Molecular dynamics simulation of reversible electroporation with Martini force field.

BACKGROUND: After the discovery of membrane-reversible electroporation decades ago, the procedure has been used extensively in biology, biotechnology and medicine. The research on the basic mechanism has increasingly attracted attention. Although most research has focused on models that consider all atomic and molecular interactions and much atomic-level information can be obtained, the huge computational demand limits the models to simulations of only a few nanometers on the spatial scale and a few nanoseconds on the time scale. In order to more comprehensively study the reversible electroporation mechanism of phospholipid membrane on the nanoscale and at longer time intervals of up to 100 ns, we developed a dipalmitoylphosphatidylcholine (DPPC) phospholipid membrane model with the coarse-grained Martini force field. The model was tested by separately examining the morphology of the phospholipid membrane, the hydrophilic channel size, the distribution of the voltage potential on both sides of the membrane, and the movement of water molecules and ions during electroporation. RESULTS: The results showed that the process went through several stages: (1) the formation of the pore with defects originating on the surface. (2) The maintenance of the pore. The defects expanded to large pores and the size remains unchanged for several nanoseconds. (3) Pore healing stage due to self-assembly. Phospholipid membrane shrunk and the pore size decreased until completely closed. The pores were not circular in cross-section for most of the time and the potential difference across the membrane decreased dramatically after the pores formed, with almost no restoration of membrane integrity even when the pores started to close. CONCLUSIONS: The mechanism of the reversible electroporation process on the nanoscale level, including defects, expansion, stability, and pore closing stages on a longer time scale of up to 100 ns was demonstrated more comprehensively with the coarse-grained Martini force field, which took both the necessary molecular information and the calculation efficiency into account.

Avoiding the side effects of electric current pulse application to electroporated cells in disposable small volume cuvettes assures good cell survival.

BACKGROUND: The harmful side effects of electroporation to cells due to local changes in pH, the appearance of toxic electrode products, temperature increase, and the heterogeneity of the electric field acting on cells in the cuvettes used for electroporation were observed and discussed in several laboratories. If cells are subjected to weak electric fields for prolonged periods, for example in experiments on cell electrophoresis or galvanotaxis the same effects are seen. In these experiments investigators managed to reduce or eliminate the harmful side effects of electric current application. METHODS: For the experiments, disposable 20 µl cuvettes with two walls made of dialysis membranes were constructed and placed in a locally focused electric field at a considerable distance from the electrodes. Cuvettes were mounted into an apparatus for horizontal electrophoresis and the cells were subjected to direct current electric field (dcEF) pulses from a commercial pulse generator of exponentially declining pulses and from a custom-made generator of double and single rectangular pulses. RESULTS: More than 80% of the electroporated cells survived the dcEF pulses in both systems. Side effects related to electrodes were eliminated in both the flow through the dcEF and in the disposable cuvettes placed in the focused dcEFs. With a disposable cuvette system, we also confirmed the sensitization of cells to a dcEF using procaine by observing the loading of AT2 cells with calceine and using a square pulse generator, applying 50 ms single rectangular pulses. CONCLUSIONS: We suggest that the same methods of avoiding the side effects of electric current pulse application as in cell electrophoresis and galvanotaxis should also be used for electroporation. This conclusion was confirmed in our electroporation experiments performed in conditions assuring survival of over 80% of the electroporated cells. If the amplitude, duration, and shape of the dcEF pulse are known, then electroporation does not depend on the type of pulse generator. This knowledge of the characteristics of the pulse assures reproducibility of electroporation experiments using different equipment.

A microdosimetry analysis of reversible electroporation in scattered, overlapping, and cancerous cervical cells.

We present a numerical method for studying reversible electroporation on normal and cancerous cervical cells. This microdosimetry analysis builds on a unique approach for extracting contours of free and overlapping cervical cells in the cluster from the Extended Depth of Field (EDF) images. The algorithm used for extracting the contours is a joint optimization of multiple-level set function along with the Gaussian mixture model and Maximally Stable Extremal Regions. These contours are then exported to a multi-physics domain solver, where a variable frequency pulsed electric field is applied. The trans-Membrane voltage (TMV) developed across the cell membrane is computed using the Maxwell equation coupled with a statistical approach, employing the asymptotic Smoluchowski equation. The numerical model was validated by successful replication of existing experimental configurations that employed low-frequency uni-polar pulses on the overlapping cells to obtain reversible electroporation, wherein, several overlapping clumps of cervical cells were targeted. For high-frequency calculation, a combination of normal and cancerous cells is introduced to the computational domain. The cells are assumed to be dispersive and the Debye dispersion equation is used for further calculations. We also present the resulting strength-duration relationship for achieving the threshold value of electroporation between the normal and cancerous cervical cells due to their size and conductivity differences. The dye uptake modulation during the high-frequency electric field electroporation is further advocated by a mathematical model.

