Synergistic Effect of Electrical and Biochemical Stimulation on Human iPSC-Derived Neural Differentiation in a Microfluidic Electrode Array Chip.

Neural differentiation is crucial for advancing our understanding of the nervous system and developing treatments for neurological disorders. The advanced methods and the ability to manipulate the alignment, proliferation, and differentiation of stem cells are essential for studying neuronal development and synaptic interactions. However, the utilization of human induced pluripotent stem cells (iPSCs) for disease modeling of neurodegenerative conditions may be constrained by the prolonged duration and uncontrolled cell differentiation required for functional neural cell differentiation. Here, we developed a microfluidic chip to enhance the differentiation and maturation of specific neural lineages by placing aligned microelectrodes on the glass surface to regulate the neural differentiation of human iPSCs. The utilization of electrical stimulation (ES) in conjunction with neurotrophic factors (NF) significantly enhanced the efficiency in generating functional neurons from human iPSCs. We also observed that the simultaneous application of NF and ES to human iPSCs promoted their differentiation and maturation into functional neurons while increasing synaptic interactions. Our research demonstrated the effect of combining NF and ES on human iPSC-derived neural differentiation.

Effect of gut microbiota-derived metabolites and extracellular vesicles on neurodegenerative disease in a gut-brain axis chip.

A new perspective suggests that a dynamic bidirectional communication system, often referred to as the microbiome-gut-brain axis, exists among the gut, its microbiome, and the central nervous system (CNS). This system may influence brain health and various brain-related diseases, especially in the realms of neurodevelopmental and neurodegenerative conditions. However, the exact mechanism is not yet understood. Metabolites or extracellular vesicles derived from microbes in the gut have the capacity to traverse the intestinal epithelial barrier or blood-brain barrier, gaining access to the systemic circulation. This phenomenon can initiate the physiological responses that directly or indirectly impact the CNS and its function. However, reliable and controllable tools are required to demonstrate the causal effects of gut microbial-derived substances on neurogenesis and neurodegenerative diseases. The integration of microfluidics enhances scientific research by providing advanced in vitro engineering models. In this study, we investigated the impact of microbe-derived metabolites and exosomes on neurodevelopment and neurodegenerative disorders using human induced pluripotent stem cells (iPSCs)-derived neurons in a gut-brain axis chip. While strain-specific, our findings indicate that both microbial-derived metabolites and exosomes exert the significant effects on neural growth, maturation, and synaptic plasticity. Therefore, our results suggest that metabolites and exosomes derived from microbes hold promise as potential candidates and strategies for addressing neurodevelopmental and neurodegenerative disorders.

Electro-Responsive Conductive Blended Hydrogel Patch.

The proposed electro-responsive hydrogel has great benefit for transdermal drug delivery system (TDDS) applications. To improve the physical or chemical properties of hydrogels, a number of researchers have previously studied the mixing efficiencies of the blended hydrogels. However, few studies have focused on improving the electrical conductivity and drug delivery of the hydrogels. We developed a conductive blended hydrogel by mixing alginate with gelatin methacrylate (GelMA) and silver nanowire (AgNW). We demonstrated that and the tensile strength of blended hydrogels were increased by a factor of 1.8 by blending GelMA and the electrical conductivity was enhanced by a factor of 18 by the addition of AgNW. Furthermore, the GelMA-alginate-AgNW (Gel-Alg-AgNW) blended hydrogel patch enabled on-off controllable drug release, indicating 57% doxorubicin release in response to electrical stimulation (ES) application. Therefore, this electro-responsive blended hydrogel patch could be useful for smart drug delivery applications.

Effect of gut microbiome-derived metabolites and extracellular vesicles on hepatocyte functions in a gut-liver axis chip.

Metabolism, is a complex process involving the gut and the liver tissue, is difficult to be reproduced in vitro with conventional single cell culture systems. To tackle this challenge, we developed a gut-liver-axis chip consisting of the gut epithelial cell chamber and three-dimensional (3D) uniform-sized liver spheroid chamber. Two cell culture chamber compartments were separated with a porous membrane to prevent microorganisms from passing through the chamber. When the hepG2 spheroids cultured with microbiota-derived metabolites, we observed the changes in the physiological function of hepG2 spheroids, showing that the albumin and urea secretion activity of liver spheroids was significantly enhanced. Additionally, the functional validation of hepG2 spheroids treated with microbiota-derived exosome was evaluated that the treatment of the microbiota-derived exosome significantly enhanced albumin and urea in hepG2 spheroids in a gut-liver axis chip. Therefore, this gut-liver axis chip could be a potentially powerful co-culture platform to study the interaction of microbiota and host cells.

