Reversing Effect Of EGCG On Sorafenib Resistance Of Human Hepatocellular Carcinoma Cell Lines

OBJECTIVE Establishment of an Sorafenib-resistant Human Hepatoma Cell Li nes Bel-7402/Sora and to explor the reversal effects of EGCG and its related mechanism.METHODS 1.The effects of Sorafenib on Bel-7402 cell survival rate was dete rmined by CCK8 method,and the models of Sorafenib-resistant Human Hepato ma Cell Lines Bel-7402/Sora were established. Morphological changes were obs erved by inverted microscope between drug resistant cell line and parental cell line. The IC50 was detected by CCK8 method in hepato cellular carcinoma cel 1 line Bel-7402/Sora, and index of multidrug resistant was calculated.Western-bl ot method was used to detect the expression of protein GRP78 and Beclinl in two cell lines with different concentrations of Sorafenib.2.CCK8 method was used to observe the inhibitory effects of different con centrations of EGCG on the resistant cell lines and the parental cell lines, and to calculate the non cytotoxic concentration of EGCG. Compare with the diffe rent effect of different concentration of Sorafenib combined with non-toxic con centration of EGCG before and after on the survival rate of the resistant cell 1 ines, and to calculate its reversal ratio and relative reversal efficiency. Western-blot method was used to detect the expression of protein GRP78 and Beclinl i n the resistant cell lines with different concentrations of Sorafenib combined wi th non cytotoxic concentrations of EGCG before and after.RESULTS 1. Sorafenib can inhibit the proliferation of hepatoma cell line Bel-7402, and showed concentration dependent, its IC50 value is 1.654 μmol/L. Co mpared with parental cell lines, the drug resistant cell lines were larger in size and irregular in shape, and a large number of vacuoles and granules appeared in the cytoplasm.The IC50 value of Sorafenib to hepatocellular carcinoma cell line Bel-7402/Sora was 10.344 μmol/L, and the drug resistance ratio was 6.25. With the increase of the concentration of Sorafenib, the expression of GRP78 was increased in two kinds of cells, the expression of Beclinl was decreased at first and then increased.The expression of the two proteins was higher in th e drug resistant cell lines than that in the parental cell lines.2. The cell survival rate was lower than before with different concentratio ns of Sorafenib combined with non cytotoxic concentration of EGCG, and the combination of IC50 value was 7.312 μmol/L, the reversing multiple was about 1.415, the relative reversal rate was 34.89%. The expression of GRP78 and B eclinl were all decreased by giving the combination of certain concentration S orafenib and non cytotoxic concentrations of EGCG in on the resistant cell lin es.CONCLUSION 1. Successfully established sorafenib resistant hepatocellular car cinoma cell line Bel-7402/Sora,and GRP78 and Beclinl are involved in the me chanism of drug resistance.2. EGCG can reverse the drug resistance of hepatocellular carcinoma cell 1 ines to Sorafenib may be related with the changes of the GRP78 and Beclinl.