Multicompartmental coacervate-based protocell by spontaneous droplet evaporation.

Hierarchical compartmentalization, a hallmark of both primitive and modern cells, enables the concentration and isolation of biomolecules, and facilitates spatial organization of biochemical reactions. Coacervate-based compartments can sequester and recruit a large variety of molecules, making it an attractive protocell model. In this work, we report the spontaneous formation of core-shell cell-sized coacervate-based compartments driven by spontaneous evaporation of a sessile droplet on a thin-oil-coated substrate. Our analysis reveals that such far-from-equilibrium architectures arise from multiple, coupled segregative and associative liquid-liquid phase separation, and are stabilized by stagnation points within the evaporating droplet. The formation of stagnation points results from convective capillary flows induced by the maximum evaporation rate at the liquid-liquid-air contact line. This work provides valuable insights into the spontaneous formation and maintenance of hierarchical compartments under non-equilibrium conditions, offering a glimpse into the real-life scenario.

TET3-mediated novel regulatory mechanism affecting trophoblast invasion and migration: Implications for preeclampsia development.

INTRODUCTION: Aberrant expression of genes has been demonstrated to be related to the abnormal function of trophoblasts and lead to the occurrence and progression of Preeclampsia (PE). However, the underlying mechanism of PE has not been elucidated. METHODS: We performed PCR analysis to investigate TET3 expression in PE placental tissues. Cell assays were performed in HTR-8/SVneo and JAR. Cell invasion and migration events were investigated by transwell assays in vitro. ChIP-PCR and Targeted bisulfite sequencing were conducted to detect the demethylation of related CpG sites in the KLF13 promoter after inhibition of TET3. In conjunction with bioinformatics analysis, luciferase reporter assays were performed to elucidate the mechanism by which miR-544 binds to TET3/KLF13 mRNA. RESULTS: In this study, we identified genes associated with human extravillous trophoblasts by conducting sc-seq analysis from the GEO. Then, we measured the expression of TET3 in a larger clinical sample. The results showed that TET3, a DNA demethylase, was found to be expressed at much higher levels in the preeclamptic placenta compared to the control. Then, the inhibition of TET3 significantly promoted trophoblast cell migration and invasion. Conversely, TET3 overexpression suppressed cell migration and invasion in vitro. Further RNA sequencing and mechanism analysis indicated that the inhibition of TET3 suppressed the activation of KLF13 by reducing the demethylation of related CpG sites in the KLF13 promoter, thereby transcriptionally inactivating KLF13 expression. Moreover, luciferase reporter assay indicate that TET3 and KLF13 were direct targets of miR-544. DISCUSSION: This study uncovers a TET3-mediated regulatory mechanism in PE progression and suggests that targeting the placental miR-544-TET3-KLF13-axis might provide new diagnostic and therapeutic strategies for PE.

Facile and Programmable Capillary-Induced Assembly of Prototissues via Hanging Drop Arrays.

An important goal for bottom-up synthetic biology is to construct tissue-like structures from artificial cells. The key is the ability to control the assembly of the individual artificial cells. Unlike most methods resorting to external fields or sophisticated devices, inspired by the hanging drop method used for culturing spheroids of biological cells, we employ a capillary-driven approach to assemble giant unilamellar vesicles (GUVs)-based protocells into colonized prototissue arrays by means of a coverslip with patterned wettability. By spatially confining and controllably merging a mixed population of lipid-coated double-emulsion droplets that hang on a water/oil interface, an array of synthetic tissue-like constructs can be obtained. Each prototissue module in the array comprises multiple tightly packed droplet compartments where interfacial lipid bilayers are self-assembled at the interfaces both between two neighboring droplets and between the droplet and the external aqueous environment. The number, shape, and composition of the interconnected droplet compartments can be precisely controlled. Each prototissue module functions as a processer, in which fast signal transports of molecules via cell-cell and cell-environment communications have been demonstrated by molecular diffusions and cascade enzyme reactions, exhibiting the ability to be used as biochemical sensing and microreactor arrays. Our work provides a simple yet scalable and programmable method to form arrays of prototissues for synthetic biology, tissue engineering, and high-throughput assays.

Improved algorithm for determining cable saddle pre-offsets considering the coupling effect of tower and splay saddles.

By analysing the mechanical and geometrical relations between the main cable, tower, and splay saddles, and considering the coupling effect of the tower and splay saddles, an improved algorithm is proposed to determine the cable saddles pre-offsets of suspension bridges. The equilibrium relationship of the cable saddles, the compatible deformation condition, and the basic equation of the main cable shape are considered to establish several coupled non-linear equations up to 19, and the tower and splay saddle pre-offsets are obtained by solving the above equations with the Newton-Raphson method. This paper presents the initial value selection principle and the constraint conditions for solving the cable saddle pre-offsets of the plane cable suspension bridge and the calculation process ensures convergence. The calculation example demonstrates that the improved algorithm without an exact initial value can achieve excellent convergence.