Evaluation of pulsed field ablation lesion characteristics using an in vitro vegetable model.

BACKGROUND: In vitro models to evaluate cardiac pulsed field ablation (PFA) have not been well established. We sought to create a standardized vegetable model and staining protocol for assessing unipolar PFA using a surface electrode. METHODS: We exposed potato slabs to unipolar PFA in a saline bath using a 3.5 mm electrode catheter and grounding pad connected to a custom-built high-voltage generator. Lesions were clearly visualized after staining with 2,3,5-triphenyltetrazolium chloride (TTC) using a timed protocol to reveal a necrotic center and a periphery of electroporated cells with intact mitochondria. RESULTS: Lesion volume increased linearly with increasing voltage and logarithmically with repetitive PFA applications. CONCLUSION: The findings observed in this vegetable model using a TTC staining protocol are consistent with findings observed with cardiomyocytes.

Engineering of Multivalent Membrane-Anchored DNA Frameworks for Precise Profiling of Variable Membrane Permeability During Reversible Electroporation.

Electroporation techniques have emerged as attractive tools for intracellular delivery, rendering promising prospects towards clinical therapies. Transient disruption of membrane permeability is the critical process for efficient electroporation-based cargo delivery. However, smart nanotools for precise characterization of transient membrane changes induced by strong electric pulses are extremely limited. Herein, multivalent membrane-anchored fluorescent nanoprobes (MMFNPs) that take advantages of flexible functionalization and spatial arrangement of DNA frameworks are developed for in situ evaluation of electric field-induced membrane permeability during reversible electroporation . Single-molecule fluorescence imaging techniques are adopted to precisely  verify the excellent analytical performance of the engineered MMFNPs. Benefited from tight membrane anchoring and sensitive adenosine triphosphate (ATP) profiling, varying degrees of membrane disturbances are visually exhibited under different intensities of the microsecond pulse electric field (µsPEF). Significantly, the dynamic process of membrane repair during reversible electroporation is well demonstrated via ATP fluctuations monitored by the designed MMFNPs. Furthermore, molecular dynamics (MD) simulations are performed for accurate verification of electroporation-driven dynamic cargo entry via membrane nanopores. This work provides an avenue for effectively capturing transient fluctuations of membrane permeability under external stimuli, offering valuable guidance for developing efficient and safe electroporation-driven delivery strategies for clinical diagnosis and therapeutics.

Dynamics of High-Density Unipolar Epicardial Electrograms During PFA.

BACKGROUND: Pulsed field ablation (PFA) is a novel nonthermal cardiac ablation technology based on irreversible electroporation (IRE). While areas of IRE lead to durable lesions, the surrounding regions, where reversible electroporation occurs, recover. The behavior of local electrograms in areas of different electroporation levels remains unknown. The goal of this study is to characterize electrogram dynamics after PFA in IRE and reversible electroporation areas. METHODS: A total of 6 domestic swine were used. PFA was applied in the epicardium of the right and left ventricles using a focal monopolar catheter. Additional radiofrequency ablations were performed. Epicardial unipolar electrograms were acquired at baseline and for 60 minutes post PFA/radiofrequency ablation using a high-density electrode matrix attached to the epicardium. Electrogram dynamics were analyzed in areas corresponding to different levels of electroporation. Acute lesion formation was assessed after 3 to 5 hours by triphenyl tetrazolium chloride staining. RESULTS: Electrogram analysis demonstrated a clear association between electrogram changes and the level of electroporation. Immediately after PFA, electrograms displayed the following: a significant decrease in R/S-wave amplitude; a large elevation of the ST-segment; and a large decrease in their |(dV/dt)|max. Marked changes in electrograms were observed beyond the lesion area. Thereafter, a gradual recovery was observed. The evolution of all the electrogram parameters throughout the 60 minutes after PFA was significantly different (P<0.05) between the IRE and reversible electroporation areas. Acute lesion staining showed significantly larger depth for PFA lesions compared with radiofrequency ablation. CONCLUSIONS: This study shows that unipolar electrograms can differentiate between reversible electroporation and IRE areas during the first 30 minutes post ablation. Differences after the first 30 minutes are less evident. Our findings could result useful for immediate lesion assessment after PFA and warrant further investigation.

