Construction of the Glycolysis Metabolic Pathway Inside an Artificial Cell for the Synthesis of Amino Acid and Its Reversible Deformation.

The bottom-up construction of artificial cells is beneficial for understanding cell working mechanisms. The glycolysis metabolism mimicry inside artificial cells is challenging. Herein, the glycolytic pathway (Entner-Doudoroff pathway in archaea) is reconstituted inside artificial cells. The glycolytic pathway comprising glucose dehydrogenase (GDH), gluconate dehydratase (GAD), and 2-keto-3-deoxygluconate aldolase (KDGA) converts glucose molecules to pyruvate molecules. Inside artificial cells, pyruvate molecules are further converted into alanine with the help of alanine dehydrogenase (AlaDH) to build a metabolic pathway for synthesizing amino acid. On the other hand, the pyruvate molecules from glycolysis stimulate the living mitochondria to produce ATP inside artificial cells, which further trigger actin monomers to polymerize to form actin filaments. With the addition of methylcellulose inside the artificial cell, the actin filaments form adjacent to the inner lipid bilayer, deforming the artificial cell from a spherical shape to a spindle shape. The spindle-shaped artificial cell reverses to a spherical shape by depolymerizing the actin filament upon laser irradiation. The glycolytic pathway and its further extension to produce amino acids (or ATP) inside artificial cells pave the path to build functional artificial cells with more complicated metabolic pathways.

Phospholipid Epitaxial Assembly Behavior on a Hydrophobic Highly Ordered Pyrolytic Graphite Surface.

Supported lipid bilayers (SLBs) are excellent models of cell membranes. However, most SLBs exist in the form of phospholipid molecules standing on a substrate, making it difficult to have a side view of the phospholipid membranes. In this study, the phospholipid striped lamella with the arrangement of their alkane tails lying on highly ordered pyrolytic graphite (HOPG) was constructed by a spin coating method. Atomic force microscopy and molecular dynamics simulations are utilized to study the self-assembly of phospholipids on HOPG. Results show that various phospholipids with different packing parameters and electrical property are able to epitaxially adsorb on HOPG. 0.1 mg/mL Plasm PC (0.1 mg/mL) could form a striped monolayer with a width of 5.93 ± 0.21 nm and form relatively stable four striped layers with the concentration increasing to 1 mg/mL. The width of the DOPS multilayer is more than that of electroneutral lipids due to the static electrical repulsion force. This universal strategy sheds light on direct observation of the membrane structure from the side view and modification of 2D materials with amphiphilic biomolecules.

Light-Harvesting Artificial Cells Containing Cyanobacteria for CO2 Fixation and Further Metabolism Mimicking.

The bottom-up constructed artificial cells help to understand the cell working mechanism and provide the evolution clues for organisms. The energy supply and metabolism mimicry are the key issues in the field of artificial cells. Herein, an artificial cell containing cyanobacteria capable of light harvesting and carbon dioxide fixation is demonstrated to produce glucose molecules by converting light energy into chemical energy. Two downstream "metabolic" pathways starting from glucose molecules are investigated. One involves enzyme cascade reaction to produce H2 O2 (assisted by glucose oxidase) first, followed by converting Amplex red to resorufin (assisted by horseradish peroxidase). The other pathway is more biologically relevant. Glucose molecules are dehydrogenated to transfer hydrogens to nicotinamide adenine dinucleotide (NAD+ ) for the production of nicotinamide adenine dinucleotide hydride (NADH) molecules in the presence of glucose dehydrogenase. Further, NADH molecules are oxidized into NAD+ by pyruvate catalyzed by lactate dehydrogenase, meanwhile, lactate is obtained. Therefore, the cascade cycling of NADH/NAD+ is built. The artificial cells built here pave the way for investigating more complicated energy-supplied metabolism inside artificial cells.

Mimicking Cellular Metabolism in Artificial Cells: Universal Molecule Transport across the Membrane through Vesicle Fusion.

