Effect and underlying mechanism of glutamine on radiosensitivity of colon cancer cells

Abstract

Objective To observe the effect of different concentrations of glutamine (Gln) on the radiosensitivity of colorectal cancer HT-29 cells and explore the possible mechanism. Methods According to different Gln concentrations, HT-29 cells at logarithmical growth were divided into control group (2 mmol/L, as the basal medium concentration group) and experimental groups Ⅰ, Ⅱ and Ⅲ (4, 6 and 8 mmol/L). After a 2-hour pre-treatment, all groups were exposed to 8 Gy irradiation of a Co-60 radiation source. CCK-8 assay and clonal formation assay were used respectively to explore the effects of different Gln concentrations on cell viability and cell radiosensitivity after irradiation. The level of reactive oxygen species (ROS) in each group was measured in 24 h after irradiation, and the apoptotic rate was detected with flow cytometry in 48 h after irradiation. The protein expression levels of Nrf2, HO-1, and cleaved-Caspase3 were determined by Western blotting. Results In 24 h after Gln intervention, the cell viability of experimental groups Ⅱ and Ⅲ of non-irradiated HT-29 cells was significantly higher than that of the control group and of experimental group Ⅰ (P < 0.05). In 24 h after radiation, the cell viability of each experimental group was significantly higher than that of the control group (P < 0.05). In 14 d after radiation, there were more clone formation in each experimental group than the control group (P < 0.05). The ROS level was significantly lower in each experimental group than the control group in 24 h after radiation (P < 0.05). After 48 h of radiation, the apoptotic rate was notably lower in each experimental group than the control group (P < 0.05). The expression level of Nrf2 in the experimental group Ⅰ was higher than that of the control group (P < 0.05), those of Nrf2 and HO-1 in the experimental groups Ⅱ and Ⅲ were higher than those of the control group and experimental group Ⅰ (P < 0.05). While the expression of cleaved-Caspase3 in the experimental groups Ⅱ and Ⅲ was lower than the control group and experimental group Ⅰ (P < 0.05), and it in the experimental group Ⅲ was lower than that of experimental group Ⅱ (P < 0.05). Conclusion Gln can significantly reduce the radiosensitivity of HT-29 cells, which is associated with its reducing oxidative stress damage and reducing cell apoptosis. Our results suggest that Gln might be detrimental to radiation therapy in patients with colorectal cancer.

Keywords

• glutamine
• ht-29 cells
• oxidative stress damage
• cell apoptosis
• radiosensitivity