Dispersive Modeling of Normal and Cancerous Cervical Cell Responses to Nanosecond Electric Fields in Reversible Electroporation Using a Drift-Step Rectifier Diode Generator.

This paper creates an approximate three-dimensional model for normal and cancerous cervical cells using image processing and computer-aided design (CAD) tools. The model is then exposed to low-frequency electric pulses to verify the work with experimental data. The transmembrane potential, pore density, and pore radius evolution are analyzed. This work adds a study of the electrodeformation of cells under an electric field to investigate cytoskeleton integrity. The Maxwell stress tensor is calculated for the dispersive bi-lipid layer plasma membrane. The solid displacement is calculated under electric stress to observe cytoskeleton integrity. After verifying the results with previous experiments, the cells are exposed to a nanosecond pulsed electric field. The nanosecond pulse is applied using a drift-step rectifier diode (DSRD)-based generator circuit. The cells' transmembrane voltage (TMV), pore density, pore radius evolution, displacement of the membrane under electric stress, and strain energy are calculated. A thermal analysis of the cells under a nanosecond pulse is also carried out to prove that it constitutes a non-thermal process. The results showed differences in normal and cancerous cell responses to electric pulses due to changes in morphology and differences in the cells' electrical and mechanical properties. This work is a model-driven microdosimetry method that could be used for diagnostic and therapeutic purposes.

The Enlargement of Ablation Area by Electrolytic Irreversible Electroporation (E-IRE) Using Pulsed Field with Bias DC Field.

Irreversible electroporation (IRE) by high-strength electric pulses is a biomedical technique that has been effectively used for minimally invasive tumor therapy while maintaining the functionality of adjacent important tissues, such as blood vessels and nerves. In general, pulse delivery using needle electrodes can create a reversible electroporation region beyond both the ablation area and the vicinity of the needle electrodes, limiting enlargement of the ablation area. Electrochemical therapy (EChT) can also be used to ablate a tumor near electrodes by electrolysis using a direct field with a constant current or voltage (DC field). Recently, reversible electroporated cells have been shown to be susceptible to electrolysis at relatively low doses. Reversible electroporation can also be combined with electrolysis for tissue ablation. Therefore, the objective of this study is to use electrolysis to remove the reversible electroporation area and thereby enlarge the ablation area in potato slices in vitro using a pulsed field with a bias DC field (constant voltage). We call this protocol electrolytic irreversible electroporation (E-IRE). The area over which the electrolytic effect induced a pH change was also measured. The results show that decreasing the pulse frequency using IRE alone is found to enlarge the ablation area. The ablation area generated by E-IRE is significantly larger than that generated by using IRE or EChT alone. The ablation area generated by E-IRE at 1 Hz is 109.5% larger than that generated by IRE, showing that the reversible electroporation region is transformed into an ablation region by electrolysis. The area with a pH change produced by E-IRE is larger than that produced by EChT alone. Decreasing the pulse frequency in the E-IRE protocol can further enlarge the ablation area. The results of this study are a preliminary indication that the E-IRE protocol can effectively enlarge the ablation area and enhance the efficacy of traditional IRE for use in ablating large tumors.

Laser Machined Fiber-based Microprobe: Application in Microscale Electroporation.

