The Role Of Cytoskeleton In Bioelectrical Effects Of Tumor Cells Induced By Steep Pulse

Malignant tumor is an universal disease threatening human in the world. Accordding to the statistics of WHO, millions of people died in cancer every year and the disease tendency in china is increasing.At present,there are many traditional tumor therapies such as surgery, radiotherapy, chemotherapy and hormonal therapy. But all of them have disadvantages. As the development of electric pulse power technology, electric pulse has been applied in many areas such as disease cure, gene transfection and medical equipment recently. Moreover, steep pulse has been used to open up a new way in tumor therapy, owing to its bioelectrical effects of cell electroporation, intracellular manipulation and irreversible electrical breakdown. In order to overcome the limitation of traditional therapies and get rid of the chemicals in tumor therapy, the basic theories of steep pulse tumor therapy were investigated basing on the electric new technology, biomedicine and mechanical subjects.In this paper, the subcellular level mechanisms of steep pulse tumor therapy including apoptosis effect, perforation effect, mitochondria membrane potential changes and the role of cytoskeleton were investigated using steep pulse tumor therapy instrument with fluorescence microscopy, flow cytometry,fluorescence spectrophoto- meter technologies.Results: Steep pulse electric field with strength of 8KV/cm, durations of 450ns and frequency of 1HZ was applied to adherent cells of HepG2 and L02. Flow cytometry apoptosis results indicated that the early apoptosis rate of HepG2 was higher than L02 after steep pulse application and the early apoptosis and necrosis rates of both cells were decreased with cytochalasin B pretreatment; Trypan blue exclusion results indicated that the cell motility rate of HepG2 was lower than L02 after steep pulse application and the cell motility rate of both cells were increased with cytochalasin B pretreatment; Fluorescence spectrophotometer results indicated that the mitochondria membrane potential of HepG2 cell was gradually decreased (from greatly to placidly)at the several time points(0h,0.5h,1h,2h,4h) after steep pulse application and the mitochondria membrane potential decreased degree was reduced with cytochalasin B pretreatment; Perforation results indicated that cell membrane of HepG2 cell was perforated at approximately 7 minutes after steep pulse application which was still occurred after 4 hours and the perforation time of HepG2 cell with cytochalasin B pretreatment was no significant difference.Conclusion: This electric field parameter of steep pulse can induce apoptosis of hepatoma carcinoma cell via mitochondria pathway and the hepatoma carcinoma cells were more sensitive than normal liver cells; The steep pulse can also induce necrosis of hepatoma carcinoma cells, possibly through the destruction of cell membrane; The loss of actin cytoskeleton strongly protected against the apoptosis effects induced by steep pulse possibly through suppressing the decrease of mitochondria membrane potential and was no significant difference on necrosis effects. From these results, it was concluded that cytoskeleton can modulate the apoptosis signal induced by steep pulse and have an important role in the bioelectrical effects of tumor cells by steep pulse.