A Microphysiological Model to Mimic the Placental Remodeling during Early Stage of Pregnancy under Hypoxia-Induced Trophoblast Invasion.

Placental trophoblast invasion is critical for establishing the maternal-fetal interface, yet the mechanisms driving trophoblast-induced maternal arterial remodeling remain elusive. To address this gap, we developed a three-dimensional microfluidic placenta-on-chip model that mimics early pregnancy placentation in a hypoxic environment. By studying human umbilical vein endothelial cells (HUVECs) under oxygen-deprived conditions upon trophoblast invasion, we observed significant HUVEC artery remodeling, suggesting the critical role of hypoxia in placentation. In particular, we found that trophoblasts secrete matrix metalloproteinase (MMP) proteins under hypoxic conditions, which contribute to arterial remodeling by the degradation of extracellular matrix components. This MMP-mediated remodeling is critical for facilitating trophoblast invasion and proper establishment of the maternal-fetal interface. In addition, our platform allows real-time monitoring of HUVEC vessel contraction during trophoblast interaction, providing valuable insights into the dynamic interplay between trophoblasts and maternal vasculature. Collectively, our findings highlight the importance of MMP-mediated arterial remodeling in placental development and underscore the potential of our platform to study pregnancy-related complications and evaluate therapeutic interventions.

3D Artificial Skin Platform for Investigating Pregnancy-Related Skin Pigmentation.

In this study, we created a 3D Artificial Skin Platform that can be used for the treatment of pigmentation by artificially realizing the skin of pregnant women. For the stable realization of 3D artificial skin, a bilayer hydrogel composed of collagen type I and fibrin was designed and applied to the study to reduce the tension-induced contraction of collagen type I, the extracellular matrix (ECM) of artificial skin, by dynamic culture. Oxygen concentration and 17ß-Estradiol (E2) concentration, which are highly related to melanin production, were selected as parameters of the pregnancy environment and applied to cell culture. Oxygen concentration, which is locally reduced in the first trimester (2.5-3%), and E2, which is upregulated in the third trimester, were applied to the cell culture process. We analyzed whether the 3D artificial skin implemented in the 3D Artificial Skin Platform could better represent the tendency of melanin expression in pregnant women than cells cultured under the same conditions in 2D. The expression levels of melanin and melanin-related genes in the 2D cell culture did not show a significant trend that was similar to the melanin expression trend in pregnant women. However, the 3D artificial skin platform showed a significant trend towards a 2-6-fold increase in melanin expression in response to low oxygen concentrations (2.5%) and E2 concentrations (17 ng/mL), which was similar to the trend in pregnant women in vivo. These results suggest that 3D artificial skin cultured on the Artificial Skin Platform has the potential to be used as a substitute for human pregnant skin in various research fields related to the treatment of pigmentation.

Microfluidics in High-Throughput Drug Screening: Organ-on-a-Chip and C. elegans-Based Innovations.

The development of therapeutic interventions for diseases necessitates a crucial step known as drug screening, wherein potential substances with medicinal properties are rigorously evaluated. This process has undergone a transformative evolution, driven by the imperative need for more efficient, rapid, and high-throughput screening platforms. Among these, microfluidic systems have emerged as the epitome of efficiency, enabling the screening of drug candidates with unprecedented speed and minimal sample consumption. This review paper explores the cutting-edge landscape of microfluidic-based drug screening platforms, with a specific emphasis on two pioneering approaches: organ-on-a-chip and C. elegans-based chips. Organ-on-a-chip technology harnesses human-derived cells to recreate the physiological functions of human organs, offering an invaluable tool for assessing drug efficacy and toxicity. In parallel, C. elegans-based chips, boasting up to 60% genetic homology with humans and a remarkable affinity for microfluidic systems, have proven to be robust models for drug screening. Our comprehensive review endeavors to provide readers with a profound understanding of the fundamental principles, advantages, and challenges associated with these innovative drug screening platforms. We delve into the latest breakthroughs and practical applications in this burgeoning field, illuminating the pivotal role these platforms play in expediting drug discovery and development. Furthermore, we engage in a forward-looking discussion to delineate the future directions and untapped potential inherent in these transformative technologies. Through this review, we aim to contribute to the collective knowledge base in the realm of drug screening, providing valuable insights to researchers, clinicians, and stakeholders alike. We invite readers to embark on a journey into the realm of microfluidic-based drug screening platforms, fostering a deeper appreciation for their significance and promising avenues yet to be explored.

