Nondestructive introduction of genes, proteins, and small molecules into mammalian cells with high efficiency is a challenging, yet critical, process. Here we demonstrate a simple nanoelectroporation ...platform to achieve highly efficient molecular delivery and high transfection yields with excellent uniformity and cell viability. The system is built on alumina nanostraws extending from a track-etched membrane, forming an array of hollow nanowires connected to an underlying microfluidic channel. Cellular engulfment of the nanostraws provides an intimate contact, significantly reducing the necessary electroporation voltage and increasing homogeneity over a large area. Biomolecule delivery is achieved by diffusion through the nanostraws and enhanced by electrophoresis during pulsing. The system was demonstrated to offer excellent spatial, temporal, and dose control for delivery, as well as providing high-yield cotransfection and sequential transfection.
This study introduces a new method of targeting acidosis (low pH) within the tumor microenvironment (TME) through the use of cathodic electrochemical reactions (CER). Low pH is oncogenic by ...supporting immunosuppression. Electrochemical reactions create local pH effects when a current passes through an electrolytic substrate such as biological tissue. Electrolysis has been used with electroporation (destabilization of the lipid bilayer via an applied electric potential) to increase cell death areas. However, the regulated increase of pH through only the cathode electrode has been ignored as a possible method to alleviate TME acidosis, which could provide substantial immunotherapeutic benefits. Here, we show through
ex vivo
modeling that CERs can intentionally elevate pH to an anti-tumor level and that increased alkalinity promotes activation of naïve macrophages. This study shows the potential of CERs to improve acidity within the TME and that it has the potential to be paired with existing electric field-based cancer therapies or as a stand-alone therapy.
Cell electroporation is an important cell manipulation technology to artificially transfer specific extracellular components into cells. However, the consistency of substance transport during the ...electroporation process is still an issue due to the wide size distribution of the natural cells. In this study, a cell electroporation microfluidic chip based on a microtrap array is proposed. The microtrap structure was optimized for single-cell capture and electric field focusing. The effects of the cell size on the cell electroporation in the microchip were investigated through simulation and experiment methods using the giant unilamellar vesicle as the simplified cell model, and a numerical model of a uniform electric field was used as a comparison. Compared with the uniform electric field, a lower threshold electric field is required to induce electroporation and produces a higher transmembrane voltage on the cell under a specific electric field in the microchip, showing an improvement in cell viability and electroporation efficiency. The larger perforated area produced on the cells in the microchip under a specific electric field allows a higher substance transfer efficiency, and the electroporation results are less affected by the cell size, which is beneficial for improving substance transfer consistency. Furthermore, the relative perforation area increases with the decrease of the cell diameter in the microchip, which is exactly opposite to that in a uniform electric field. By manipulating the electric field applied to the microtrap individually, a consistent proportion of substance transfer during electroporation of cells with different sizes can be achieved.
In this paper, a new application of Piezoelectric Transformer (PT)-based power converters to generate high-voltage (HV) bipolar pulses for medical electroporation therapy is proposed. In particular, ...PT-based power conversion is investigated as an alternative to magnetics-based approaches of generating HV from a relatively low-voltage (LV) input source for application in electroporation therapy. The detailed PT-based system design and selection of wide bandgap semiconductor switches such as GaN FETs, high-voltage SiC diodes and SiC MOSFETs, as well as simulation results to demonstrate proof-of-concept using LTSpice are presented. Preliminary experimental results of the PT-based capacitor charger are shown, and work is ongoing to develop a complete hardware prototype of the proposed HV pulse generator.
In this paper, a theoretical study on the sensitivity of the nuclear electroporation model to real values is developed. This work has two main objectives. The first one is to study the sensitivity of ...a nuclear electroporation model to pulses of the order of ns. The second objective is to provide a guide for cell membrane and nuclear envelope electroporation for nanosecond pulses applications. A bibliographic review is performed to gather data used in experiments and other theoretical studies, and this data is used within an optimization routine to find the electroporation threshold as a function of stimulation pulse parameters. The nuclear electroporation threshold is plotted, and the model sensitivity is calculated and compared. The model is more sensitive to nuclear electrical properties, although some cellular parameters show a slight, but not negligible, influence. These results are summarized in two graphs that serve as an electroporation guide for ns pulses.
