Gene transfer and drug delivery with electric pulse generators.

Electroporation is a process that involves a series of intense electric pulses used to overcome the barrier of the cell membrane. The application of an external electric field may result in reversible breakdown of cell membrane. This transient state helps to introduce cells with a variety of different molecules, such as DNA and drug molecules, and is known as the fundamental mechanisms of gene transfer and drug delivery with electric pulses.

[Effects of electric pulses on liver cancer cells: apoptosis induction and decrease of mitochondrial transmembrane potential].

In order to investigate the effects of electric pulses on cancer cells, we carried out the experiments with exposing HepG2 and L02 to electric pulses (1 kV/cm, l00 micros, 1 Hz) for different lengths of time (8 s, 15 s, 30 s, 60 s). Annexin V-FITC Kit and Flow cytometry were used to study the apoptosis of treated cells. The results showed that the electric pulses of 1 kV/cm, l00 micros, 1 Hz for 8 s could not induce tumor cells apoptosis.

[Ca(2+) is an important mediator of nanosecond steep pulse-induced apoptosis in human ovarian cancer SKOV3 cells].

Objective: To explore the role of Ca(2+) in nanosecond steep pulse (NSP)-induced apoptosis of human ovarian carcinoma cell line SKOV3 in vitro.

Methods: The early apoptotic rate of SKOV3 cells treated with NSP was detected by Annexin V/PI double staining and flow cytometry. MTT assay was used to detect the viability of the cells pretreated with BAPTA-AM (0, 25, 50 and 100 µmol/L) chelation for 1 h to increase the intracellular free Ca(2+) prior to NSP exposure, and the cell morphological changes and caspase 12 expression were detected using Hoechst 33342 staining and Western blotting, respectively.

[Analysis of frequency-domain and window effect for cellular inner and outer membranes subjected to pulsatile electric field].

Based on multi-layer dielectric model of spherical biological cell, a simulating method of frequency characteristics of inner and outer membranes is presented in this paper. Frequency-domain analysis showed that inner and outer membranes subjected to pulsed electric field exhibit band-pass and low-pass filter characteristics, respectively. A calculating method of the transmembrane potential of inner and outer membranes induced by time-varying electric field was introduced, and the window effect between electric field and transmembrane potential was also analyzed.

[Focusing properties of picosecond electric pulses in non-invasive cancer treatment].

In the light of optical theory, we advanc an ultra-wideband impulse radiating antenna (IRA) which is composed of an ellipsoidal reflector and a cone radiator. The high-intensity ultra-short electric pulses radiated by IRA can be transferred into the deep target in tissue non-invasively and be focused effectively. With the focused picosecond electric pulses, the organelles (mitochondria) transmembrane potential shall change to collapse under which the tumor cells will be targetly induced to apoptosis, so the method of non-invasive treatment of tumors would be achieved.

Effect of actin cytoskeleton disruption on electric pulse-induced apoptosis and electroporation in tumour cells.

Electric pulses are known to affect the outer membrane and intracellular structures of tumour cells. By applying electrical pulses of 450 ns duration with electric field intensity of 8 kV/cm to HepG2 cells for 30 s, electric pulse-induced changes in the integrity of the plasma membrane, apoptosis, viability and mitochondrial transmembrane potential were investigated. Results demonstrated that electric pulses induced cell apoptosis and necrosis accompanied with the decrease of mitochondrial transmembrane potential and the formation of pores in the membrane.

[The proliferating ability inhibited by energy controllable steep pulse in breast cancer cell line and its mechanism].

This was a study aimed to observe the proliferating ability inhibited by energy controllable steep pulse (ECSP) and to detect the expression of gene with relation to the proliferating ability of the tumor in breast cancer cell line; the possible mechanisms were also addressed. Human breast cancer cell line MDA-MB-231 was treated with ECSP; the apoptosis and the expression of tumor suppressor gene--Rb genes and E2F1 genes in ECSP group and control group were detected by TUNEL staining and Reverse Transcripitional PCR respectively. ECSP was found to inhibit the proliferating ability of breast cancer cells markedly, the cell amount in ECSP group decreased and the TUNEL positive cells increased obviously, compared to control; 24 hours after treatment the expression of Rb genes mRNA increased, whereas the expression of E2F1 mRNA decreased.

Apoptosis induction with electric pulses - a new approach to cancer therapy with drug free.

Electrical pulses have been widely used in biomedical fields, whose applications depend on the parameters such as durations and electric intensity. Conventional electroporation (0.1-1kV/cm, 100micros) has been used in cell fusion, transfection and electrochemotherapy.