Microscale electroporation devices are mostly restricted to in vitro experiments (i.e., microchannel and microcapillary). Novel fiber-based microprobes can enable in vivo microscale electroporation and arbitrarily select the cell groups of interest to electroporate. We developed a flexible, fiber-based microscale electroporation device through a thermal drawing process and femtosecond laser micromachining techniques. The fiber consists of four copper electrodes (80 µm), one microfluidic channel (30 µm), and has an overall diameter of 400 µm. The dimensions of the exposed electrodes and channel were customizable through a delicate femtosecond laser setup. The feasibility of the fiber probe was validated through numerical simulations and in vitro experiments. Successful reversible and irreversible microscale electroporation was observed in a 3D collagen scaffold (seeded with U251 human glioma cells) using fluorescent staining. The ablation regions were estimated by performing the covariance error ellipse method and compared with the numerical simulations. The computational and experimental results of the working fiber-based microprobe suggest the feasibility of in vivo microscale electroporation in space-sensitive areas, such as the deep brain.

Scanning electrochemical microscope as a tool for the electroporation of living yeast cells.

In this study, a scanning electrochemical microscope (SECM) was for the first time adapted to perform the electroporation process of living yeast cells. We have demonstrated that relatively low voltage pulses of 1-2 V vs. Ag/AglCl,Cl-sat applied to gold-based ultramicroelectrode (Au-UME) are performing reversible electroporation of yeast cells immobilized on fluorine-doped tin oxide (FTO)/glass surface. SECM and electrochemical impedance spectroscopy (EIS) were used for the determination of quantitative electrochemical characteristics before and after the electroporation. The electrochemical impedance spectroscopy (EIS) illustrated significant electrochemical changes of electroporated yeast cells, while SECM feedback mode surface vertical scan current-distance curves showed that the diameter of the area affected by the electrical pulse is about 25 times larger than the diameter of the Au-UME used for the electroporation process. The results presented in this research open up a possibility to develop a targeted electroporation system which will affect only the selected area of tissue or some other cell-covered surface. Such model is promising for the selective treatment of selected cells in tissues and/or other sensitive biological systems while selecting the location and size of electroporated areas.

Reversible electroporation study of realistic normal and cancerous cervical cells model using avalanche transistor-based nano pulse generator.

In this paper, we study the reversible electroporation process on normal and cancerous cervical cells. The 2D contour of the cervical cells is extracted using image processing techniques from the Pap smear images. The conductivity change in the cancer cell model has been used to differentiate the effects of the high-frequency electric field on normal and cancerous cells. The cells' dielectric constant modulates when this high-frequency pulse is applied based on the Debye relaxation. To computationally visualize the effects of the electroporation on the cell membrane, the Smoluchowski equation is employed to estimate pore density, and Maxwell equations are used to determine the electric potential developed across the membrane of the cervical cell. The results demonstrate the suitability of this mathematical model for studying the response of normal and cancerous cells under electric stress. The electric field is supplied with the help of a realistic pulse generator which is designed on the principle of Marx circuit and avalanche transistor-based operations to produce a Gaussian pulse. The paper here uses a strength-duration curve to differentiate the electric field and time in nanoseconds required to electroporate normal and cancerous cells.

Clinical Phase I/II Study: Local Disease Control and Survival in Locally Advanced Pancreatic Cancer Treated with Electrochemotherapy.

OBJECTIVE: To assess local disease control rates (LDCR) and overall survival (OS) in locally advanced pancreatic cancer (LAPC) treated with electrochemotherapy (ECT). METHODS: Electrochemotherapy with bleomycin was performed in 25 LAPC patients who underwent baseline Magnetic Resonance Imaging (MRI) and/or Computed Tomography (CT) and Position Emission Tomography (PET) scans before ECT and 1 and 6 months post ECT. LDCR were assessed using Response Evaluation Criteria in Solid Tumors (RECIST 1.1) and Choi criteria. Needle electrodes with fixed linear (N-30-4B) or fixed hexagonal configurations (N-30-HG or I-40-HG or H-30-ST) or variable geometry (VGD1230 or VGD1240) (IGEA S.p.A., Carpi, Italy) were used to apply electric pulses. Pain evaluation was performed pre-ECT, after 1 month and after 6 months with ECT. Overall survival estimates were calculated by means of a Kaplan-Meier analysis. RESULTS: At 1 month after ECT, 76% of patients were in partial response (PR) and 20% in stable disease (SD). Six months after ECT, 44.0% patients were still in PR and 12.0% in SD. A LDCR of 56.0% was reached six months after ECT: 13 patients treated with fixed geometry had a LDCR of 46.1%, while for the 12 patients treated with variable geometry, the LDCR was 66.7%. The overall survival median value was 11.5 months: for patients treated with fixed geometry the OS was 6 months, while for patients treated with variable geometry it was 12 months. Electrochemotherapy was well-tolerated and abdominal pain was rapidly resolved. CONCLUSIONS: Electrochemotherapy obtained good results in terms of LDCR and OS in LAPC. Multiple needle insertion in a variable geometry configuration optimized by pre-treatment planning determined an increase in LDCR and OS compared to a fixed geometry configuration.