Portable Colorimetric Hydrogel Beads for Point-of-Care Antimicrobial Susceptibility Testing.

Sepsis is a life-threatening condition with systemic inflammatory responses caused by bacterial infections. Considering the emergence of antibiotic-resistant bacteria such as methicillin-resistant Staphylococcus aureus (MRSA), sepsis is a great threat to public health. The gold standard methods for antimicrobial susceptibility testing (AST), however, take at least approximately 3 days to implement the entire blood culture, pure culture, and AST processes. To overcome the time-consuming nature of conventional AST, a method employing a chromatic biosensor composed of poly(diacetylene), alginate, and LB broth (PAL) is introduced in this study. Compared to the gold standards, AST with PAL biosensors can be completed within a time frame as short as 16 h. Such a significant reduction in time is possible because the consecutive cultures and AST are carried out simultaneously by encapsulating the bacterial nutrients and detection molecules into a single component. The bead-like hydrogel sensors were used in their freeze-dried form, which endows them with portability and stability, thus making them adequate for point-of-care testing. The PAL biosensor yields minimum inhibitory concentrations comparable to those from the Clinical and Laboratory Standards Institute, and the applicability of the biosensor is further shown in MRSA-infected mice.

Elucidating the Molecular Interactions between Lipids and Lysozyme: Evaporation Resistance and Bacterial Barriers for Dry Eye Disease.

Dry eye disease (DED) is a chronic condition characterized by ocular dryness and inflammation. The tear film lipid layer (TFLL) is the outermost layer composed of lipids and proteins that protect the ocular surface. However, environmental contaminants can disrupt its structure, potentially leading to DED. Although the importance of tear proteins in the TFLL functionality has been clinically recognized, the molecular mechanisms underlying TFLL-protein interactions remain unclear. In this study, we investigated tear protein-lipid interactions and analyzed their role in the TFLL functionality. The results show that lysozyme (LYZ) increases the stability of the TFLL by reducing its surface tension and increasing its surface pressure, resulting in increased TFLL evaporation and bacterial invasion resistance, with improved wettability and lubrication performance. These findings highlight the critical role of LYZ in maintaining ocular health and provide potential avenues for investigating novel approaches to DED treatment and patient well-being.

Multicomponent-Loaded Vesosomal Drug Carrier for Controlled and Sustained Compound Release.

Liposomes have been extensively adopted in drug delivery systems with clinically approved formulations. However, hurdles remain in terms of loading multiple components and precisely controlling their release. Herein, we report a vesosomal carrier composed of liposomes encapsulated inside the core of another liposome for the controlled and sustained release of multiple contents. The inner liposomes are made of lipids with different compositions and are co-encapsulated with a photosensitizer. Upon induction of reactive oxygen species (ROS), the contents of the liposomes are released, with each type of liposome displaying distinct kinetics due to the variance in lipid peroxidation for differential structural deformation. In vitro experiments demonstrated immediate content release from ROS-vulnerable liposomes, followed by sustained release from ROS-nonvulnerable liposomes. Moreover, the release trigger was validated at the organismal level using Caenorhabditis elegans. This study demonstrates a promising platform for more precisely controlling the release of multiple components.

Trophoblast Migration with Different Oxygen Levels in a Gel-Patterned Microfluidic System.

In the placenta, substances such as nutrients, oxygen, and by-products are exchanged between the mother and the fetus, and the proper formation of the placenta determines the success of pregnancy, including the growth of the fetus. Preeclampsia is an obstetric disease in which the incomplete formation of the placenta occurs, which is known to occur when there is an abnormality in the invasion of trophoblast cells. The invasion of trophoblast cells is controlled by oxygen concentration, and HIF-1α changes according to oxygen concentration, showing a difference in cell mobility. MMP-2 and MMP-9 are observed to be high in the endometrium involved in trophoblast invasion, and the expression is regulated according to the oxygen concentration. In this experiment, cell culture was conducted using a gel-patterned system with a hypoxic chamber. Before the chip experiment, the difference in the expression of MMP-2 and MMP-9 according to the oxygen concentration was confirmed using a hypoxia chamber. After that, trophoblast cells (HTR8/SVneo) and endothelial cells (HUVECs) were separated and cultured through a physical barrier through a hydrogel on a microfluidic chip. Cells were cultured in a hypoxic chamber under controlled oxygen levels. It was confirmed that the mobility of trophoblast cells in culture on the chip was upregulated in a hypoxic environment through oxygen control. This suggests that the formation of a hypoxic environment in the endometrium where the invasion of trophoblast cells occurs plays a role in increasing cell mobility.

