| Electrophysiological activity is one of the most common phenomena throughout the entire cell lifetime. It has close relationship with cell proliferation cycles as well as plays an important role in recognition, conglutination, phagocytosis, differentiation, metastasis and malignant transformation. Recent researches had discovered that there was significant difference in membranous charges between normal cells and tumor cells. Moreover, the malignant phenotypes of tumor cells could be inhibited by outside electromagnetic field. So the changes of membranous charges was not only direct manifestation but also consequence of cell malignant transformation, furthermore it may probably participate in the process of canceration.It is more and more important and necessary to measure membrane charge, membrane potential and membrane electric capacity with understood cellular electrophysiological activities deeply, The general methods used in membrane charges research were Laser Doppler Electrophoresis and Single-cell Gel Electrophoresis, but they were relative quantification methods which had difficulties in accurately measuring the electrophysiological activities of hepatocellular carcinoma cells. So great significance still needs for measurement the electrophysiological activities accurately and exploration variability in membrane charges among variably differentiated hepatocellular carcinoma cells.Objectives:To compare the difference in membrane charge, membrane potential and membrane electric capacity between the hepatocellar carcinoma cells (HCC) and normal hepatocyte so as to elucidate variability of membrane charges in process of HCC canceration.Methods:(1) Six cell lines were cultured in recommended condition including normal hepatocyte HL-7702, well-differentiated HCC cells of Huh-7and HepG2, mid-differentiated HCC cell of BEL-7402and poor-differentiated HCC cells of SK-HEP-1and SMMC-7721, ten cells of each cell line at G1and M phases were separated by Flow Cytometery, and maintained their quiescent condition.(2) The established "Optical Tweezers-Cell Electrophoresis" system was used to measure the membrane charges of yeast cell for confirmation its accuracy and stability.(3) Both membrane potential and membrane electric capacity of six cell lines were measured by Patch Clamp technique.(4) All the data were statisticed by independent sample T-test using SPSS17.0software to show statistical significance (P<0.01).Results:(1) The membrane charge of yeast cell was5.11±0.24×10-10C as well as the membrane charge density was6.25±0.26×10-12C/βm2measured by the established "Optical Tweezers-Cell Electrophoresis" system.(2) The membrane potentials of HL-7702, Huh-7, HepG2, BEL-7402, SK-HEP-1and SMMC-7721were respectively-5.4±1.71mv,-35.88±4.20mv,-36.90±1.86mv,-38.50±5.15mv,-44.70±3.95mv,-47.20±4.64mv at G1phase as well as-12.6±1.63mv,-56.90±5.20mv,-57.00±4.54mv,-58.45±5.16mv,-75.70±3.59mv,-74.70±5.64mv at M. phase; The membrane electric capacities were respectively15.32±1.81pf,19.88±1.30pf,19.70±1.21pf,19.96±1.44pf,25.72±2.35pf and25.93±2.36pf at G1phase as well as20.50±1.44pf,28.86±1.29pf,28.88±1.22pf,28.96±1.54pf,40.69±2.33pf and40.91±2.34pf at M phase.(3) The membrane potentials of HCCs significantly decreased compared with that of normal hepatocyte at the same cell phases (G1and M phases)(P<0.01). The membrane potentials of poor-differentiated HCCs significantly decreased (P<0.01) compared with the other two kinds HCCs including mid-differentiated and well-differentiated ones. There was little difference in membrane potential between mid-differentiated and well-differentiated HCCs, so was in the cell lines at the same differentiated status (P>0.05).(4) The membrane electric capacities of HCCs significant increased compared with that of normal hepatocyte at the same cell phases (G1and M phases)(P<0.01), and those of poor-differentiated HCCs significantly increased compared with those of mid-differentiated and well-differentiated ones as well (P<0.01). But little difference in membrane electric capacity was shown between mid-differentiated and well-differentiated HCCs, so was in the cell lines at the same differentiated status (P>0.05).(5) The membrane potentials of six cell lines in M phase significantly decreased (P<0.01) as well as the membrane electric capacities significant increased in M phase (P<0.01) compared with those in Gl phase, especially those of5HCCs.Conclusion:(1) The membrane charges of yeast cells could be accurately measured by the established "Optical Tweezers-Cell Electrophoresis" system which was good for further used in measuring the membrane charges of HCCs and normal hepatocyte.(2) The membrane charges of HCCs were significantly lower than that of normal hepatocyte in quiescent condition at the same cell phase, the more poorly differentiated HCC were, the lower the membrane charges were. (3) The membrane electric capacities of HCCs were significantly higher than that of normal hepatocyte in quiescent condition at the same cell phase; the more poorly differentiated HCC were, the higher the membrane electric capacities were.(4) Both of membrane charges and membrane electric capacities of6cell lines at M phase were significantly different from those at Gl phase, especially those of HCCs probably because of much more active metabolism of HCC at M phase. |
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