Transcatheter Intraarterial Perfusion MRI Approaches to Differentiate Reversibly Electroporated Penumbra From Irreversibly Electroporated Zones in Rabbit Liver.

RATIONALE AND OBJECTIVES: To investigate whether transcatheter intraarterial perfusion (TRIP) magnetic resonance imaging (MRI) can differentiate reversible electroporation (RE) zones from irreversible electroporation (IRE) zones immediately after IRE procedure in the rabbit liver. MATERIALS AND METHODS: All studies were approved by the institutional animal care and use committee and performed in accordance with institutional guidelines. A total of 13 healthy New Zealand White rabbits were used. After selective catheterization of the hepatic artery under X-ray fluoroscopy, we acquired TRIP-MRI at 20 minutes post-IRE using 3 mL of 5% intraarterial gadopentetate dimeglumine. Semi-quantitative (peak enhancement, PE; time to peak, TTP; wash-in slope, WIS; areas under the time-intensity curve, AUT, over 30, 60, 90, 120, 150, and 180 seconds after the initiation of enhancement) and quantitative (Ktrans, ve, and vp) TRIP-MRI parameters were calculated. The relationships between TRIP-MRI parameters and histological measurements and the differential ability of TRIP-MRI parameters was assessed. RESULTS: PE, AUT60, AUT90, AUT120, AUT150, AUT180, Ktrans, and ve were significantly higher in RE zones than in IRE zones (all P < 0.05), and AUC for these parameters ranged from 0.91(95% CI, 0.80, 1.00) to 0.99 (95% CI, 0.98, 1.00). There was no significant difference in AUC between any two parameters (Z, 0-1.47; P, 0.14-1.00). Hepatocyte apoptosis strongly correlated with PE, AUT60, AUT90, AUT120, AUT150, AUT180, Ktrans, and vp (the absolute value r, 0.6-0.7, all P < 0.0001). CONCLUSION: AUT150 or AUT180 could be a potential imaging biomarker to differentiate RE from IRE zones, and TRIP-MRI permits to differentiate RE from IRE zones immediately after IRE procedure in the rabbit liver.

Understanding the role of calcium-mediated cell death in high-frequency irreversible electroporation.

High-frequency irreversible electroporation (H-FIRE) is an emerging electroporation-based therapy used to ablate cancerous tissue. Treatment consists of delivering short, bipolar pulses (1-10µs) in a series of 80-100 bursts (1 burst/s, 100µs on-time). Reducing pulse duration leads to reduced treatment volumes compared to traditional IRE, therefore larger voltages must be applied to generate ablations comparable in size. We show that adjuvant calcium enhances ablation area in vitro for H-FIRE treatments of several pulse durations (1, 2, 5, 10µs). Furthermore, H-FIRE treatment using 10µs pulses delivered with 1mM CaCl2 results in cell death thresholds (771±129V/cm) comparable to IRE thresholds without calcium (698±103V/cm). Quantifying the reversible electroporation threshold revealed that CaCl2 enhances the permeabilization of cells compared to a NaCl control. Gene expression analysis determined that CaCl2 upregulates expression of eIFB5 and 60S ribosomal subunit genes while downregulating NOX1/4, leading to increased signaling in pathways that may cause necroptosis. The opposite was found for control treatment without CaCl2 suggesting cells experience an increase in pro survival signaling. Our study is the first to identify key genes and signaling pathways responsible for differences in cell response to H-FIRE treatment with and without calcium.

Nanosecond pulsed electrical fields enhance product recovery in plant cell fermentation.