[Experimental study on effects of energy controllable steep pulses on cytoskeleton of human ovarian cancer cells SKOV3].

The aim of our study was to determine the effects of energy controllable steep pulse (ECSP) on the cytoskeleton of human ovarian cancer cells SKOV3. SKOV3 cells were divided into five groups under ECSP treatment with different parameters (frequency, pulse duration, peak value of voltage). The positive control group included SKOV3 cells treated with volchicine; the negative control group included SKOV3 cells subjected to sham-lightning stroke.

The effect of high frequency steep pulsed electric fields on in vitro and in vivo antitumor efficiency of ovarian cancer cell line skov3 and potential use in electrochemotherapy.

Background: Patients received electrochemotherapy often associated with unpleasant sensations mainly result from low-frequency electric pulse induced muscle contractions. Increasing the repetition frequency of electric pulse can reduce unpleasant sensations. However, due to the specificity of SPEF, frequency related antitumor efficiency need to be further clarified.

Experiment and mechanism research of SKOV3 cancer cell apoptosis induced by nanosecond pulsed electric field.

This paper studies the apoptosis of human ovarian carcinoma cell Line (SKOV3) induced by the nanosecond pulsed electric field (10kV/cm, 100ns, 1 Hz) and its effect on intracellular calcium concentration ([Ca2+]i). These cells were doubly marked by Annexin V-FITC/PI, and the apoptosis rate was analyzed with flow cytometry. After AO/EB staining the morphological changes were observed under fluorescent microscope, and their ultrastructural changes were observed under scanning electron microscope (SEM).

Study on time-frequency characteristics of cellular transmenbrane potentials based on equivalent circuit model.

An equivalent circuit model of spherical biological cell subjected to pulsed electric field (PEF) is introduced, on the basis of multilayer dielectric model and complex-domain analysis. The transfer functions of inner and outer membranes are also presented. By introducing the time-domain solutions of transmembrane potentials induced by PEF, the relationship between electric field parameters and transmembrane potentials is discussed.

[Research progress of nanosecond pulsed electric field applied to intracellular electromanipulation].

In recent years, many experts have done some researches on experiment and mechanism of intracellular electromanipulation (IEM) under nanosecond pulsed electric field (nsPEF). The experiment results have shown that nsPEF could not induce electroporation of cell membrane, but could induce intracellular effects such as apoptosis, calcium release, enhancement of gene expression, and fragmentation of DNA and chromosome. In order to account for the phenomenon, researchers believe that when the pulse width of the pulsed electric field is larger than the charging time of plasma membrane, the pulsed electric field mainly targets on the outer membrane of cell; and that the effect of the pulsed electric field on nucleus and nuclear membrane increases with the decrease of the pulse width.

[The study of apoptosis and mechanism of cells exposed to steep pulse].

This experiment was designed to study the apoptosis and related mechanism of adherent liver tumor cells (SMMC-7721) and adherent normal liver cells (HL-7702) when they were exposed to the steep pulse generated by the steep pulse apparatus for tumor treatment. The results showed that the steep pulse of 200 V could induce tumor cells apoptosis. The tumor cells presented with their apoptosis when they were exposed to the steep pulse from 200 V to 250 V.

Study of transmembrane potentials on cellular inner and outer membrane--frequency response model and its filter characteristic simulation.

Based on the multilayer dielectric model for a spherical cell, a frequency response model of transmembrane potentials on cellular inner and outer membranes is established with a simulating method. The simulating results indicate that transmembrane potential on the inner membrane shows first-order bandpass filter characteristic, while transmembrane potential on the outer membrane shows first-order low-pass filter characteristic approximately. It could be found that the transmembrane potential on the inner membrane is greater than that on the outer membrane, and can keep a higher value in the range from a center frequency to an upper cutoff frequency, which is desirable to induce intracellular electromanipulation.

Frequency response model and simulation of transmembrane potentials on cellular inner and outer membranes.

In order to calculate transmembrane potentials of both inner and outer membranes more accurately, this paper presented a multi-shelled dielectric model for spherical cell according to the structure of cell, because the influence of organelles such as nuclear on the electric field distribution should be account for. Based on this model, the general method for analyzing the frequency response of the transmembrane potential on cellular inner and outer membrane is proposed. From the simulation and analysis of the frequency response, it can be drawn that the frequency response of the transmembrane potential on cellular outer membrane shows first-order low-pass filter characteristic, while that on inner membrane can be treated as a first-order band-pass filter.

Effects of steep pulsed electric fields (SPEF) on mitochondrial transmembrane potential of human liver cancer cell.

