The gene expression differs in the nuclei of normal and malignant mammalian cells, and transcription is a critical initial step, which defines the difference. The mechanical properties of ...transcriptionally active chromatin are still poorly understood. Recently we have probed transcriptionally active chromatin of the nuclei subjected to mechanical stress, by Atomic Force Microscopy (AFM) 1. Nonetheless, a systematic study of the phenomenon is needed.
Nuclei were deformed and studied by AFM. Non-deformed nuclei were studied by fluorescence confocal microscopy. Their transcriptional activity was studied by RNA electrophoresis.
The malignant nuclei under the study were stable to deformation and assembled of 100–300 nm beads-like units, while normal cell nuclei were prone to deformation. The difference in stability to deformation of the nuclei correlated with DNA supercoiling, and transcription-depended units were responsive to supercoils breakage. The inhibitors of the topoisomerases I and II disrupted supercoiling and made the malignant nucleus prone to deformation. Cell nuclei treatment with histone deacetylase inhibitors (HDACIs) preserved the mechanical stability of deformed malignant nuclei and, at the same time, made it possible to observe chromatin decondensation up to 20–60 nm units. The AFM results were supplemented with confocal microscopy and RNA electrophoresis data.
Self-assembly of transcriptionally active chromatin and its decondensation, driven by DNA supercoiling-dependent rigidity, was visualized by AFM in the mechanically deformed nuclei.
We demonstrated that supercoiled DNA defines the transcription mechanics, and hypothesized the nuclear mechanics in vivo should depend on the chromatin architecture.
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•Mechanically deformed normal and malignant cells nuclei were assessed by AFM.•The nuclei of the malignant cells revealed a landscape assembled from 100 to 300 nm beads-like rigid units.•Topoisomerases I and II inhibitors broke supercoils, making nuclear chromatin prone to deformation.•Chromatin decondensation, induced by HDACIs treatment, revealed ∼20 nm rigid units.•The landscape represented transcription events, dependent on the DNA supercoiling.
Nowadays droplet microfluidics is widely used to perform high throughput assays and for the synthesis of micro- and nanoparticles. These applications usually require packaging several reagents into ...droplets and their mixing to start a biochemical reaction. For rapid mixing microfluidic devices usually require additional functional elements that make their designs more complex. Here we perform a series of 2D numerical simulations, followed by experimental studies, and introduce a novel asymmetric flow-focusing droplet generator, which enhances mixing during droplet formation due to a 2D or 3D asymmetric vortex, located in the droplet formation area of the microfluidic device. Our results suggest that 2D numerical simulations can be used for qualitative analysis of two-phase flows and droplet generation process in quasi-two-dimensional devices, while the relative simplicity of such simulations allows them to be easily applied to fairly complicated microfluidic geometries. Mixing inside droplets formed in the asymmetric generator occurs up to six times faster than in a conventional symmetric one. The best mixing efficiency is achieved in a specific range of droplet volumes, which can be changed by scaling the geometry of the device. Thus, the droplet generator suggested here can significantly simplify designs of microfluidic devices because it enables both the droplet formation and fast mixing of the reagents within droplets. Moreover, it can be used to precisely estimate reaction kinetics.
To create a high throughput system for screening single cells in "water-in-oil" droplets, a key role is played by the method of controlling fluid flows in a microfluidic droplet generator. This is ...due to the fact that it is required to obtain monodisperse drops-microcapsules, place cells in them and be able to manipulate them. For these purposes, not only systems based on syringe pumps are suitable, but also pressure control units may be used. Unfortunately, pressure control instruments on the market for researchers are represented by only a few companies and are quite expensive. In this work, we develop a homemade microfluidic pressure controller based on SMC ITV 0010 pneumatic regulators and show its operability. Also, we investigated the conditions for reproducible generation of water-in-oil emulsions in a microfluidic flow focusing droplet generator by pressure driven fluid flows.