Continuous microfluidic fabrication of anisotropic microparticles for enhanced wastewater purification.

Anisotropic microparticles containing functional nanomaterials have attracted growing interest due to their enhanced performance in diverse applications ranging from catalysts to environmental remediation. However, the preparation of anisotropic microparticles with highly controlled morphologies and dimensions usually suffers from a limited material choice. Here, we develop a facile strategy to continuously prepare anisotropic microparticles with their shapes changing from spherical to pear-like, maraca-like and rod-like for enhanced water decontamination. Anisotropic microparticles are produced by deforming oil-droplet templates within microfibers and then locking their shapes via thermo/photo-polymerization. The sizes and geometries of the oil-droplet templates are precisely controlled by varying the fluid flow conditions. In addition, porous spherical and rod-like microparticles are functionalized for photocatalytic degradation of organic contaminants by incorporating functional TiO2 and Fe3O4 nanoparticles. Compared to spherical microparticles with equal volume, functionalized rod-like microparticles exhibit better performance in removal of contaminants due to their larger specific surface area, which facilitates the contact between the loaded catalysts and organic pollutants. Moreover, the magnetic rod-like microparticles can be easily recovered and reused without deterioration of catalytic performance. The proposed strategy in this study is useful for producing anisotropic microparticles with well-tailored shapes via different polymerization methods and extending their potential applications.

Dielectrophoresis Response of Water-in-Oil-in-Water Double Emulsion Droplets with Singular or Dual Cores.

Microfluidic technologies have enabled generation of exquisite multiple emulsion droplets, which have been used in many fields, including single-cell assays, micro-sized chemical reactions, and material syntheses. Electrical controlling is an important technique for droplet manipulation in microfluidic systems, but the dielectrophoretic behaviors of multiple emulsion droplets in electrical fields are rarely studied. Here, we report on the dielectrophoresis response of double emulsion droplets in AC electric fields in microfluidic channel. A core-shell model is utilized for analyzing the polarization of droplet interfaces and the overall dielectrophoresis (DEP) force. The water-in-oil-in-water droplets, generated by glass capillary devices, experience negative DEP at low field frequency. At high frequency, however, the polarity of DEP is tunable by adjusting droplet shell thickness or core conductivity. Then, the behavior of droplets with two inner cores is investigated, where the droplets undergo rotation before being repelled or attracted by the strong field area. This work should benefit a wide range of applications that require manipulation of double emulsion droplets by electric fields.

Comparative proteome analysis of amniotic fluids and placentas from patients with idiopathic polyhydramnios.

INTRODUCTION: Idiopathic polyhydramnios (IPH) is an abnormal increase in amniotic fluid volume (AFV). This condition has unknown etiologies and is associated with various adverse pregnancy outcomes including maternal and fetal complication. This study aims to establish a comparative proteome profile for the human amniotic fluid (AF) of IPH and normal pregnancies and identify the responsible mediators and pathways that regulate AFV. METHODS: We first employed coupled isobaric tags for relative and absolute quantitation (iTRAQ) proteomics and bioinformatics analysis to examine the differentially expression proteins (DEPs) in the AF of IPH and normal pregnancies. Second, CUL5, HIP1, FSTL3, and LAMP2 proteins were selected for verification in amnion, chorion, and placental tissues by Western blot analysis. RESULTS: We identified 357 DEPs with 282 upregulated and 75 downregulated. Bioinformatics analysis revealed that cell, cellular process, and binding were the most enriched Gene Ontology terms. Amoebiasis, hematopoietic cell lineage, and NF-kappa B signaling pathway were the top significant pathways. In the verification procedure, FSTL3 protein had a highly significant expression in the amnion, chorion, and placentas of IPH and normal AFV groups (p < 0.05). DISCUSSION: Our results provide new insights into idiopathic polyhydramnios and offer fundamental points for future studies on AFV.

Efficient particle and droplet manipulation utilizing the combined thermal buoyancy convection and temperature-enhanced rotating induced-charge electroosmotic flow.

