Geobacillus thermoglucosidasius is a kind of Gram-positive facultative anaerobic bacteria. The fast growth rate under high temperature and less susceptibility to microbial contamination enable G. thermoglucosidasius to be a desirable producer of biofuels and high-value-added chemicals for the next-generation industrial biotechnology. However, compared with the classical model strain Escherichia coli, the applications of G. thermoglucosidasius are hampered by its low transformation efficiency. This study aimed at obtaining competent cells with high transformation efficiency through inactivating restriction enzymes, adding cell membrane inhibitors and cell wall weakening agents. The results showed that the electro-transformation efficiency achieved 1.2×10⁴ CFU/(μg DNA) by knocking out four genes encoding restriction enzymes. Adding a certain amount of tween 80, dl-threonine and glycine further increased the competent efficiency about 22.5, 44, and 334 times, respectively. The electro-transformation efficiency was enhanced to 4.6×10⁶ CFU/(μg DNA) under the optimized conditions, laying a foundation for genetic manipulation and metabolic engineering of G. thermoglucosidasius.
Electroporation ; Electroporation Therapies ; Bacillaceae ; Cell Membrane ; Escherichia coli/genetics*

In these years, the electrical technology is widely applied in the study of biomedical engineering. Using electroporation therapy (EPT) to treat tumor is associated with biomedical engineering, electrical new technology, computer technology and microelectronic technology, which is a new marginal subject. Many experts have studied the mechanism and clinical treatment of the cell membrane electroporation phenomenon under electrical fields. These researches have shown that the membrane electroporation can stimulate the transport and intake of various drugs, which improves the tumoricidal effect of these drugs. The researchers have also been exploring the phenomenon that irreversible electrical breakdown (IREB) of cell membrane under high electrical fields and steep pulses leads to the death of tumor.
Cell Membrane ; ultrastructure ; Electrochemotherapy ; Humans ; Neoplasms ; drug therapy ; Research ; trends

Pulsed electric fields (PEFs) with fixed frequency, width and gradually increased peak value of voltage was applied to 30 healthy rabbit liver tissues. The specific aims were to explore the feasibility of establishing a model of in vivo PEFs distribution in healthy rabbit liver tissues and to provide important references for clinical electrochemotherapy and for electrotransfer. Repeated experiment and self-comparison statistics design were implemented. The rabbit underwent the experiment under intravenous anesthesia and their liver tissues, after exposure to PEFs, were sent for HE staining. Necrotic borderline was visible 3 days after PEFs application, the necrotic shape of concentric circle was evident around the electrodes under optical microscope at lower voltage, as voltage increasing, two necroses in the shape of concentric circle gradually enlarged; nuclei with chromatin condensation, fragmentation and lysis alterations were seen in the middle region between the needles; concentric circles changed into ellipse fusiform and finally overlaped each other forming irregular necrosis contours. Cell cavitation and tissues ischemia were also observed within electric field. The shape of tissue necrosis from the experiment was noted to correlate with theoretic simulation of electric field distribution. Therefore, rabbit liver tissues can be a good carrier for in vivo modeling of electric field distribution when the lethal effects of PEFs in tissues are investigated. PEFs also show safety for the surrounding normal tissue while causing damage or injury to the target area therapeutically.
Animals ; Electricity ; adverse effects ; Electrochemotherapy ; Liver ; pathology ; Male ; Models, Theoretical ; Necrosis ; Rabbits

OBJECTIVETo estimate electroporation (EP) influence on malignant and normal cells.

METHODSTwo cell lines including human malignant melanoma (Me-45) and normal human gingival fibroblast (HGFs) were used. EP parameters were the following: 250, 1 000, 1 750, 2 500 V/cm; 50 µs by 5 impulses for every case. The viability of cells after EP was estimated by MTT assay. The ultrastructural analysis was observed by transmission electron microscope (Zeiss EM 900).

RESULTSIn the current study we observed the intracellular effect following EP on Me-45 and HGF cells. At the conditions applied, we did not observe any significant damage of mitochondrial activity in both cell lines treated by EP. Conversely, we showed that EP in some conditions can stimulate cells to proliferation. Some changes induced by EP were only visible in electron microscopy. In fibroblast cells we observed significant changes in lower parameters of EP (250 and 1 000 V/cm). After applying higher electric field intensities (2 500 V/cm) we detected many vacuoles, myelin-like bodies and swallowed endoplasmic reticulum. In melanoma cells such strong pathological modifications after EP were not observed, in comparison with control cells. The ultrastructure of both treated cell lines was changed according to the applied parameters of EP.