The results of an experimental study on the gas-liquid flow structure and interphase mass transfer are presented for a periodic slug flow of ethanol-CO
2
mixture in a straight microchannel and a ...microchannel with a sudden change in the cross-section. The experiments were carried out for a microchannel with successive contractions and expansions along its length and average cross-section of
m
2
. For determination of the change in the volume of the gas slug along the microchannel length because of CO
2
absorption, the method of high-speed flow visualization was used, followed by digital image processing. The frequency of generation, speed, size, and volume variation of the gas bubbles were measured along the length of the channel. From the data obtained, the volumetric mass transfer coefficient from the liquid side was determined. A comparison was made of the obtained volumetric mass transfer coefficient for the straight channel and for the channel with sudden enlargement of the cross-section: slight enhancement of the mass transfer in the channel with the sudden change in the cross-section, resulting from the pulsating shape of the bubbles, has been shown.
The structure of a slug gas-liquid flow and interfacial mass transfer during the flow of ethanol-N
2
and ethanol-CO
2
mixtures in a horizontal rectangular microchannel have been experimentally ...studied. The experiments were carried out in a straight microchannel with a cross section of 380·190 µm. To determine a change in the volume of a gas bubble along the microchannel length, the method of high-speed visualization with digital processing was applied. In a wide range of gas flow rates, the bubble frequency and the volume of gas bubbles and their velocity were measured, and the volumetric coefficient of mass transfer from liquid was determined for the ethanol—CO
2
mixture. A physically substantiated model of interfacial mass transfer for a slug flow in a channel of rectangular cross section, which takes into account the circulation flow in a liquid slug, was proposed.
To characterize the effect of oxidative stress (OS) on the ability of erythrocytes to pass through microvessels and capillaries, we studied the dynamics of the movement of human erythrocytes in ...vitro, in the channels of a microfluidic device, under the action of an OS inducer, tert-butyl hydroperoxide (tBH), and compared it with a cytological assessment of the transformation of cell membranes. OS induced the impairment of the control over the shape and volume of cells, the decrease in passage speed, and the occlusions of microchannels. The developed microfluidic device provided an assessment of the microreology of erythrocytes under the action of hydroperoxide. In the future, it is possible to use microfluidic analysis of erythrocytes in patients to assess the effect of xenobiotics on microrheology.
Monodisperse polymeric particles have great potential in biomedical and physical applications. Modern high-throughput droplet microfluidic technologies make it possible to produce monodisperse ...water-in-oil macroemulsions with desired properties. Polymerization in a macroemulsion transforms it to a suspension of microparticles. These particles may be viewed as containers for targeted delivery of drugs and also as bioink for 3D printing of tissues and organs. Conditions for formation of PEGDA and polyacrylamide particles using a microfluidic flow-focusing emulsion generator have been studied. Manufactured microparticles have been characterized by their geometrical sizes and mechanical properties. In addition, the diffusion escape of small molecules from microparticles has been studied using Rhodamine B fluorescent dye.
Nowadays isolation and sorting of biological objects plays an important role in life science and medicine. Magnetic sorting has big opportunities because of its advantages such as sufficient ...selectivity and bioinerticity. In this work we studied formation and handling of polyacrylamide microparticles doped with magnetic nanoparticles using droplet microfluidics. Using a flow focusing droplet generator magnetic nanoparticles were encapsulated to polyacrylamide droplets which were polymerized inside the device. It was shown that these magnetic droplets could be efficiently sorted by magnetic field in a microfluidic device.
Abstract
In this study, we developed a new model of neuronal cells plating into a developed neural network to study functional integration using microfluidic methods. The integration was modeled in a ...three-chamber microfluidic chip by growing two weakly coupled neuronal networks and enhancing its connectivity by plating new dissociated cells. The direction of connections was formed by the asymmetric design of the chip. Such technology can be used to develop a new type of scaffold to recover the modular structure of the network.
Polyethylene glycol diacrylate (PEGDA) microparticles (MPs) have a great potential in a wider range of applications including cell carriers, drug delivery agents, and multiplexing assays. Among ...existing MP fabricating methods, only the microfluidic droplet-based methods provide both morphology complexity and substance diversity. This work is concerned with the production PEGDA-based droplets with the photoinitiator (Irgacure 2959) in a flow-focusing microfluidic generator. The kinetics of MPs solidification in mineral oil with various common surfactants (Abil EM 180, Span 80) were investigated. Also, the Young's modulus of solid PEGDA were obtained.
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