Effects Of Steep Pulsed Electric Field On Biorheological Properties Of Hepatoma Cells

Malignant tumor is one of the most severe diseases which does harm to human's health. At present, the main treatments for tumor include chemical therapy, radiation therapy, biologic therapy and electroporotion therapy. Electroporotion therapy is a new developed technique to kill malignant tissues or cells and has become one of the efficient methods of tumor comprehensive treatment. However there is lack of all-round understanding about the mechanisms of electroporotion therapy to tumor.In this thesis, we combined biorheology and electrical technique, and investigated the growth, proliferation and apoptosis related gene expression of hepatoma cells treated by EPEF. We also studied the change of viscoelasticity,adhesive force of hepatoma cells to human umbilical vein endothelial cells(HUVECs) using of micropipette aspiration technique. The main works and results in this paper are as follows:1. SPEF equipment and experimental parametersSPEF equipment was self-assembled by Key Laboratory of High Voltage & Electrical New Technique of Ministry of Education of China, Chongqing University. It consists of circuit of direct current, producing and output of SPEF and collecting and processing system controlled by computer. In our experiments, we fixed voltage(200V), frequency (100Hz), treanment time (10min) and changed the capacitance values (0.01μF and 0.1μF)to investigate the effects of SPEF on biorheological properties.2.Effects of SPEF on proliferating dynamics of hepatoma cellsWith MTT assay, the growth and proliferation of hepatoma cells treated by SPEF were evaluted. Results showed that SPEF could inhibit the growth and proliferation of hepatoma cells and high dosage treatment showed a stronger inhibition than that of low dasage treatment. It also showed that G0/G1 phase increase but S phase and G2/M phase decrease after treated by SPEF, which suggested that SPEF could change the distribution of cell cycle and block hepatoma cells entering S phase.3. Effects of SPEF on expression of gene involved apoptosis in hepatoma cellsAfter exposure to SPEF, Expression of bcl-2 and p53 gene in hepatoma cells was measured by FCM and FITC marked monoclonal antibody quantificationally. It found that SPEF could down-regulate the expression of bcl-2 gene, but expression of p53 gene don't have any change. It indicated that the apoptosis of hepatoma cells induced by SPEF maybe performed through down-regulate the expression of bcl-2 gene. 4. Effects of SPEF on viscoelastical properties of hepatoma cellsCompared with control, both K1 and K2 values of hepatoma cells treated by low dosage SPEF have no change, while K1 of hepatoma cells treated by high dosage SPEF showed an obvious decrease and K2 significantly increase. Viscous coefficientμshowed an increase trend in all treatment groups after exposure to SPEF. As for the eigenvalues of viscoelasticity of hepatoma cells, it showed that the primary deformation have no defference but the maximum deformation of high dasage treatment hepatoma cells increase markedly when compared with control cells. It also showed a smaller rigidity and longer time of creeping deformation than that of control cells.5. Effects of SPEF on adhesive force of hepatoma cells to HUVECsAdhesive force of low dasage treatment cells to HUVECs present a higher value when compared with control cells. However, adhesive force of high dasage treatment cells to HUVECs reveals no significant difference compared with control cells.