Efficient granular sample manipulation is crucial for various microfluidic-based applications such as material synthesis and drug delivery. Herein we present a novel method to efficiently manipulate microbeads and droplets using the combined thermal buoyancy convection and temperature-enhanced rotating induced-charge electroosmotic flow. Within the granular fluid, a pair of counter-rotating microvortices is formed above the floating electrode, leading to the formation of a flow stagnation region at the bottom center. Granular samples then can be effectively transported to this region by the Stokes drag, and the concentration performance can be flexibly manipulated by adjusting the energization strategies of the chip. The contributions of fluid convection, dielectrophoresis, thermophoresis, and gravity force to particle migration are first studied and compared, proving that the convection flow and gravity force are mainly responsible for particle migration and deposition respectively. Then the systematic enriching experiments of 4-µm silica particles demonstrate that the particle migration velocity can be highly improved by the combined thermal-electrical field. Finally, the effective concentration of nanocopper particles and the assembly of oil-in-water/water-in-oil-in-water droplets indicate that this approach is capable of manipulating diverse granular samples. Therefore, this strategy can be attractive for lots of microfluidic-based applications because of its high efficiency and simplicity.

Compound-Droplet-Pairs-Filled Hydrogel Microfiber for Electric-Field-Induced Selective Release.

The separate co-encapsulation and selective controlled release of multiple encapsulants in a predetermined sequence has potentially important applications for drug delivery and tissue engineering. However, the selective controlled release of distinct contents upon one triggering event for most existing microcarriers still remains challenging. Here, novel microfluidic fabrication of compound-droplet-pairs-filled hydrogel microfibers (C-Fibers) is presented for two-step selective controlled release under AC electric field. The parallel arranged compound droplets enable the separate co-encapsulation of distinct contents in a single microfiber, and the release sequence is guaranteed by the discrepancy of the shell thickness or core conductivity of the encapsulated droplets. This is demonstrated by using a high-frequency electric field to trigger the first burst release of droplets with higher conductivity or thinner shell, followed by the second release of the other droplets under low-frequency electric field. The reported C-Fibers provide novel multidelivery system for a wide range of applications that require controlled release of multiple ingredients in a prescribed sequence.

Electric Field-Induced Cutting of Hydrogel Microfibers with Precise Length Control for Micromotors and Building Blocks.

Microfiber modules with controllable lengths emerged as novel biomimetic platforms and are significant for many tissue engineering applications. However, accurately controlling the length of microfibers on the scale of millimeter or even micrometer still remains challenging. Here, a novel and scalable strategy to generate microfiber modules with precisely tunable lengths ranging from 100 to 3500 µm via an alternating current (AC) electric field is presented. To control the microfiber length, double-emulsion droplets containing a chelating agent (sodium citrate) as a spacing node are first uniformly embedded in the microfibers in a controllable spatial arrangement. This process is precisely tuned by adjusting the flow rates, thus, tailoring the resulting multicompartmental microfiber structure. Next, an AC voltage signal is used to trigger the electric field-induced cutting process, where the time-averaged electrical force acting on the induced bipolar charge from the Maxwell-Wagner structural polarization mechanism breaks the stress balance at the interfaces, rupturing the double-emulsion droplets, and resulting in the burst release of the encapsulated chelating agents into the hydrogel cavity. The outer hydrogel shell is quickly dissolved by a chemical reaction, cutting the long fiber into a series of microfiber units of given length. Furthermore, adding magnetic nanoparticles endows magnetic functionality with these microfiber modules, which are allowed to serve as micromotors and building blocks. This electro-induced cutting method provides a facile strategy for the fabrication of microfibers with desired lengths, showing considerable promise for various chemical and biological applications.

Electrically controlled rapid release of actives encapsulated in double-emulsion droplets.

Controlled release of multiple actives after encapsulation in a microenvironment is significant for various biological and chemical applications such as controlled drug delivery and transplantation of encapsulated cells. However, traditional systems often lack efficient encapsulation and release of multiple actives, especially when incorporated substances must be released at a targeted location. Here, we present a straightforward approach to release multiple actives at a prescribed position in microfluidic systems; one or two actives are encapsulated in water-in-oil-in-water emulsion droplets, followed by controlled release of the actives via an alternating current electric field. An electric field-induced compression due to Maxwell-Wagner interfacial polarization overcomes the disjoining pressure at the thin shell and leads to the thinning and rupture of the oil layer of the droplets, resulting in the release of the encapsulated actives to the suspending medium. This technique is feasible for encapsulation and release of various reagents in terms of ion species and ion concentrations. Moreover, polymer nanoparticles and yeast cells can also be included in the droplets and then be released at targeted locations. This versatile method should be well-suited for targeted delivery of various active ingredients such as functional chemical reagents and biological cells.

Sequential Coalescence Enabled Two-Step Microreactions in Triple-Core Double-Emulsion Droplets Triggered by an Electric Field.