Microfluidic electrode array chip for electrical stimulation-mediated axonal regeneration.

The precise manipulation of the neural stem cell (NSC)-derived neural differentiation is still challenging, and there is a technological barrier to regulate the axonal regeneration in a controlled manner. Here, we developed a microfluidic chip integrated with a microelectrode array as an axonal guidance platform. The microfluidic electrode array chip consisted of two compartments and a bridge microchannel that could isolate and guide the axons. We demonstrated that the NSCs were largely differentiated into neural cells as the electric field was applied to the microfluidic electrode array chip. We also confirmed the synergistic effects of the electrical stimulation (ES) and neurotrophic factor (NF) on axonal outgrowth. This microfluidic electrode array chip can serve as a central nervous system (CNS) model for axonal injury and regeneration. Therefore, it could be a potentially powerful tool for an in vitro model of the axonal regeneration.

Contributions of the microbiome to intestinal inflammation in a gut-on-a-chip.

The intestinal microbiome affects a number of biological functions of the organism. Although the animal model is a powerful tool to study the relationship between the host and microbe, a physiologically relevant in vitro human intestinal system has still unmet needs. Thus, the establishment of an in vitro living cell-based system of the intestine that can mimic the mechanical, structural, absorptive, transport and pathophysiological properties of the human intestinal environment along with its commensal bacterial strains can promote pharmaceutical development and potentially replace animal testing. In this paper, we present a microfluidic-based gut model which allows co-culture of human and microbial cells to mimic the gastrointestinal structure. The gut microenvironment is recreated by flowing fluid at a low rate (21 µL/h) over the microchannels. Under these conditions, we demonstrated the capability of gut-on-a-chip to recapitulate in vivo relevance epithelial cell differentiation including highly polarized epithelium, mucus secretion, and tight membrane integrity. Additionally, we observed that the co-culture of damaged epithelial layer with the probiotics resulted in a substantial responded recovery of barrier function without bacterial overgrowth in a gut-on-a-chip. Therefore, this gut-on-a-chip could promote explorations interaction with host between microbe and provide the insights into questions of fundamental research linking the intestinal microbiome to human health and disease.

Conductive GelMA-Collagen-AgNW Blended Hydrogel for Smart Actuator.

Blended hydrogels play an important role in enhancing the properties (e.g., mechanical properties and conductivity) of hydrogels. In this study, we generated a conductive blended hydrogel, which was achieved by mixing gelatin methacrylate (GelMA) with collagen, and silver nanowire (AgNW). The ratio of GelMA, collagen and AgNW was optimized and was subsequently gelated by ultraviolet light (UV) and heat. The scanning electron microscope (SEM) image of the conductive blended hydrogels showed that collagen and AgNW were present in the GelMA hydrogel. Additionally, rheological analysis indicated that the mechanical properties of the conductive GelMA-collagen-AgNW blended hydrogels improved. Biocompatibility analysis confirmed that the human umbilical vein endothelial cells (HUVECs) encapsulated within the three-dimensional (3D), conductive blended hydrogels were highly viable. Furthermore, we confirmed that the molecule in the conductive blended hydrogel was released by electrical stimuli-mediated structural deformation. Therefore, this conductive GelMA-collagen-AgNW blended hydrogel could be potentially used as a smart actuator for drug delivery applications.

Electro-responsive hydrogel-based microfluidic actuator platform for photothermal therapy.

Electrical stimuli play an important role in regulating the delivery of plasmonic nanomaterials with cancer targeting peptides. Here, we developed an electro-responsive hydrogel-based microfluidic actuator platform for brain tumor targeting and photothermal therapy (PTT) applications. The electro-responsive hydrogels consisted of highly conductive silver nanowires (AgNWs) and biocompatible collagen I gels. We confirmed that an electrically conductive hydrogel could be used as an effective actuator by applying an electrical signal in the microfluidic platform. Furthermore, we successfully demonstrated PTT efficacy for brain tumor cells using targetable Arg-Gly-Asp (RGD) peptide-conjugated gold nanorods (GNRs). Therefore, our electro-responsive hydrogel-based microfluidic actuator platform could be useful for electro-responsive intelligent nanomaterial delivery and PTT applications.