Highly Specific Peptide-Mediated Cuvette-Form Localized Surface Plasmon Resonance (LSPR)-Based Fipronil Detection in Egg.

Herein, we have developed peptide-coated gold nanoparticles (AuNPs) based on localized surface plasmon resonance (LSPR) sensor chips that can detect fipronil with high sensitivity and selectivity. The phage display technique has been exploited for the screening of highly specific fipronil-binding peptides for the selective detection of the molecule. LSPR sensor chips are fabricated initially by attaching uniformly synthesized AuNPs on the glass substrate, followed by the addition of screened peptides. The parameters, such as the peptide concentration of 20 µg mL-1 and the reaction time of 30 min, are further optimized to maximize the efficacy of the fabricated LSPR sensor chips. The sensing analysis is performed systematically under standard fipronil solutions and spike samples from eggs. The developed sensor has shown excellent sensitivity towards both standard solutions and spike samples with limit of detection (LOD) values of 0.01 ppb, respectively. Significantly, the developed LSPR sensor chips offer distinct features, such as a facile fabrication approach, on-site sensing, rapid analysis, cost-effectiveness, and the possibility of mass production, in which the chips can be effectively used as a promising and potential on-site detection tool for the estimation of fipronil.

Multi-Modal Locomotion of Caenorhabditis elegans by Magnetic Reconfiguration of 3D Microtopography.

Miniaturized untethered soft robots are recently exploited to imitate multi-modal curvilinear locomotion of living creatures that perceive change of surrounding environments. Herein, the use of Caenorhabditis elegans (C. elegans) is proposed as a microscale model capable of curvilinear locomotion with mechanosensing, controlled by magnetically reconfigured 3D microtopography. Static entropic microbarriers prevent C. elegans from randomly swimming with the omega turns and provide linear translational locomotion with velocity of ≈0.14 BL s-1 . This velocity varies from ≈0.09 (for circumventing movement) to ≈0.46 (for climbing) BL s-1 , depending on magnetic bending and twisting actuation coupled with assembly of microbarriers. Furthermore, different types of neuronal mutants prevent C. elegans from implementing certain locomotion modes, indicating the potential for investigating the correlation between neurons and mechanosensing functions. This strategy promotes a platform for the contactless manipulation of miniaturized biobots and initiates interdisciplinary research for investigating sensory neurons and human diseases.

Elucidation of the Interactions of Reactive Oxygen Species and Antioxidants in Model Membranes Mimicking Cancer Cells and Normal Cells.

Photosensitizers (PSs) used in photodynamic therapy (PDT) have been developed to selectively destroy tumor cells. However, PSs recurrently reside on the extracellular matrix or affect normal cells in the vicinity, causing side effects. Additionally, the membrane stability of tumor cells and normal cells in the presence of reactive oxygen species (ROS) has not been studied, and the effects of ROS at the membrane level are unclear. In this work, we elucidate the stabilities of model membranes mimicking tumor cells and normal cells in the presence of ROS. The model membranes are constructed according to the degree of saturation in lipids and the bilayers are prepared either in symmetric or asymmetric form. Interestingly, membranes mimicking normal cells are the most vulnerable to ROS, while membranes mimicking tumor cells remain relatively stable. The instability of normal cell membranes may be one cause of the side effects of PDT. Moreover, we also show that ROS levels are controlled by antioxidants, helping to maintain an appropriate amount of ROS when PDT is applied.

Simple and rapid detection of ractopamine in pork with comparison of LSPR and LFIA sensors.

This study developed a simple and rapid strategic technique to detect ractopamine (chemical growth-promoting agent) in pork. Two highly sensitive and specific gold nanoparticle-based portable sensors, i.e., localized surface plasmon resonance (LSPR) sensors, and lateral flow immunoassay (LFIA) strips were developed to detect veterinary drug residues in food products, that have detrimental effects on humans. Optimization studies were conducted on several sensor devices to improve sensitivity. Each sensor comprised functionalized gold nanoparticles conjugated with ractopamine antibodies. The LSPR sensor chip achieved excellent detection sensitivity = 1.19 fg/mL and was advantageous for quantitative analysis due to its wide dynamic range. On the other hand, LFIA strips provided visual test confirmation and achieved 2.27 ng/mL detection sensitivity, significantly less sensitive than LSPR. The complementary sensors help overcome each other's shortcomings with both the techniques offering ease of use, affordability and rapid diagnosis. Thus, these sensors can be applied on-site for routine screening of harmful drug residues in pork meat. They also provide useful direction for advanced technologies to enhance assay performance for detecting various other food drug contaminants.

