Effects of gamma irradiation on cell cycle, apoptosis and telomerase activity in p53 wild-type and deficient HCT116 colon cancer cell lines

Abstract

Radiotherapy serves as adjunctive treatment to chemotherapy and surgical resection of colorectal cancer. However, the cellular response to irradiation varies depending on the expression of tumor suppressor p53, which plays a significant role in the regulation of cell cycle arrest, apoptosis and telomerase activity in various cancers. The present study aimed to investigate cell cycle arrest, apoptosis and telomerase activity with respect to p53 expression in p53 wild-type (+/+) and deficient (-/-) HCT116 colon cancer cell lines following 5 Gy γ-irradiation. Cell cycle arrest and apoptosis were evaluated using flow cytometry. The telomerase activity was measured using a TRAP (telomerase repeat amplification protocol) assay. Following treatment with irradiation, G₁/S cell cycle arrest occurred in the p53+/+ cells, whereas the p53-/- cells accumulated in the G₂ phase. No differences were observed in the apoptotic ratios between the two cell lines following irradiation. Decreased telomerase activity was observed in the p53+/+ cells, whereas telomerase activity was increased in the p53-/- cells. The results showed that while telomerase activity and G₁ cell cycle arrest were regulated depending on the p53 status, G₂ arrest and the apoptotic response were promoted via a p53-independent pathway.

Keywords: apoptosis; cell proliferation; irradiation; p53; telomerase activity.

Figure 1

(A) Average apoptotic percentages obtained from three independent experiments in irradiated and non-irradiated p53 wild-type (+/+) and deficient (−/−) HCT116 cells. (B) The average apoptotic cell numbers of irradiated and non-irradiated control cells. Values of <10% were considered to be the threshold in the irradiated control p53−/− and p53 +/+ cells. IR, irradiated.

Figure 2

Normal distribution of G₁, S and G₂ stages of the cell cycle in non-irradiated (A) p53 wild-type (+/+) and (B) p53 deficient (−/−) HCT116 cells at 0 and 24 h.

Figure 3

(A) G₁ phase arrest at 24 h and accumulation in G₂ phase arrest at 48 h in irradiated p53 deficient (−/−) HCT116 cells. (B) G₁ and S phase arrest in irradiated p53 wild-type (+/+) HCT116 cells at 24 and 48 h. Blue areas of the histogram illustrate apoptotic cells associated with the sub-G₀ DNA content.

Figure 4

Comparison of telomerase activity in p53 wild-type (+/+) and p53 deficient (−/−) cells in (A) irradiated and (B) non-irradiated cells. IR, irradiated.