Resistance reverse effect of anemoside B4 and tetrandrine on oxaliplatin-resistant human colon cancer cells and the mechanism

Abstract

Background and purpose: Oxaliplatin(L-OHP) is one of the most commonly used chemotherapy drugs in colorectal cancer and L-OHP-resistance is very common in colorectal cancer therapy. This research was to discuss the reversal effect in L-OHP-resistant human colon cancer cell line Lo Vo/L-OHP by anemoside B4(AB4) and tetrandrine(Tet) and to clarify their molecular mechanism. Methods: Lo Vo/L-OHP cells were treated for 48 h by AB4 and Tet at different concentrations to get non-toxic dose. Drug sensitivity was measured by MTT. After the treatment, the cell cycle and apoptosis of the cells were detected. Expression of P-gp m RNA, z DHHC9 m RNA and SMAD4 m RNA were detected by RT-PCR. Expression of P-gp, z DHHC9 and SMAD4 protein were detected by Western blot. Results: The IC50 of Lo Vo/L-OHP for L-OHP was(112.5±23.6) μg/m L, and the IC50 decreased to(62.8±21.4) μg/m L(P0.05) and(58.9±26.3) μg/m L(P0.05) after the treatment with 0.71 μg/m L AB4 and 0.45 μg/m L Tet(non-toxic dose) separately. The cell cycle experiment showed that the cells in G1 stage decreased and in S stage increased but with no statistical significance(P0.05). The cell apoptosis experiment showed that the apoptotic rate increased after the treatment with AB4 and Tet with non-toxic dose but with no statistical significance(P0.05). The RT-PCR experiment showed that the expressions of P-gp m RNA and z DHHC9 m RNA were increased and SMAD4 m RNA was decreased in Lo Vo/L-OHP cells compared with in Lo Vo cells(P0.05), while it was found that P-gp m RNA in Lo Vo/L-OHP cells was decreased after the treatment with AB4 at non-toxic dose(P0.05). Western blot experiment showed the protein of P-gp in Lo Vo/L-OHP cells was decreased after treatment with AB4(P0.05), which was accordant to the PCR result. The expression of z DHHC9 protein in Lo Vo/L-OHP cells was decreased in Lo Vo/L-OHP cells after treatment with Tet(P0.05). Conclusion: AB4 and Tet have some reversal effect on resistant to L-OHP in Lo Vo/L-OHP cells. The molecular mechanism of the resistance reverse effect was related to down-regulation of P-gp for AB4 and down-regulation of z DHHC9 for Tet.