A simple strategy for signal enhancement in lateral flow assays using superabsorbent polymers.

To enhance the sensitivity of lateral flow assays (LFAs), a simple strategy is proposed using a nitrocellulose membrane modified with a superabsorbent polymer (SAP). SAP was incorporated into a nitrocellulose membrane for the flow control of detection probes. When absorbing aqueous solutions, SAP promoted the formation of biomolecule complexes to achieve up to a tenfold sensitivity improvement for the detection of human IgG. The assay time was optimized experimentally and numerically to within 20 min using this strategy. Moreover, fluid saturation in LFAs modified with SAP was mathematically simulated to better understand the underlying process, and molecular dynamics simulations were carried out to determine the effect of SAP. The proposed design was also applied to samples spiked with human immunoglobulin-depleted serum to test its applicability. The strategy presented is unique in that it preserves the characteristics of conventional LFAs, as it minimizes user intervention and is simple to manufacture at scale.

Quantitative Analysis of the Membrane Affinity of Local Anesthetics Using a Model Cell Membrane.

Local anesthesia is a drug that penetrates the nerve cell membrane and binds to the voltage gate sodium channel, inhibiting the membrane potential and neurotransmission. It is mainly used in clinical uses to address the pain of surgical procedures in the local area. Local anesthetics (LAs), however, can be incorporated into the membrane, reducing the thermal stability of the membrane as well as altering membrane properties such as fluidity, permeability, and lipid packing order. The effects of LAs on the membrane are not yet fully understood, despite a number of previous studies. In particular, it is necessary to analyze which is the more dominant factor, the membrane affinity or the structural perturbation of the membrane. To analyze the effects of LAs on the cell membrane and compare the results with those from model membranes, morphological analysis and 50% inhibitory concentration (IC50) measurement of CCD-1064sk (fibroblast, human skin) membranes were carried out for lidocaine (LDC) and tetracaine (TTC), the most popular LAs in clinical use. Furthermore, the membrane affinity of the LAs was quantitatively analyzed using a colorimetric polydiacetylene assay, where the color shift represents their distribution in the membrane. Further, to confirm the membrane affinity and structural effects of the membranes, we performed an electrophysiological study using a model protein (gramicidin A, gA) and measured the channel lifetime of the model protein on the free-standing lipid bilayer according to the concentration of each LA. Our results show that when LAs interact with cell membranes, membrane affinity is a more dominant factor than steric or conformational effects of the membrane.

Surface-directed engineering of tissue anisotropy in microphysiological models of musculoskeletal tissue.

Here, we present an approach to model and adapt the mechanical regulation of morphogenesis that uses contractile cells as sculptors of engineered tissue anisotropy in vitro. Our method uses heterobifunctional cross-linkers to create mechanical boundary constraints that guide surface-directed sculpting of cell-laden extracellular matrix hydrogel constructs. Using this approach, we engineered linearly aligned tissues with structural and mechanical anisotropy. A multiscale in silico model of the sculpting process was developed to reveal that cell contractility increases as a function of principal stress polarization in anisotropic tissues. We also show that the anisotropic biophysical microenvironment of linearly aligned tissues potentiates soluble factor-mediated tenogenic and myogenic differentiation of mesenchymal stem cells. The application of our method is demonstrated by (i) skeletal muscle arrays to screen therapeutic modulators of acute oxidative injury and (ii) a 3D microphysiological model of lung cancer cachexia to study inflammatory and oxidative muscle injury induced by tumor-derived signals.

Multilayered film for the controlled formation of freestanding lipid bilayers.

A freestanding lipid bilayer or black lipid membrane is a powerful tool for studying ion channels and for biophysical studies of other membrane proteins under controlled chemical and physical conditions. Even though the lipid bilayer has been considered an excellent sensing platform to detect diverse single molecules from nucleotides to cells, it is not yet widely used, mainly due to its low stability and the expertise needed for membrane formation. To ameliorate the issues of conventional membrane formation techniques, we report a novel layered film that consists of a nonporous layer sandwiched between two porous layers to facilitate bilayer formation. Moreover, the absorption of excess solvent present in the membrane precursor solution can be achieved by the film, enabling control over the membrane formation process. Through this layered design, we could obtain an ideal film that has a reduced and controlled membrane formation time (<30 min) and a sufficient bilayer lifetime (3 h) for ion channel studies and biosensing.