The potential of pharmacologically active secondary plant metabolites is limited by the low yield from often rare plants, and the lack of economically feasible chemical synthesis of these complex compounds. Plant cell fermentation offers an alternative strategy to overcome these constraints. However, the efficiency of this approach is limited by intracellular sequestration of the products, such that continuous bioprocessing is not possible. As a precondition for such a, more attractive, continuous process, it is of great importance to stimulate the export of the product into the medium without impairing viability and, thus, the productivity of the cells. Using nicotine alkaloids of tobacco as a case study, an alternative strategy is explored, where nanosecond pulsed electric fields (nsPEFs) are applied for the efficient downstream recovery of the products. To maintain cell viability and allow for the further use of biomass, cells were exposed to strong (1-20 kV·cm-1), but very short (10-100 ns) electric pulses, which leads to a temporary permeabilisation of cell membranes. Using two transgenic cell lines, where two key genes involved in the metabolism of the anti-Alzheimer compound nornicotine were overexpressed, we could show that this nsPEF treatment improved the partitioning of some nicotine alkaloids to the culture medium without impairing viability, nor the synthesis of alkaloids. However, this release was only partial and did not work for nornicotine. Thus, nsPEFs produced a fractionation of alkaloids. We explain this electrofractionation by a working model considering the differential intracellular compartmentalization of nicotineic alkaloids.

Impact of the number of electric pulses on cell electrochemotherapy in vitro: Limits of linearity and saturation.

In this study the evolution in the efficiency of electrochemotherapy (reversible electroporation) with pulse number was assessed in vitro. Experiments were performed using 100 µs pulses at different electric field intensities and the chemotherapeutic agent bleomycin. Additionally, electrical impedance spectroscopy measurements were used as a different method to study in real time the changes produced on cells with pulse number during trains of consecutive pulses. Our results show that the relation between pulse number and the observed outcome is complex and difficult to fully characterize. This relation can display a highly linear behaviour up to a certain number of pulses and/or field intensity applied. However, the relation between the number of pulses and the observed outcome always evolves to a saturation or at least a reduction in the electric field effects that is displayed when either electric field intensity or pulse number are increased. An exponential model was found to best describe this relation within the range of experimental conditions considered. Electrical impedance measurements confirmed the results and gave a more precise quantification of this dependence. The study highlights the importance that pulse number has in the electrochemotherapy protocols and establishes some limits in the use of this parameter.

Electrochemotherapy as a New Modality in Interventional Oncology: A Review.

Electroporation is a well-known phenomenon that occurs at the cell membrane when cells are exposed to high-intensity electric pulses. Depending on electric pulse amplitude and number of pulses, applied electroporation can be reversible with membrane permeability recovery or irreversible. Reversible electroporation is used to introduce drugs or genetic material into the cell without affecting cell viability. Electrochemotherapy refers to a combined treatment: electroporation and drug injection to enhance its cytotoxic effect up to 1000-fold for bleomycin. Since several years, electrochemotherapy is gaining popularity as minimally invasive oncologic treatment. The adoption of electrochemotherapy procedure in interventional oncology poses several unsolved questions, since suitable tumor histology and size as well as therapeutic efficacy still needs to be deepen. Electrochemotherapy is usually applied in palliative settings for the treatment of patients with unresectable tumors to relieve pain and ameliorate quality of life. In most cases, it is used in the treatment of advanced stages of neoplasia when radical surgical treatment is not possible (eg, due to lesion location, size, and/or number). Further, electrochemotherapy allows treating tumor nodules in the proximity of important structures like vessels and nerves as the treatment does not involve tissue heating. Overall, the safety profile of electrochemotherapy is favorable. Most of the observed adverse events are local and transient, moderate local pain, erythema, edema, and muscle contractions during electroporation. The aim of this article is to review the recent published clinical experiences of electrochemotherapy use in deep-seated tumors with particular focus on liver cases. The principle of electrochemotherapy as well as the application to cutaneous metastases is briefly described. A short insight in the treatment of bone metastases, unresectable pancreas cancer, and soft tissue sarcoma will be given. Preclinical and clinical studies on treatment efficacy with electrochemotherapy of hepatic lesions and safety of the procedure adopted are discussed.

Single exponential decay waveform; a synergistic combination of electroporation and electrolysis (E2) for tissue ablation.

