Analysis On The Mechanism Of Cell Electropration And Its Applications In Tumor Treatment

Cell electroporation refers to a biophysical process that transient pores appeare on the lipid bilayer of a cell due to the radiation of transient electrical field, which leads to the instantly increased permeability of the cell membrane. When the cell eletroporation comes forth, many large-size molecules such as hydrophilic molecules, virosomes, DNA, protein and drug particles are able to enter the cell through the membrane although they are unable to enter under normal conditions.Employing the technique of the cell electroporation for tumor treatment is an important application in various utilizations of cell electroporation. The method, which uses electric pulses in conjunction with chemotherapy drugs in tumor treatment, is named electrochemotherapy (ECT). It produces pores in the membranes of tumor cells by using electric pulses, and so the permeability of tumor cells goes up, which boosts drugs to enter cells. Hence, the toxicity of drugs to tumor cells is enhanced and the therapeutic effects of drugs can be improved.Various factors such as the intensity and last time of the pulse, the size of cells, the types of cells, the suspension, surrounding tissues and etc., can affect the cell electroporation. Therefore, experiments are usually necessary to determine the optimal factors when cell electroporation is used for different cells or for treating different tumors. Nowadays, neither the mechanism of the cell electroporation and the electrochemotherapy, nor the distribution of the electrical field in tumors for the electrochemotherapy as well as the relation between the therapeutic effects and the distribution of electrical field, is known very well.The main research content of this paper is as follows:(1) We has studied the influence to many kinds of in vitro cells through the experimental contrast the long time low intensity (20V/cm) periodic transient electric pulses and the short time high intensity (kV/cm) electric pulses. It proved that two kinds of signals all have electroporation effect to cells, and perforation rate and hole size besides with pulse parameters related, but also with cell types, structure, composition, shape and cell activity. The experiments discovered that, electroporation of low intensity electric pulses on cell membrane is reversible. In the irradiation 20～60min time, the perforation rate was not correlated with the irradiation time obviously, the holes in the membrane addressed to emerge unceasingly to appear, and unceasingly voluntarily repairs, not cell necrosis. The cell electroporation probability of high intensity electric pulses is proportional to the pulse amplitude, pulse duration, and pulse number. In individual cells appears electroporation quantity to be many in a certain range. Perforation rate and pulse amplitude and the pulse duration product has the direct ratio, the product may cause excessive cell irreversible electric breakdown, cell death.(2) By selecting different tumor cells in vitro, anti-cancer drugs, drug dose, electric pulse parameters and approach, experimental study pulse united the anticancer drugs to the tumor cell toxicity. Proved electrochemotherapy (ECT) may obviously enhance the toxicity of drugs on tumor cells. Especially electric pulse unites low-dose drugs can raise cell toxicity more than three times, and different tumor cell sensitivity to various anticancer drugs. Therefore, electrochemotherapy is in accordance with the appropriate type of drug.(3) We had established the sarcoma 180 Kun Ming mice model, combined pulse with low-dose cyclophosphamide, doxorubicin, and bleomycin in the treatment of sarcoma 180 in mice, the experimental results indicated: ECT group, inhibition rate to reach 66～95ï¼…, tumor regression ratio is 100ï¼…, and completely vanishes the proportion is 41～73ï¼…. The results are superior to other groups obviously. Slices of tumor tissue staining and histochemical analysis, discovered electrochemotherapy is the strongest to tumor cells inhibitory action, the major part tumor cells suffers injury even necrosis, shows that electric pulses combined with chemotherapy drugs to treat the sarcoma 180 mice to have the function which synergistic effect.(4) Pulsed through low-dose Cisplatin in the treatment of ovarian cancer xenograft SKOV₃ nude mice, achieved good results for the electrochemical clinical therapy in ovarian cancer, and laid a foundation for application.The above tumor electrochemotherapy experiments show: the high intensity electric pulse has a strong inhibition to tumor, which has the selectivity to the tumor treatment function, such selectivity characteristics of the tumor tissue and tumor cells and the biological characters. Therefore, electrochemical characteristics of the tumor should be chosen suitable to pulse parameters in tumor electrochemotherapy.The dose used in the experiment is far lower than conventional dose of chemotherapy, and directly gave drug in the tumor site, reduced the total amount of drugs and the whole cycle of drug. The toxicity and side effect are very small. By doing kidney pathologic examination of the mice after chemotherapy, it shows that low dose of the drug did not cause renal injury in nude mice.High intensity electric pulse has the obvious inhibitory action to tumor blood vessel growth. The tumor surface processes which after the electric field nearly does not have blood vessel distribution, but has rich blood vessel without electric field processing. Used the immunohistochemical technique to examine in the tumor pathological section the vascular endothelial growth factor VEGF acceptor the expression of KDR, obviously decreased after electric field processing tumor VEGF aceeptor KDR, tumor vascular growth was inhibited. It shows that high intensity electric field has inhibited the tumor vascular growth, reduced or has completely cut off the tumor's nutrition supply, urged the tumor cell apoptosis or necrosis, achieved the purpose of the treatment of tumors.(5) Based on the importance of electric field distribution in tumor tissue electrochemotherapy, we considered the mice xenograft tumor model and the condition of experimental, had independently developed an electrode array that used in the tumor electrochemotherapy, had analyzed the field distribution around the electrode to different electrode array and different loading methods. We had established a simplified model of tumor tissue, union tumor and surrounding electric parameters, analyzed the electric field which produces in the tumor tissue by electrode array, analyzed the experimental results and given the electrode array selection principles in electrochemotherapy.(6) According to the dielectric medium properties of cells, we had analyzed the relations of electric field and the cell shape. It shows that the cell should have the concentration effect of electric field when the electric field through a convexconcave cell. From the perspective of the electric field to enhance, we explained the reason of rabbit blood red cell electroporation that had the concave-convex structure occurred in the center place.(7) We has adopted the cell kinetics model to preliminary analyze the mechanism of electroporation, theoretically analyzed the main factors of electroporation. Finally on the electroporation and the electric field intensity as well as temperature relations, we had conducted the electroporation experimental study to yeast cells. Separately changes the pulse voltage and temperature in the study of yeast cells by electroporation. The results showed: pulse amplitude is the main factor to exert influence perforation, but the ambient temperature change is very small to the electroporation influence, has some reference value to the electroporation and electrochemotherapy of the in-depth study.This article innovation spot:(1) From the cell shape of the electric field distribution in perspective, we had analyzed the relationship between cell shape and electroporation for the first time.(2) We had analyzed the electric field distribution in tumor to different electrode array structure and different imposed pulse mode for the first time.(3) We had established a xenograft model of human ovarian cancer SKOV₃ in nude mice, first electrochemotherapy for treatment.(4) We had contrasted research the electroporation of the low intensity and the high intensity electric pulse. (5) We had experimental studied the influence of pulse amplitude and the environmental temperature to cell electroporation, had pointed out the main reasons of electroporation.