Production of Flavonoids in Callus Cultures of Sophora flavescens Aiton.

Flavonoids, including maackiain (Maac) from Sophora flavescens Aiton roots, have many pharmacological properties, such as antitumor, antimicrobial, and antifungal activities. This research aimed to develop an in vitro plant and callus culture system for S. flavescens for the purpose of generating an alternative production system for enhancing Maac production, as Maac is usually present in very small amounts in S. flavescens' roots. We arranged the optimal conditions of different tissues of S. flavescens and supplemented the medium with various plant growth regulators (PGRs). The highest induction and proliferation rates of callus was shown in combination treatments of all concentrations of thidiazuron (TDZ) and picloram. In addition, calli induced with leaf explants cultured on 2.0 mg/L picloram and 0.5 mg/L 6-benzyladenine (BA) in Murashige and Skoog (MS) medium had the highest accumulation of the active metabolite Maac. In vitro shoots were regenerated on medium containing combinations of TDZ and α-Naphthalene acetic acid (NAA). A reliable protocol for the mass production of secondary metabolites using a callus culture of S. flavescens was successfully established.

rGO nanomaterial-mediated cancer targeting and photothermal therapy in a microfluidic co-culture platform.

We developed the microfluidic co-culture platform to study photothermal therapy applications. We conjugated folic acid (FA) to target breast cancer cells using reduced graphene oxide (rGO)-based functional nanomaterials. To characterize the structure of rGO-based nanomaterials, we analyzed the molecular spectrum using UV-visible and Fourier-transform infrared spectroscopy (FT-IR). We demonstrated the effect of rGO-FA-based nanomaterials on photothermal therapy of breast cancer cells in the microfluidic co-culture platform. From the microfluidic co-culture platform with breast cancer cells and human umbilical vein endothelial cells (HUVECs), we observed that the viability of breast cancer cells treated with rGO-FA-based functional nanomaterials was significantly decreased after near-infrared (NIR) laser irradiation. Therefore, this microfluidic co-culture platform could be a potentially powerful tool for studying cancer cell targeting and photothermal therapy.

Generation of tumor spheroids using a droplet-based microfluidic device for photothermal therapy.

Despite their simplicity, monolayer cell cultures are not able to accurately predict drug behavior in vivo due to their inability to accurately mimic cell-cell and cell-matrix interactions. In contrast, cell spheroids are able to reproduce these interactions and thus would be a viable tool for testing drug behavior. However, the generation of homogenous and reproducible cell spheroids on a large scale is a labor intensive and slow process compared to monolayer cell cultures. Here, we present a droplet-based microfluidic device for the automated, large-scale generation of homogenous cell spheroids in a uniform manner. Using the microfluidic system, the size of the spheroids can be tuned to between 100 and 130 µm with generation frequencies of 70 Hz. We demonstrated the photothermal therapy (PTT) application of brain tumor spheroids generated by the microfluidic device using a reduced graphene oxide-branched polyethyleneimine-polyethylene glycol (rGO-BPEI-PEG) nanocomposite as the PTT agent. Furthermore, we generated uniformly sized neural stem cell (NSC)-derived neurospheres in the droplet-based microfluidic device. We also confirmed that the neurites were regulated by neurotoxins. Therefore, this droplet-based microfluidic device could be a powerful tool for photothermal therapy and drug screening applications.

Dual-nozzle microfluidic droplet generator.

The droplet-generating microfluidics has become an important technique for a variety of applications ranging from single cell analysis to nanoparticle synthesis. Although there are a large number of methods for generating and experimenting with droplets on microfluidic devices, the dispensing of droplets from these microfluidic devices is a challenge due to aggregation and merging of droplets at the interface of microfluidic devices. Here, we present a microfluidic dual-nozzle device for the generation and dispensing of uniform-sized droplets. The first nozzle of the microfluidic device is used for the generation of the droplets, while the second nozzle can accelerate the droplets and increase the spacing between them, allowing for facile dispensing of droplets. Computational fluid dynamic simulations were conducted to optimize the design parameters of the microfluidic device.