Mass transport across cell membranes is a primary process for cellular metabolism. For this purpose, electrostatically mediated membrane fusion is exploited to transport various small molecules including glucose-6-phosphate, isopropyl ß-D-thiogalactoside, and macromolecules such as DNA plasmids from negatively charged large unilamellar vesicles (LUVs) to positively charged giant unilamellar vesicles (GUVs). After membrane fusion between these oppositely charged vesicles, molecules are transported into GUVs to trigger the NAD+ involved enzyme reaction, bacterial gene expression, and in vitro gene expression of green fluorescent protein from a DNA plasmid. The optimized charged lipid percentages are 10% for both positively charged GUVs and negatively charged LUVs to ensure the fusion process. The experimental results demonstrate a universal way for mass transport into the artificial cells through vesicle fusions, which paves a crucial step for the investigation of complicated cellular metabolism.

Micrometer-size double-helical structures from phospholipid-modified carbon nanotubes.

Biomolecular self-assembly plays a key role in the life system. Herein, double-helical phospholipid-modified carbon nanotube structures were constructed via the self-assembly of phospholipids on carbon nanotubes. These micrometer size spring structures may find potential applications in biocompatible microrobots.

Engineering tunable terahertz radiation from an electron bunch using graphene metasurfaces.

We propose an approach to generate tunable terahertz (THz) radiation from an electron bunch passing over the unique graphene metasurface. We not only control the frequency of the THz radiation but also tune the amplitude and direction of the radiation by varying the chemical potential of the graphene. Several new phenomena are observed. The radiation has the same frequency with the resonant frequency of the graphene metasurface at normal incidence. The radiation frequency meets the linear relationship with the chemical potential. The radiation magnitude is the inverse to the reflection magnitude, and the sum of them is close to being a constant. The strong Smith-Purcell radiation on the graphene metasurface is due to the interaction between the electron bunch and periodic surface plasmon polaritons (SPPs). The stronger the SPP, the higher is the radiation magnitude that is obtained. These results would provide a promising way for developing tunable radiation in the THz band.

Principles and Applications of Single Particle Tracking in Cell Research.

It is a tough challenge for many decades to decipher the complex relationships between cell behaviors and cellular physical properties. Single particle tracking (SPT) with high spatial and temporal resolution has been applied extensively in cell research to understand physicochemical properties of cells and their bio-functions by tracking endogenous or exogenous probes. This review describes the fundamental principles of SPT as well as its applications in intracellular mechanics, membrane dynamics, organelles distribution, and processes of internalization and transport. Finally, challenges and future directions of SPT are also discussed.

Recoverable peroxidase-like Fe3O4@MoS2-Ag nanozyme with enhanced antibacterial ability.

Antibacterial agents with enzyme-like properties and bacteria-binding ability have provided an alternative method to efficiently disinfect drug-resistance microorganism. Herein, a Fe3O4@MoS2-Ag nanozyme with defect-rich rough surface was constructed by a simple hydrothermal method and in-situ photodeposition of Ag nanoparticles. The nanozyme exhibited good antibacterial performance against E. coli (~69.4%) by the generated ROS and released Ag+, while the nanozyme could further achieve an excellent synergistic disinfection (~100%) by combining with the near-infrared photothermal property of Fe3O4@MoS2-Ag. The antibacterial mechanism study showed that the antibacterial process was determined by the collaborative work of peroxidase-like activity, photothermal effect and leakage of Ag+. The defect-rich rough surface of MoS2 layers facilitated the capture of bacteria, which enhanced the accurate and rapid attack of •OH and Ag+ to the membrane of E. coli with the assistance of local hyperthermia. This method showed broad-spectrum antibacterial performance against Gram-negative bacteria, Gram-positive bacteria, drug-resistant bacteria and fungal bacteria. Meanwhile, the magnetism of Fe3O4 was used to recycle the nanozyme. This work showed great potential of engineered nanozymes for efficient disinfection treatment.

Effect of hyperbaric oxygen combined with alprostadil in the treatment of elderly diabetic nephropathy and effects on serum miR-126 and miR-342 levels.