CONCLUSIONSWe can claim that EP conditions are cell line dependent. In terms of the intracellular morphology, human fibroblasts are more sensitive to electric field as compared with melanoma cells. Optimal conditions should be determined for each cell line. Summarizing our study, we can conclude that EP is not an invasive method for human normal and malignant cells. This technique can be safely applied in chemotherapy for delivering drugs into tumor cells.

Adult ; Cell Line ; Cell Line, Tumor ; Cell Shape ; Cell Survival ; Cells, Cultured ; Electrochemotherapy ; Electroporation ; Fibroblasts ; chemistry ; cytology ; Gingiva ; chemistry ; cytology ; Humans ; Male ; Melanoma ; chemistry ; physiopathology ; therapy

OBJECTIVETo evaluate the feasibility of electroporation-mediated transfection of recombinant plasmid to mandibular distraction area of rabbit in vivo.

METHODSNew-Zeland rabbit were employed. The mandible was distracted 3 days after operation at a rate of 0.8 mm per day for 7 days. The rabbits were randomly divided into 3 groups as group A (recombinant plasmid pIRES-VEGF165-EGFP), group B (recombinant plasmid plRES-VEGF165-EGFP) and group C (normal saline). The rabbits were sacrified at 3 hours, 1, 3, 7 and 14 d after injection respectively. The tissue at the distraction area was taken out for frozen section. The gene expression was assessed by the detection of expression of green fluorescence protein (GFP) using fluorescence microscope. The liver and kidney function test (ALT, AST, BUN, Scr) and the histological examination of heart, liver and kidney were also performed.

RESULTSGFP was seen in the distraction area in group A and group B 3 hours after injection, which increased at the 1st day, reached peak value at the 3rd day, decreased at the 7th day and was very lower at 14th day. The GFP expression was much stronger in group A than in group B. GFP was not expressed in group C. There was no statistical difference in the concentration of ALT, AST, BUN and Scr in serum of rabbits among the three groups.

CONCLUSIONSElectroporation-mediated transfection of recombinant plasmid can be expressed in the distraction area of rabbits, and there was no toxicity to the liver and kidney of rabbits. Electroporation could obviously improve transfection efficiency in vivo. It indicates that electroporation-mediated transfection of recombinant plasmid to distraction area tissue of rabbits is feasible.

Animals ; Electrochemotherapy ; Female ; Genetic Therapy ; Green Fluorescent Proteins ; genetics ; Male ; Mandible ; surgery ; Models, Animal ; Osteogenesis, Distraction ; Rabbits ; Vascular Endothelial Growth Factor A ; genetics

In this paper is reported a new approach for the treatment of sarcoma--electroporation therapy. Electroporation can accelerate pharmacal molecules into cytoplasm by transient electromagnetic pulses. We have utilized the phenomenon of electroporation treating the S-180 sarcomas in the hind legs of the Kunming mice by intratumoral injection of anti-tumor agent at low dose. From the experiment, we learned that this approach can bring about remarkable effect. The technical procedure is easy to do and easy to control. Especially, it is useful in curing the flat tumor and has little untoward side effect. It deserves to be recommended as a new approach to treating the tumor in clinics.
Animals ; Antineoplastic Agents ; therapeutic use ; Apoptosis ; drug effects ; Cell Line, Tumor ; Cyclophosphamide ; therapeutic use ; Electrochemotherapy ; methods ; Mice ; Neoplasm Transplantation ; Sarcoma 180 ; drug therapy ; pathology

Electrochemotherapy was instituted for sarcoma, and the tumor inhibitory ratio, curing ratio, vascular endothelial growth factor, microvessel density and mechanism were measured and analyzed. The results indicate that the curing ratio of electrochemotherapy for sarcoma is 84.6%. The present research provides experimental evidence for the security, mechanism and feasibility of electrochemotherapy in clinical practice.
Animals ; Antineoplastic Agents ; administration & dosage ; Bleomycin ; administration & dosage ; Drug Delivery Systems ; methods ; Electrochemotherapy ; methods ; Electroporation ; methods ; Mice ; Sarcoma 180 ; therapy ; Vascular Endothelial Growth Factor A ; analysis

OBJECTIVETo explore the therapeutic effect of electrochemical therapy(ECT) for high-flow vascular malformation in maxillofacial region.

