Modulation of multidrug resistance by andrographolid in a HCT-8/5-FU multidrug-resistant colorectal cancer cell line.

摘要

OBJECTIVE: To develop an HCT-8/5-FU multidrug-resistant colorectal cancer cell line and to elucidate the effect of Andrographolid (AG), an extract from Andrographis paniculate, a medicinal herb on the HCT-8/5-FU multidrug-resistant colorectal cancer cell line. METHODS: An HCT-8 colorectal cancer cell line was used and a high concentration of 5-Fluorouracid (5-FU) was introduced at the beginning to induce drug resistance, then the concentration of 5-FU was increased in gradients. Approximately 7 months later, the cells grew stably in 2.0 microg/mL of 5-FU, and the cell line was named HCT-8/5-FU multidrug-resistant colorectal cancer cell line. The resistant index of HCT-8/5-FU cells to 5-FU, adriamycin (ADM), cisplatin (DDP) was checked by MTT test, and a growth curve was drawn. The morphological changes were observed by both light and electron microscope. The function of P-170 was detected by rhodamine staining. After the application of AG and co-administration of 5-FU, ADM and DDP, the growth curves and inhibition rate as well as apoptosis rate of HCT-8/5-FU at different concentrations of AG were evaluated by MTT and flow cytometry. Rhodamine staining was used to investigate the possible mechanism involved by AG. RESULTS: The resistance index of HCT-8/5-FU to 5-FU was 16.6, and a cross-resistance to ADM and DDP was noticed. Compared with parental cells, HCT-8/5-FU cell's growth rate did not change significantly but the cell's morphology was remarkably changed as compared with parental cells. Overexpression of P-170 by HCT-8/5-FU cell was indicated through rhodamine staining. AG at a low concentration showed weak inhibitory effect on HCT-8/5-FU. However, a remarkable inhibitory and apoptosis rate was shown when AG was co-administered with 5-FU, ADM and DDP, respectively. Interestingly AG alone could not induce apoptosis and change the cell cycles. AG might affect the expression of P-170, which was indicated by rhodamine staining. CONCLUSIONS: The HCT-8/5-FU multidrug-resistant colorectal cancer cell line has been successfully developed and because it has cross-resistance to 5-FU, ADM and DDP, it might serve as an ideal multidrug resistance (MDR) model for colorectal cancer research. The mechanism of HCT-8/5-FU resistance to chemotherapeutic agents might be related to the overexpression of P-170. Low concentrations of AG alone have no significant inhibition on HCT-8/5-FU and fail to induce apoptosis and to change cell cycles. AG might act as a chemosensitizer when co-administered with 5-FU, ADM and DDP, and the mechanism of reversal modulation of multidrug resistance by AG in the HCT-8/5-FU resistant cell line might be related to its downregulation of overexpression of P-170.