Genetic engineering in the rat has been revolutionized by the development of CRISPR-based genome editing tools. Conventional methods for inserting genome editing elements such as CRISPR/Cas9 reagents ...into rat zygotes include cytoplasmic or pronuclear microinjections. These techniques are labor-intensive, require specialized micromanipulator equipment, and are technically challenging. Here, we describe a simple and effective method for zygote electroporation in which CRISPR/Cas9 reagents are introduced into rat zygotes via pores produced by precise electrical pulses applied to the cells. Zygote electroporation allows for high-throughput efficient genome editing in rat embryos.
Irreversible electroporation has raised great interest in the past decade as a means of destroying cancers in a way that does not involve heat. Irreversible electroporation is a novel ablation ...technology that uses short high-voltage electrical pulses to enhance the permeability of tumor cell membranes and generate irreversible nano-sized structural defects or pores, thus leading to cell death. Irreversible electroporation has many advantages over thermal therapies due to its nonthermal mechanism: (1) reduced risk of injury to surrounding organs and (2) no “heat-sink” effect due to nearby blood vessels. However, so far, it has been difficult for irreversible electroporation to completely ablate large tumors (eg, >3 cm in diameter). In order to overcome this problem, many preclinical and clinical studies have been performed to improve the efficacy of IRE in the treatment of large size of tumors through a chemical perspective. Due to the distribution of electric field, irreversible electroporation region, reversible electroporation region, and intact region can be found in the treatment of irreversible electroporation. Thus, 2 types of chemical enhancements of irreversible electroporation were discussed in the article, such as the reversible electroporation region enhanced and the irreversible electroporation region enhanced. Specifically, the state-of-the-art results regarding the following approaches that have the potential to be used in the enhancement of irreversible electroporation were systematically reviewed in the article, including (1) combination with cytotoxic drugs, (2) calcium electroporation, (3) modification of cell membrane, and (4) modification of the tumor cell microenvironment. In the end, we concluded with 4 issues that should be addressed in the future for improving irreversible electroporation further in a chemical way.
•Physical NW-EP and chemical Cl2 were compared to remove culturable and viable cells.•NW-EP had ∼3–5 times lower energy consumption for culturable cell removal than Cl2.•NW-EP inhibited the formation ...of VBNC cells via destroying cell wall and membrane.•Osmotic pressure caused gradual inactivation of VBNC cells with cell wall damage.•NW-EP showed excellent adaptability to control VBNC cells in DI, tap and lake waters.
The induction of viable but nonculturable (VBNC) bacteria with cellular integrity and low metabolic activity by chemical disinfection causes a significant underestimation of potential microbiological risks in drinking water. Herein, a physical Co3O4 nanowire-assisted electroporation (NW-EP) was developed to induce cell damage via the locally enhanced electric field over nanowire tips, potentially achieving effective inhibition of VBNC cells as compared with chemical chlorination (Cl2). NW-EP enabled over 5-log removal of culturable cell for various G+/G- bacteria under voltage of 1.0 V and hydraulic retention time of 180 s, and with ∼3–6 times lower energy consumption than Cl2. NW-EP also achieved much higher removals (∼84.6 % and 89.5 %) of viable Bacillus cereus (G+) and Acinetobacter schindleri (G-) via generating unrecoverable pores on cell wall and reversible/irreversible pores on cell membrane than Cl2 (∼28.6 % and 41.1 %) with insignificant cell damage. The residual VBNC bacteria with cell wall damage and membrane pore resealing exhibited gradual inactivation by osmotic stress, leading to ∼99.8 % cell inactivation after 24 h storage (∼59.4 % for Cl2). Characterizations of cell membrane integrity and cell morphology revealed that osmotic stress promoted cell membrane damage for the gradual inactivation of VBNC cells during storage. The excellent adaptability of NW-EP for controlling VBNC cells in DI, tap and lake waters suggested its promising application potentials for drinking water, such as design of an external device on household taps.
Display omitted
PDF
