Effect of HMGB1 knockdown by RNA interference on cell proliferation and migration of esophageal squamous cell carcinoma after X-ray radiation

Abstract

Objective To evaluate the effect of X-ray radiation on cell proliferation, migration, survival ability and cell cycle of human esophageal squamous cell carcinoma after RNA interference-mediated down-regulation of HMGB1 gene expression. Methods The expression of HMGB1 at mRNA and protein levels in the human esophageal squamous cell carcinoma cell lines ECA109 and KYSE30 was determined using RT-PCR and Western blot assays. MTS and Transwell assays were employed to examine the proliferation and migration of ECA109 and KYSE30 cell lines. The cellular survival ability in vitro was assessed by clone formation assay. The cell cycle after X-ray radiation in different groups was detected by flow cytometry. Results The expression of HMGB1 at mRNA and protein levels in ECA109 and KYSE30 cells were markedly higher in a dose-dependent and time-dependent manner in the radiation group than that in the control group (all P<0.05). MTS results demonstrated that the proliferation of ECA109 and KYSE30 cells was obviously lower at each time point after radiation than that in the group without radiation (all P<0.01). The expression of HMGB1 at mRNA and protein levels was significantly inhibited in the HMGB1 siRNA group than those in the control and NC groups (both P<0.01). The data from the clone formation assay revealed that the radiosensitivity was significantly increased after down-regulation of HMGB1 expression (P<0.01). Transwell migration assay revealed that the number of migrating cells at the fourth hour after X-ray irradiation in the HMGB1 siRNA group was significantly lower than those in the control and negative groups (both P<0.01). In the HMGB1 siRNA group, the percentage of cells at G0/G1 phase was obviously higher, whereas the percentage of S phase was significantly lower than those in the control and NC groups, and the trend was even more significant after X-ray radiation (all P<0.01). Conclusion Inhibition of HMGB1 expression by siRNA can suppress the proliferation and migration of ECA109 and KYSE30 cells and enhance the radiosensitivity by increasing the cell cycle arrest at G0/G1 stage after X-ray irradiation in vitro. Key words: HMGB1 gene; Cell proliferation and migration; Cell cycle; Radiosensitivity