Treatment mechanism of matrine in combination with irinotecan for colon cancer

Abstract

The inhibitory effect of matrine (MA) was studied in combination with irinotecan (CPT-11) on proliferation of human colon carcinoma cell line HT29. We also explored the mechanism of cell apoptosis induction in HT29. HT29 cells were treated with different concentrations of MA and CPT-11 alone and in combination. The growth inhibition in HT29 cells was evaluated using MTT assay. Apoptosis was detected using AV-PI double staining flow cytometry. Transmission electron microscopy was used to detect structural changes in cells. Topoisomerase (TOPO) I, Bax and Caspase-3 expression levels were evaluated using western blot analysis. MA and CPT-11 alone and in combination, inhibited the proliferation of HT29 cells, whereas the combination treatment exhibited higher inhibitory effect (P<0.01). This suggests the existence of synergistic cytotoxicity. Compared with each treatment alone, the combination treatment caused more significant damage to cell structure, and caused a significantly higher apoptosis rate (P<0.01). Additionally, the combination treatment increased TOPO I, Bax and Caspase-3 expression levels (P<0.01). In conclusion, MA in combination with CPT-11 synergistically inhibited HT29 cell proliferation and induced apoptosis in these cells. The mechanism may be related to upregulation of the TOPO I, Bax and Caspase-3 protein expression.

Keywords: HT29 cells; apoptosis; colon cancer; irinotecan; matrine.

Figure 1.

Flow cytometry measurement of apoptosis rates of different treatment groups. **P<0.01, compared with normal control group; ^&&P<0.01, combination treatment group compared with MA treatment alone group; ^##P<0.01, combination treatment group compared with CPT-11 treatment alone group. MA, matrine; CPT-11, irinotecan.

Figure 2.

Ultrastructures of HT29 cells in different treatment groups by TEM. (A) The normal control group, (B) the MA treatment alone group, (C) the CPT-11 treatment alone group, and (D) the combination treatment group. TEM, transmission electron microscopy; MA, matrine; CPT-11, irinotecan.

Figure 3.

Western blot analysis of protein expression in HT29 cells. (A) Western blot analysis bands of TOPO I, Bax and Caspase-3, and (B) gray-scale quantification of TOPO I, Bax and Caspase-3. **P<0.01, compared with the normal control group; ^&&P<0.01, the combination treatment group compared with the MA treatment alone group; ^##P<0.01, the combination treatment group compared with the CPT-11 treatment alone group. TOPO, topoisomerase; MA, matrine; CPT-11, irinotecan.