Prediction analysis and quality assessment of microwell array images.

Microwell arrays are widely used for the analysis of fluorescent-labelled biomaterials. For rapid detection and automated analysis of microwell arrays, the computational image analysis is required. Support Vector Machines (SVM) can be used for this task. Here, we present a SVM-based approach for the analysis of microwell arrays consisting of three distinct steps: labeling, training for feature selection, and classification into three classes. The three classes are filled, partially filled, and unfilled microwells. Next, the partially filled wells are analyzed by SVM and their tendency towards filled or unfilled tested through applying a Gaussian filter. Through this, all microwells can be categorized as either filled or unfilled by our algorithm. Therefore, this SVM-based computational image analysis allows for an accurate and simple classification of microwell arrays.

Nicotiana benthamiana Matrix Metalloprotease 1 (NMMP1) gene confers disease resistance to Phytophthora infestans in tobacco and potato plants.

We previously isolated Nicotiana benthamiana matrix metalloprotease 1 (NMMP1) from tobacco leaves. The NMMP1 gene encodes a highly conserved, Zn-containing catalytic protease domain that functions as a factor in the plant's defense against bacterial pathogens. Expression of NMMP1 was strongly induced during interactions between tobacco and one of its pathogens, Phytophthora infestans. To elucidate the role of the NMMP1 in defense of N. benthamiana against fungal pathogens, we performed gain-of-function and loss-of-function studies. NMMP1-overexpressing plants had stronger resistance responses against P. infestans infections than control plants, while silencing of NMMP1 resulted in greater susceptibility of the plants to the pathogen. This greater susceptibility correlated with fewer NMMP1 transcripts than the non-silenced control. We also examined cell death as a measure of disease. The amount of cell death induced by the necrosis-inducing P. infestans protein 1, PiNPP1, was dependent on NMMP1 in N. benthamiana. Potato plants overexpressing NMMP1 also had enhanced disease resistance against P. infestans. RT-PCR analysis of these transgenic potato plants revealed constitutive up-regulation of the potato defense gene NbPR5. NMMP1-overexpressing potato plants were taller and produced heavier tubers than control plants. We suggest a role for NMMP1in pathogen defense and development.

Recombinant Human Acidic Fibroblast Growth Factor (aFGF) Expressed in Nicotiana benthamiana Potentially Inhibits Skin Photoaging.

Responding to the need for recombinant acidic fibroblast growth factor in the pharmaceutical and cosmetic industries, we established a scalable expression system for recombinant human aFGF using transient and a DNA replicon vector expression in Nicotiana benthamiana. Recombinant human-acidic fibroblast growth factor was recovered following Agrobacterium infiltration of N. benthamiana. The optimal time point at which to harvest recombinant human acidic fibroblast growth factor expressing leaves was found to be 4 days post-infiltration, before necrosis was evident. Commassie-stained SDS-PAGE gels of His-tag column eluates, concentrated using a 10 000 molecular weight cut-off column, showed an intense band at the expected molecular weight for recombinant human acidic fibroblast growth factor. An immunoblot confirmed that this band was recombinant human acidic fibroblast growth factor. Up to 10 µg recombinant human-acidic fibroblast growth factor/g of fresh leaves were achieved by a simple affinity purification protocol using protein extract from the leaves of agroinfiltrated N. benthamiana. The purified recombinant human acidic fibroblast growth factor improved the survival rate of UVB-irradiated HaCaT and CCD-986sk cells approximately 89 and 81 %, respectively. N. benthamiana-derived recombinant human acidic fibroblast growth factor showed similar effects on skin cell proliferation and UVB protection compared to those of Escherichia coli-derived recombinant human acidic fibroblast growth factor. Additionally, N. benthamiana-derived recombinant human acidic fibroblast growth factor increased type 1 procollagen synthesis up to 30 % as well as reduced UVB-induced intracellular reactive oxygen species generation in fibroblast (CCD-986sk) cells.UVB is a well-known factor that causes various types of skin damage and premature aging. Therefore, the present study demonstrated that N. benthamiana-derived recombinant human acidic fibroblast growth factor effectively protects skin cell from UVB, suggesting its potential use as a cosmetic or therapeutic agent against skin photoaging.