BACKGROUND: Electrolytic ablation and electroporation based ablation are minimally invasive, non-thermal surgical technologies that employ electrical currents and electric fields to ablate undesirable cells in a volume of tissue. In this study, we explore the attributes of a new tissue ablation technology that simultaneously delivers a synergistic combination of electroporation and electrolysis (E2). METHOD: A new device that delivers a controlled dose of electroporation field and electrolysis currents in the form of a single exponential decay waveform (EDW) was applied to the pig liver, and the effect of various parameters on the extent of tissue ablation was examined with histology. RESULTS: Histological analysis shows that E2 delivered as EDW can produce tissue ablation in volumes of clinical significance, using electrical and temporal parameters which, if used in electroporation or electrolysis separately, cannot ablate the tissue. DISCUSSION: The E2 combination has advantages over the three basic technologies of non-thermal ablation: electrolytic ablation, electrochemical ablation (reversible electroporation with injection of drugs) and irreversible electroporation. E2 ablates clinically relevant volumes of tissue in a shorter period of time than electrolysis and electroporation, without the need to inject drugs as in reversible electroporation or use paralyzing anesthesia as in irreversible electroporation.

Early radiological assessment of locally advanced pancreatic cancer treated with electrochemotherapy.

AIM: To report early imaging assessment of ablated area post electrochemotherapy (ECT) in patients with locally advanced pancreatic cancer (LAPC). METHODS: ECT was performed in 19 LAPC patients enrolled in an approved ongoing clinical phase I/II study. Before and after ECT, 18 patients underwent computed tomography (CT) scan, 11 patients underwent morphological and functional magnetic resonance (MR) scan (dynamic contrast enhanced-MRI) calculating wash-in slope (WIS) and wash-out slope (WOS); diffusion weighted imaging calculating pseudo-diffusivity (Dp), perfusion fraction (fp) and tissue diffusivity (Dt); 10 patients underwent positron emission tomography (PET). Response evaluation criteria in solid tumour (RECIST) on MR and CT were used to assess tumour therapy response. Choi on CT, PET response criteria in solid tumors (PERCIST) on PET and functional parameters on MR were used to evaluate treatment response. RESULTS: For each patient no significant reduction was measurable by CT and MR using RECIST. According Choi criteria a partial response was obtained in 18/18 (100.0%) patients. According PERCIST criteria 6/10 (60.0%) patients showed a partial response, 3/10 (30.0%) stable disease and 1/10 (10.0%) progression disease. Moreover, using functional MR parameters, a significant reduction of viable tumour after ECT can be observed. According ΔWIS and ΔWOS 9/11 (81.8%) patients exhibited a partial response and 2/11 (18.2%) stable disease; 8/11 (72.7%) patients were considered in partial response by ΔDp evaluation and 3/11 (27.3%) in stable disease; according ΔDt 7/11 (63.6%) patients showed a partial response, 1/11 (9.1%) showed progression of disease and 3/11 (27.3%) were stable. Perfusion fraction fp showed a significant reduction after ECT only in four patients. No significant difference was observed after ECT in signal intensity of T1-weighted images and T2-weighted images, and in equilibrium-phase of contrast study, according to χ2 test was observed. A good correlation was reported between ΔHounsfield unit and Δmaximum standardized uptake value and between Δfp and ΔWOS, with a significant statistically difference (P < 0.05) using Spearman correlation coefficient. CONCLUSION: Perfusion and diffusion MR derived parameters, Choi, PERCIST criteria are more performant than morphological MR and CT criteria to assess ECT treatment response.

Tissue Ablation by a Synergistic Combination of Electroporation and Electrolysis Delivered by a Single Pulse.

A synergistic combination of electroporation and electrolysis (SEE) has been found with distinct advantages over tissue ablation by electrolysis or electroporation alone. Minimally invasive tissue ablation by electrolysis uses a low magnitude direct electric current to produce a lesion due to the creation of chemical products that result in cell death. Electroporation creates permeabilizations in the cell membrane which may lead to loss of cell homeostasis and cell death. When these two modes of tissue ablation are combined, a more effective method of cell death is achieved, likely due to the ability of electrolytic products to access the cell interior through the permeabilized cell membrane. Here, a new method of achieving SEE tissue ablation is obtained through the application of a single exponential decay pulse. This parametric study explores the mechanisms of damage as a function of the initial electric field and amount of delivered charge. It is seen that treatment parameters can dictate the mode of tissue ablation, either by SEE or by irreversible electroporation alone.