This study aims to investigate the effect of hyperbaric oxygen combined with alprostadil in the treatment of elderly diabetic nephropathy (DN) and its effect on serum miR-126 and miR-342 levels. The total effective rate of the study group was 91.53% after treatment, which was higher than that (74.58%) of the control group (p<0.05); the levels of UAER, Scr, BUN and HbA1c, FPG, 2h PG were lowered in the two groups after treatment, and the levels of these indexes were lower in the study group than those in the control group (p<0.05); the levels of vWF, ET-1, CD8+, miR-342 were lowered after treatment for the two groups, and the levels of these indexes were lower in the study group than those in the control group; the levels of NO, CD3+, CD4+ and miR-126 were increased after treatment and the levels were higher in the study group than those in the control group (p<0.05). The application of hyperbaric oxygen combined with alprostadil in the treatment of elderly DN patients can improve renal function, lower blood glucose, improve vascular endothelial function and immune function, adjust serum miR-126 and miR-342 levels, thereby increasing curative effect.

Phospholipid Self-Assemblies Shaped Like Ancient Chinese Coins for Artificial Organelles.

Phospholipid self-assemblies are ubiquitous in organisms. Nonspherical lipid-based proto-organelles bear the merits with structures similar to real organelles. It is still a challenge to mimic mass transport between organelles inside cells. Herein, unusual phospholipid self-assemblies shaped like ancient Chinese coins (ACC) were discovered by the recrystallization of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine in an ethanol/water solution from 50 to 25 °C with a certain cooling rate. Their diameter and the ratio of the square edge to the disk diameter were controlled by varying ethanol percentage, lipid concentration, and cooling rate. The ACC-shaped phospholipid bicelles expanded to stacked cisterna structures in pure water, which were regarded as artificial organelles. Mass transport among organelles in a cell was mimicked via the membrane fusion of vesicle shuttles and artificial organelles, which induced cascade enzyme reactions inside artificial organelles. The ACC-shaped phospholipid assemblies provide nice platforms for the studies of cell biology and bottom-up synthetic biology.

Chemical Information Exchange in Organized Protocells and Natural Cell Assemblies with Controllable Spatial Positions.

An ultrasound-based platform is established to prepare homogenous arrays of giant unilamellar vesicles (GUVs) or red blood cell (RBCs), or hybrid assemblies of GUV/RBCs. Due to different responses to the modulation of the acoustic standing wave pressure field between the GUVs and RBCs, various types of protocell/natural cell hybrid assemblies are prepared with the ability to undergo reversible dynamic reconfigurations from vertical to horizontal alignments, or from 1D to 2D arrangements. A two-step enzymatic cascade reaction between transmitter glucose oxidase-containing GUVs and peroxidase-active receiver RBCs is used to implement chemical signal transduction in the different hybrid micro-arrays. Taken together, the obtained results suggest that the ultrasound-based micro-array technology can be used as an alternative platform to explore chemical communication pathways between protocells and natural cells, providing new opportunities for bottom-up synthetic biology.

In situ Surface Charge Density Visualization of Self-assembled DNA Nanostructures after Ion Exchange.

The charge density of DNA is a key parameter in strand hybridization and for the interactions occurring between DNA and molecules in biological systems. Due to the intricate structure of DNA, visualization of the surface charge density of DNA nanostructures under physiological conditions was not previously possible. Here, we perform a simultaneous analysis of the topography and surface charge density of DNA nanostructures using atomic force microscopy and scanning ion conductance microscopy. The effect of in situ ion exchange using various alkali metal ions is tested with respect to the adsorption of DNA origami onto mica, and a quantitative study of surface charge density reveals ion exchange phenomena in mica as a key parameter in DNA adsorption. This is important for structure-function studies of DNA nanostructures. The research provides an efficient approach to study surface charge density of DNA origami nanostructures and other biological molecules at a single molecule level.

Chemical Signal Communication between Two Protoorganelles in a Lipid-Based Artificial Cell.

The chemical signal communication among organelles in the cell is extremely important for life. We demonstrate here the chemical signal communication between two protoorganelles using cascade enzyme reactions in a lipid-based artificial cell. Two protoorganelles inside the artificial cell are large unilamellar vesicles containing glucose oxidase (GOx-LUVs) and a vesicle containing horseradish peroxidase (HRP) and Amplex red, respectively. The glucose molecules outside the artificial cell penetrate the lipid bilayer through mellitin pores and enter into one protoroganelle (GOx-LUV) to produce H2O2, which subsequently is transported to the other protoorganelle to oxidize Amplex red into red resorufin catalyzed by HRP. The number of GOx-LUVs in an artificial cell is controlled by using a GOx-LUV solution with different density during the electroformation. The reaction rate for resorufin in the protoorganelle increases with more GOx-LUVs inside the artificial cell. The artificial cell developed here paves the way for a more complicated signal transduction mechanism study in a eukaryocyte.