Analysis of 3D multi-layer microfluidic gradient generator.

We developed a three-dimensional (3D) simple multi-layer microfluidic gradient generator to create molecular gradients on the centimeter scale with a wide range of flow rates. To create the concentration gradients, a main channel (MC) was orthogonally intersected with vertical side microchannel (SC) in a 3D multi-layer microfluidic device. Through sequential dilution from the SC, a spatial gradient was generated in the MC. Two theoretical models were created to assist in the design of the 3D multi-layer microfluidic gradient generator and to compare its performance against a two-dimensional equivalent. A first mass balance model was used to predict the steady-state concentrations reached, while a second computational fluid dynamic model was employed to predict spatial development of the gradient by considering convective as well as diffusive mass transport. Furthermore, the theoretical simulations were verified through experiments to create molecular gradients in a 3D multi-layer microfluidic gradient generator.

Alpha-ray spectrometry at high temperature by using a compound semiconductor detector.

The use of conventional radiation detectors in harsh environments is limited by radiation damage to detector materials and by temperature constraints. We fabricated a wide-band gap semiconductor radiation detector based on silicon carbide. All the detector components were considered for an application in a high temperature environment like a nuclear reactor core. The radiation response, especially to alpha particles, was measured using an (241)Am source at variable operating voltages at room temperature in the air. The temperature on detector was controlled from 30°C to 250°C. The alpha-particle spectra were measured at zero bias operation. Even though the detector is operated at high temperature, the energy resolution as a function of temperature is almost constant within 3.5% deviation.

A self-biased neutron detector based on an SiC semiconductor for a harsh environment.

Neutron detector based on radiation-hard semiconductor materials like SiC, diamond and AlN has recently emerged as an attractive device for an in-core reactor neutron flux monitoring, a spent fuel characterization, and a home land security application. For the purpose of field measurement activity, a radiation detector having a low-power consumption, a mechanical stability and a radiation hardness is required. Our research was focused on the development of a radiation-resistive neutron semiconductor detector based on a wide band-gap SiC semiconductor. And also it will be operated at a zero-biased voltage using a strong internal electric field. The charge collection efficiency (CCE) was over 80% when the biased voltage was zero. When the biased voltage was applied above 20V, the charge collection efficiency reached 100%.

Development of a beta gauge system for a fabric density measurement.

A beta gauge system was designed and fabricated for a fabric density measurement with high sensitivity in real time. An ionization chamber and 3.7GBq (85)Kr was chosen as a beta radiation sensor and a beta source. An ionization chamber, which had a 0.5microm-thick aluminized mylar film as an incident window, was constructed with a guard electrode structure in consideration of minimizing leakage currents. Xe gas was chosen as a filling gas to increase sensitivity of an ionization chamber. Linearity of the ionization currents against the densities was 0.9894 in the selected measurement range. And the deviation of the ionization currents at a standard density was below 0.02%. The ranges of the density measurement and the active area were from 20 to 999g/cm(2) with +/-1% accuracy and 40mm diameters. The fabricated beta gauge system can be applied to fields such as in the paper, film, and plastic production industries.

New concept of CZT-based hand-held radioisotope identifier.

Radiation dose can be determined with the radiation dosimeter. Recently, hand-held radioisotope identifier is being developed by many researchers to measure the radiation dose and to determine the type of the radioisotope. New design concept of the radioisotope identifier is introduced in this work. In this design, one can measure the radiation dose and the type of the radioisotope, and one can also determine the direction where the radiation comes. The hand-held radioisotope identifier is based on cadmium zinc telluride (CZT) detector. The detection principle is from the change of the energy spectrum as the direction of the incident radiation is varied. EGSnrc Monte Carlo simulation is done to calculate the energy spectrum of the CZT detector, and the result is compared with the measured one. The energy spectra are calculated when the gamma-ray is incident on the detection at different directions. The simulation results show the change of the spectrum shape as the incident radiation comes from different directions. The calculation is also done with different radiation energies and detector sizes. A prototype radioisotope identifier is being developed at Korea Atomic Energy Research Institute (KAERI). This work would be helpful to develop next generation radiation dosimeter.