Aptamer-Conjugated Polydiacetylene Colorimetric Paper Chip for the Detection of Bacillus thuringiensis Spores.

A colorimetric polydiacetylene (PDA) paper strip sensor that can specifically recognize Bacillus thuringiensis (BT) HD-73 spores is described in this work. The target-specific aptamer was combined with PDA, and the aptamer-conjugated PDA vesicles were then coated on polyvinylidene fluoride (PVDF) paper strips by a simple solvent evaporation method. The PDA-aptamer paper strips can be used to detect the target without any pre-treatment. Using the paper strip, the presence of BT spores is directly observable by the naked eye based on the unique blue-to-red color transition of the PDA. Quantitative studies using the paper strip were also carried out by analyzing the color transitions of the PDA. The specificity of this PDA sensor was verified with a high concentration of Escherichia coli, and no discernable change was observed. The observable color change in the paper strip occurs in less than 1 h, and the limit of detection is 3 × 107 CFU/mL, much below the level harmful to humans. The PDA-based paper sensor, developed in this work, does not require a separate power or detection device, making the sensor strip highly transportable and suitable for spore analysis anytime and anywhere. Moreover, this paper sensor platform is easily fabricated, can be adapted to other targets, is highly portable, and is highly specific for the detection of BT spores.

Synthetic Anion Transporters as Endoplasmic Reticulum (ER) Stress Inducers.

Cl--ion transporters (2a-2h) were synthesized based on the binding motifs of prodigiosin. Transporter 2e clearly displays Cl--ion transportation activity across both model and live cell membranes. Furthermore, 2e can disrupt Ca2+ homeostasis and increase the intracellular concentration of Ca2+ in the DLD-1 cell. This disruption can lead to Caspase-dependent apoptosis supported by CHOP expression (a marker of ER stress) and the appearance of the cleaved forms of Caspase 3 and PARP.

A Helical Polypeptide-Based Potassium Ionophore Induces Endoplasmic Reticulum Stress-Mediated Apoptosis by Perturbing Ion Homeostasis.

Perturbation of potassium homeostasis can affect various cell functions and lead to the onset of programmed cell death. Although ionophores have been intensively used as an ion homeostasis disturber, the mechanisms of cell death are unclear and the bioapplicability is limited. In this study, helical polypeptide-based potassium ionophores are developed to induce endoplasmic reticulum (ER) stress-mediated apoptosis. The polypeptide-based potassium ionophores disturb ion homeostasis and then induce prolonged ER stress in the cells. The ER stress results in oxidative environments that accelerate the activation of mitochondria-dependent apoptosis. Moreover, ER stress-mediated apoptosis is triggered in a tumor-bearing mouse model that suppresses tumor proliferation. This study provides the first evidence showing that helical polypeptide-based potassium ionophores trigger ER stress-mediated apoptosis by perturbation of potassium homeostasis.

A protamine-conjugated gold decorated graphene oxide composite as an electrochemical platform for heparin detection.

In this study, an effective electrochemical sensor was developed for heparin detection using a protamine-conjugated graphene oxide/gold (GO/Au) composite. Protamine is an antidote that can act as an affinity ligand for heparin. The GO was used as support for signal amplification, and Au nanoparticles (NPs) were employed to immobilize the protamine. This Au NPs also increasing the electron transfer rate and enhancing the signal response during protamine-heparin integration. The proposed affinity sensor had a simple fabrication process, a low detection limit (0.9 nM), a wide linear range (1.9 × 10-7 M to 1.5 × 10-9 M), high stability, and high selectivity in the detection of heparin.

Biomimetic Membranes with Transmembrane Proteins: State-of-the-Art in Transmembrane Protein Applications.

In biological cells, membrane proteins are the most crucial component for the maintenance of cell physiology and processes, including ion transportation, cell signaling, cell adhesion, and recognition of signal molecules. Therefore, researchers have proposed a number of membrane platforms to mimic the biological cell environment for transmembrane protein incorporation. The performance and selectivity of these transmembrane proteins based biomimetic platforms are far superior to those of traditional material platforms, but their lack of stability and scalability rule out their commercial presence. This review highlights the development of transmembrane protein-based biomimetic platforms for four major applications, which are biosensors, molecular interaction studies, energy harvesting, and water purification. We summarize the fundamental principles and recent progress in transmembrane protein biomimetic platforms for each application, discuss their limitations, and present future outlooks for industrial implementation.