-shaped programmable material (patterned position of SR regions) could be flexibly altered by changing a coded sequence program. We confirmed that the three-dimensional (3D) coil-like structures were self-folded at the patterned SR regions responding to the thermal stimulus and that the chirality of the self-folded 3D coil-like structures depends on the condition of the stimulus to the microfiber. Finally, interaction with objects using the programmable microfiber as a soft actuator was demonstrated. Our microfiber-shaped programmable materials expand possibilities of fiber-based materials in biomimetics and soft robotics fields.

This paper describes a microfluidic multicolor display utilizing juxtapositional color mixing of pixels. Our display has a 14 14 array of pixels (2.5 mm in pixel diameter, 8.46 ppi) on the display surface where multicolor is expressed by controlling the pattern of the four primary color inks (cyan (C), magenta (M), yellow (Y), and black (= key plate, K)) that fill the pixels. The microfluidic display has a three-layer structure composed of a top layer with pixels for displaying images, a middle layer that serves as a background screen, and a bottom layer with microchannels that connect the pixels. In order to express multicolor by combining CMYK primary colors, we optimized the concentration of the inks used as the primary colors. By designing patterns of pixels filled with CMYK ink, color gradations and multicolor images were displayed on our display. The proposed microfluidic display could be applied to eye-friendly and low-energy-consumption flexible display applications including multi-purpose sign boards used in outdoors, wearing objects, and exterior/interior of vehicles and architects.