METHODS38 patients with high-flow vascular malformation in maxillofacial region were treated with ECT. The voltage was 6-8 V, and the electric current was 80-100 mA. The electric quantity was 10-20 C/cm2.

RESULTSAmong the 38 cases, 32 cases received only one treatment, 5 cases received two treatments and one case received three treatments. The patients were followed up for 2 months to 3 years. The total effective rate was 100% with 55.26% of cure rate and 44.74% of partial cure rate. No hemolysis happened during follow-up period.

CONCLUSIONSECT is a simple and effective method for high-flow vascular malformation with less morbidity.

Adolescent ; Adult ; Child ; Child, Preschool ; Electrochemotherapy ; Facial Neoplasms ; drug therapy ; Female ; Follow-Up Studies ; Hemangioma ; drug therapy ; Humans ; Male ; Middle Aged ; Treatment Outcome ; Young Adult

OBJECTIVETo evaluate the efficacy and safety of electrochemotherapy in treating venous malformations.

METHODSElectrochemotherapy was applied to 665 patients with venous malformations of limbs and trunk, and 505 cases were followed up for half to 6 years. In this study, 228 male and 277 female patients were involved. Diagnosis was made by clinical manifestations and magnetic resonance imaging. The platinum electrodes were inserted into tumor through a trocar with plastic insulating cannula percutaneously and connected with the electrochemical therapeutic apparatus in anodes and cathodes separately. Then electricity was given. The treating voltage is 6-12 V and volume 100-180 mA, the total electricity used is in general 80-100 coulombs per 1.0 square centimeter of tumors' area. The treating time was usually from several dozen minutes to over 2 h depending on the size of the tumor. The severe cases which needed to be treated once again usually were operated after 6 months.

RESULTSThe primary efficacy end point was defined as an improvement of patients' symptoms and a reduction in size of tumor 6 months after treatment. Effects were divided into 4 grades, and the efficacy rate decreased from grade 1 to grade 4. The efficacy turned out that 30.1% (152/505) of patients was classified as grade 1; 46.3% (234/505) as grade 2; 19.0% (96/505) as grade 3 and 4.6% (23/505) as grade 4.

CONCLUSIONSElectrochemotherapy shows special superiorities in treating venous malformations. It might bring a confirmed clinical efficacy with the advantages of less injury, quick recovery, simple operation and less complications.

Adolescent ; Adult ; Child ; Child, Preschool ; Electrochemotherapy ; Female ; Humans ; Magnetic Resonance Imaging ; Male ; Middle Aged ; Neoplasms ; drug therapy ; pathology ; Retrospective Studies ; Vascular Malformations ; drug therapy

OBJECTIVE: To analyse the therapie effect of low-voltage electrochemotherapy (EP) on the head and neck cancer. METHOD: YCU-N861 human head and neck cancer cells were subcutaneously inoculated in the neck of nude mice to establish tumor-bearing models. Then the mice were treated with bleomycin (BLM, 20 microg per mouse), accompanied with electric stimulation (50 V or 100 V). RESULT: BLM treatment applied with EP can significantly inhibit the proliferation of tumor cells than BLM or EP treatment separately (P < 0.01). The tumor volume, the mitotic index (MI) and the labeling index (LI) of Ki67 in BLM+EP group were all obviously decreased than BLM group, EP group and control group (P < 0.01). CONCLUSION Low-voltage electrochemotherapy can offer promising method in the local treatment of head and neck cancer.
Animals ; Antimetabolites, Antineoplastic ; administration & dosage ; therapeutic use ; Bleomycin ; administration & dosage ; therapeutic use ; Cell Line, Tumor ; Electrochemotherapy ; Female ; Head and Neck Neoplasms ; drug therapy ; therapy ; Humans ; Mice ; Mice, Inbred BALB C ; Mice, Nude ; Neoplasm Transplantation