Giant Unilamellar Vesicle Microarrays for Cell Function Study.

Giant unilamellar vesicles (GUVs) are widely used as artificial cell models which contribute to elucidate fundamental questions on origin of life and cell functions. Herein, the GUV microarrays were developed using a point-to-plane electrode system combined with microcontact stripping technique. The biomolecules (DNA, etc.) were selectively encapsulated only inside patterned GUVs. The GUV arrays were used to investigate species mass transport across cell membranes. The release of carboxyfluorescein from GUVs showed a melittin concentration dependent manner. The diffusion coefficient were 0.37 × 10-11, 0.36 × 10-11, 0.54 × 10-11, 1.10 × 10-11, 1.74 × 10-11, 2.31 × 10-11, and 3.62 × 10-11 m2/s for 0, 0.5, 1.0, 2.0, 3.0, 4.0, and 5.0 µM melittin, respectively. The GUV arrays were also a good platform for cell metabolism investigation. Horseradish peroxidase (HRP) loaded GUV microarrays were used to mimic internal metabolism by exposing them to the substrates of H2O2 and o-PD to yield fluorescent 2,3-diaminophenazine (2,3-DAP).The proposed GUV arrays have great potential in cell function studies.

MoS2 @HKUST-1 Flower-Like Nanohybrids for Efficient Hydrogen Evolution Reactions.

A novel MoS2 -based flower-like nanohybrid for hydrogen evolution was fabricated through coating the Cu-containing metal-organic framework (HKUST-1) onto MoS2 nanosheets. It is the first time that MoS2 @HKUST-1 nanohybrids have been reported for the enhanced electrochemical performance of HER. The morphologies and components of the MoS2 @HKUST-1 flower-like nanohybrids were characterized by scanning electron microscopy, X-ray diffraction analysis and Fourier transform infrared spectroscopy. Compared with pure MoS2 , the MoS2 @HKUST-1 hybrids exhibit enhanced performance on hydrogen evolution reaction with an onset potential of -99 mV, a smaller Tafel slope of 69 mV dec-1 , and a Faradaic efficiency of nearly 100 %. The MoS2 @HKUST-1 flower-like nanohybrids exhibit excellent stability in acidic media. This design opens new possibilities to effectively synthesize non-noble metal catalysts with high performance for the hydrogen evolution reaction (HER).

Impact of Electric Fields on the Nanoscale Behavior of Lipid Monolayers at the Surface of Graphite in Solution.

The nanoscale organization and dynamics of lipid molecules in self-assembled membranes is central to the biological function of cells and in the technological development of synthetic lipid structures as well as in devices such as biosensors. Here, we explore the nanoscale molecular arrangement and dynamics of lipids assembled in monolayers at the surface of highly ordered pyrolytic graphite (HOPG), in different ionic solutions, and under electrical potentials. Using a combination of atomic force microscopy and fluorescence recovery after photobleaching, we show that HOPG is able to support fully formed and fluid lipid membranes, but mesoscale order and corrugations can be observed depending on the type of the lipid considered (1,2-dioleoyl- sn-glycero-3-phosphocholine, 1,2-dioleoyl- sn-glycero-3-phospho-l-serine (DOPS), and 1,2-dioleoyl-3-trimethylammoniumpropane) and the ion present (Na+, Ca2+, Cl-). Interfacial solvation forces and ion-specific effects dominate over the electrostatic changes induced by moderate electric fields (±1.0 V vs Ag/AgCl reference electrode) with particularly marked effects in the presence of calcium, and for DOPS. Our results provide insights into the interplay between the molecular, ionic, and electrostatic interactions and the formation of dynamical ordered structures in fluid lipid membranes.

Phospholipid-Block Copolymer Hybrid Vesicles with Lysosomal Escape Ability.