Mitochondrial plays an important role in apoptosis, and measuring the change of mitochondrial transmembrane potential (delta psi m) is a useful method for apoptosis. Exposing human liver cancer cell SMMC-7721 dyed with Rhodamine 123 to steep pulsed electric fields (SPEF), this paper observed the real-time change of delta psi m using laser scanning confocal microscope (LSCM) to study the apoptosis effect of SPEF. The experiment results showed that delta psi m decreased gradually in process of exposing to SPEF, SPEF with higher voltage (600V) and shorter width (100ns) could cause a quicker decrease than SPEF with lower voltage (200V) and longer width (1.

Transcutaneous battery recharging by volume conduction and its circuit modeling.

Many implantable devices require large capacity batteries implanted in the body. Transcutaneous battery recharging can effectively maintain the longevity of these implants. Based on this consideration we have developed a transcutaneous battery recharging circuit unit which takes advantages of skin volume conduction.

[An experimental study of energy controllable steep pulse in the treatment of rat with subcutaneous transplantive tumor].

This experimental study was designed to investigate the effects and the expressions of microvessel density (MVD), vascular endothelial growth factor (VEGF) on the transplanted tumor in the rat model with Walker-256 after energy controllable steep pulse(ECSP). The experiment revealed that the steep pulse electrical field has better effect on tumor, compared with the control. The positive cell staining intensity of VEGF in the control group was significantly higher than that in ECSP group (P < 0.

[Effect of steep pulsed electric fields on the immune response of tumor-bearing Wistar mice].

This study sought to evaluate the effect of steep pulsed electric fields (SPEFs) on the immune response of Wistar mice inoculated with Walker256 sarcoma. Thirty mice were randomly divided into three groups: control group (group A, inoculated with Walker256 sarcoma, not treated), treatment group (group B, inoculated with Walker256 sarcoma, treated by SPEFs), and normal control group (group C, inoculated with normal saline, not treated). Tumor size was measured before and every 3 days after treatment by vernier caliper.

[Biological effects and their applications in medicine of pulsed electric fields].

Pulsed electric fields can induce various kinds of biological effects that are essentially different from the normal electric fields, especially the interactions of Nanosecond Pulsed electric field (nsPEF) with cells. The biological effects of different pulsed electric fields on cell membranes, cytoplasmic matrixes, cell growth are introduced in this paper. Based on these effects, some applications of pulsed electric fields in cancer therapy, gene therapy, and delivery of drugs are reviewed in details.

Lethal Effects of Steep Pulsed Electric Field (SPEF) to Target Lymphatic Capillaries in VX2Implanted Breast Cancer of Rabbits.

This paper discussed the lethal effects of Steep Pulsed Electric Field (SPEF) to target lymphatic capillaries in VX2implanted breast cancer of rabbits using methylene blue dye injection staining, 5'-AMP-ALPase enzyme histochemical double staining, electron microscopy and VEGF-C observations. The result of methylene blue dye perfusion showed that the edge of tumor treated with SPEF was indistinct, no circularity or liner structure. Enzyme histochemical staining examination displayed that no positive staining was found in SPEF-target region of breast tumor, and lymphatic capillaries presented piecemeal structure in the edge of target region.

Analysis of Transmembrane Potentials Induced by Pulsed Electric Field with Different Durations Based on Five-shelled Dielectric Model of Cell.

In order to account for the influence of subcellular organelles on transmembrane potential, five-shelled dielectric model of spherical cell was developed in this paper. And the calculation method of transmembrane potential induced by any pulsed electric field (PEF) was put forward. Thus the effects of different pulse duration on transmembrane potentials of cell membrane and nuclear membrane were also analyzed.

Vascular effect and immunity effect of steep pulse electric field on Walker 256-bearing Wistar mice.

The vascular effect and immunity effect of SPEF on Wistar mice were studied, and the possible treatment mechanisms and clinical trial methods of SPEF were discussed. We tested lymphocyte proliferation ratio and activity of NK cell, and the contents of IFN-alpha and IL-4 in the serum lymphocyte were inspected with ELISA method. Moreover, macrophage were collected, the content of TNF-alpha excreted by macrophage were inspected with ELISA method, too.

Lethal and inhibitory effects of steep pulsed electric field on tumor-bearing BALB/c mice.

Exposing cancer cell to steep pulsed electric field (SPEF) with appropriate dosage, the cell may die due to irreversible electrical breakdown (IREB). 48 BALB/c mice inoculated with U/sub 14/ cervical cancer cell were treated with SPEF to study the lethal and inhibitory effects of SPEF on them. They were randomly divided into test group (26) and control group (22), and then four experiments were carried out.