Advances in microfluidic emulsification have enabled the generation of exquisite multiple-core droplets, which are promising structures to accommodate microreactions. An essential requirement for conducting reactions is the sequential coalescence of the multiple cores encapsulated within these droplets, therefore, mixing the reagents together in a controlled sequence. Here, a microfluidic approach is reported for the conduction of two-step microreactions by electrically fusing three cores inside double-emulsion droplets. Using a microcapillary glass device, monodisperse water-in-oil-in-water droplets are fabricated with three compartmented reagents encapsulated inside. An AC electric field is then applied through a polydimethylsiloxane chip to trigger the sequential mixing of the reagents, where the precise sequence is guaranteed by the discrepancy of the volume or conductivity of the inner cores. A two-step reaction in each droplet is ensured by two times of core coalescence, which totally takes 20-40 s depending on varying conditions. The optimal parameters of the AC signal for the sequential fusion of the inner droplets are identified. Moreover, the capability of this technique is demonstrated by conducting an enzyme-catalyzed reaction used for glucose detection with the double-emulsion droplets. This technique should benefit a wide range of applications that require multistep reactions in micrometer scale.

Continuously Electrotriggered Core Coalescence of Double-Emulsion Drops for Microreactions.

Microfluidically generated double emulsions are promising templates for microreactions, which protect the reaction from external disturbance and enable in vitro analyses with large-scale samples. Controlled combination of their inner droplets in a continuous manner is an essential requirement toward truly applications. Here, we first generate dual-cored double-emulsion drops with different inner encapsulants using a capillary microfluidic device; next, we transfer the emulsion drops into another electrode-integrated polydimethylsiloxane microfluidic device and utilize external AC electric field to continuously trigger the coalescence of inner cores inside these emulsion drops in continuous flow. Hundreds of thousands of monodisperse microreactions with nanoliter-scale reagents can be conducted using this approach. The performance of core coalescence is investigated as a function of flow rate, applied electrical signal, and core conductivity. The coalescence efficiency can reach up to 95%. We demonstrate the utility of this technology for accommodating microreactions by analyzing an enzyme catalyzed reaction and by fabricating cell-laden hydrogel particles. The presented method can be readily used for the controlled triggering of microreactions with high flexibility for a wide range of applications, especially for continuous chemical or cell assays.

A dual-core double emulsion platform for osmolarity-controlled microreactor triggered by coalescence of encapsulated droplets.

Droplet-based microfluidics has provided a means to generate multi-core double emulsions, which are versatile platforms for microreactors in materials science, synthetic biology, and chemical engineering. To provide new opportunities for double emulsion platforms, here, we report a glass capillary microfluidic approach to first fabricate osmolarity-responsive Water-in-Oil-in-Water (W/O/W) double emulsion containing two different inner droplets/cores and to then trigger the coalescence between the encapsulated droplets precisely. To achieve this, we independently control the swelling speed and size of each droplet in the dual-core double emulsion by controlling the osmotic pressure between the inner droplets and the collection solutions. When the inner two droplets in one W/O/W double emulsion swell to the same size and reach the instability of the oil film interface between the inner droplets, core-coalescence happens and this coalescence process can be controlled precisely. This microfluidic methodology enables the generation of highly monodisperse dual-core double emulsions and the osmolarity-controlled swelling behavior provides new stimuli to trigger the coalescence between the encapsulated droplets. Such swelling-caused core-coalescence behavior in dual-core double emulsion establishes a novel microreactor for nanoliter-scale reactions, which can protect reaction materials and products from being contaminated or released.

FXR antagonism of NSAIDs contributes to drug-induced liver injury identified by systems pharmacology approach.

Non-steroidal anti-inflammatory drugs (NSAIDs) are worldwide used drugs for analgesic, antipyretic, and anti-inflammatory therapeutics. However, NSAIDs often cause several serious liver injuries, such as drug-induced liver injury (DILI), and the molecular mechanisms of DILI have not been clearly elucidated. In this study, we developed a systems pharmacology approach to explore the mechanism-of-action of NSAIDs. We found that the Farnesoid X Receptor (FXR) antagonism of NSAIDs is a potential molecular mechanism of DILI through systematic network analysis and in vitro assays. Specially, the quantitative real-time PCR assay reveals that indomethacin and ibuprofen regulate FXR downstream target gene expression in HepG2 cells. Furthermore, the western blot shows that FXR antagonism by indomethacin induces the phosphorylation of STAT3 (signal transducer and activator of transcription 3), promotes the activation of caspase9, and finally causes DILI. In summary, our systems pharmacology approach provided novel insights into molecular mechanisms of DILI for NSAIDs, which may propel the ways toward the design of novel anti-inflammatory pharmacotherapeutics.