The success of nanoparticulate formulations in drug delivery depends on various aspects including their toxicity, internalization, and intracellular location. Vesicular assemblies consisting of phospholipids and amphiphilic block copolymers are an emerging platform, which combines the benefits from liposomes and polymersomes while overcoming their challenges. We report the synthesis of poly(cholesteryl methacrylate)- block-poly(2-(dimethylamino) ethyl methacrylate) (pCMA- b-pDMAEMA) block copolymers and their assembly with phospholipids into hybrid vesicles. Their geometry, their ζ-potential, and their ability to adsorb onto polymer-coated surfaces were assessed. Giant unilamellar vesicles were employed to confirm the presence of both the phospholipids and the block copolymer in the same membrane. Furthermore, the cytotoxicity of selected hybrid vesicles was determined in RAW 264.7 mouse macrophages, primary rat Kupffer cells, and human macrophages. The internalization and lysosomal escape ability of the hybrid vesicles were confirmed using RAW 264.7 mouse macrophages. Taken together, our findings illustrate that the reported hybrid vesicles are a promising complementary drug delivery platform for existing liposomes and polymersomes.

Multicompartmentalized Microreactors Containing Nuclei and Catalase-Loaded Liposomes.

Multicompartmentalized microreactors are considered as cell mimics with hierarchical structures inspired by mammalian cells. We report the successful assembly and encapsulation of purified nuclei from RAW 264.7 cells (pNuc) into alginate-based microreactors. We demonstrate the preserved function of nuclei within the microreactors for mRNA production. Further, we load catalase-loaded liposomes (Lcat) into the microreactors to break down hydrogen peroxide (H2O2) into oxygen and water. Assemblies containing both natural pNuc and synthetic Lcat show significantly higher mRNA production in the presence of H2O2 compared to microreactors without Lcat or no H2O2 present, suggesting a beneficial effect of the locally enzymatically produced oxygen for transcription. This novel type of microreactors, containing both natural and synthetic compartments, presents a substantial advancement from assemblies equipped with solely synthetic units and offers opportunities as hypoxia models or for cell-free protein synthesis.

A green method to synthesize flowerlike Fe(OH)3 microspheres for enhanced adsorption performance toward organic and heavy metal pollutants.

Dyestuffs and heavy metal ions in water are seriously harmful to the ecological environment and human health. Three-dimensional (3D) flowerlike Fe(OH)3 microspheres were synthesized through a green yet low-cost injection method, for the removal of organic dyes and heavy metal ions. The Fe(OH)3 microspheres were characterized by thermal gravimetric analysis (TGA), Fourier transform infrared (FT-IR), and transmission electron microscopy (TEM) techniques. The adsorption kinetics of Congo Red (CR) on Fe(OH)3 microspheres obeyed the pseudo-second-order model. Cr6+ and Pb2+ adsorption behaviors on Fe(OH)3 microspheres followed the Langmuir isotherm model. The maximum adsorption capacities of the synthesized Fe(OH)3 were 308, 52.94, and 75.64mg/g for CR, Cr6+, and Pb2+ respectively. The enhanced adsorption performance originated from its surface properties and large specific surface area of 250m2/g. The microspheres also have excellent adsorption stability and recyclability. Another merit of the Fe(OH)3 material is that it also acts as a Fenton-like catalyst. These twin functionalities (both as adsorbent and Fenton-like catalyst) give the synthesized Fe(OH)3 microspheres great potential in the field of water treatment.

A Fissionable Artificial Eukaryote-like Cell Model.

The use of artificial cells has attracted considerable attention in various fields from biotechnology to medicine. Here, we develop a cell-sized vesicle-in-vesicle (VIV) structure containing a separate inner vesicle (IV) that can be loaded with DNA. We use polymerase chain reaction (PCR) to successfully amplify the amount of DNA confined to the IV. Subsequent osmotic stress-induced fission of a mother VIV into two daughter VIVs successfully divides the IV content while keeping it confined to the IV of the daughter VIVs. The fission rate was estimated to be ∼20% quantified by fluorescence microscope. Our VIV structure represents a step forward toward construction of an advanced, fissionable cell model.