AIM: To investigate the expression of miR-210 and the role it plays in the cell cycle to regulate radioresistance in oesophageal squamous cell carcinoma(ESCC). METHODS: Mi R-210 expression was evaluated in 37 pairs of...AIM: To investigate the expression of miR-210 and the role it plays in the cell cycle to regulate radioresistance in oesophageal squamous cell carcinoma(ESCC). METHODS: Mi R-210 expression was evaluated in 37 pairs of ESCC tissues and matched para-tumorous normal oesophageal tissues from surgical patients who had not received neoadjuvant therapy, and in the cells of two novel radioresistant cell lines, TE-1R and Eca-109 R, using quantitative reverse transcription-polymerase chain reaction(q RT-PCR). The transient up-regulation of mi R-210 expression in TE-1R and Eca-109 R cells was studied using liposomes and was confirmed using qR T-PCR. The rate of cell survival after a series of radio-treatment doses was evaluated using the cloneformation assay. Flow cytometry was used to detect the changes to the cell cycle patterns due to radiation treatment. RT-PCR and Western blot were used to detect the expression of ataxia telangiectasia mutated(ATM) and DNA dependent protein kinase(DNA-PKcs) after irradiation, and the cell sphere formation assay was used to evaluate the proliferative ability of the cancer stem-like cells.RESULTS: The level of mi R-210 expression was significantly decreased, by 21.3% to 97.2%, with the average being 39.2% ± 16.1%, in the ESCC tissues of most patients(81.1%, 30 of 37 vs patients with high mi R-210 expression, P < 0.05). A low level of expression of miR-210 was correlated with a poorly differentiated pathological type(P < 0.01) but was not correlated with the T-stage or lymph node infiltration(both P > 0.05). Early local recurrences(< 18 mo, n = 19) after radiotherapy were significantly related with low miR-210 expression(n = 13, P < 0.05). The level of mi R-210 was decreased by approximately 73%(vs TE-1, 0.27 ± 0.10, P < 0.01) in the established radioresistant TE-IR cell line and by 52%(vs Eca-109, 0.48 ± 0.17, P < 0.05) in the corresponding Eca-109 R line. Transient transfection with a mi R-210 precursor increased the level of mi R-210 expression, leading to a significant increase in cell survival after radiotherapy(P < 0.05). Twenty-four hours after radiation, the proportion of pmiR-210 cells in S phase was increased(vs control cells, 30.4% ± 0.4%, and vs untreated TE-1R cells, 23.3% ± 0.7%, P < 0.05 for both). The levels of DNA-PKcs(0.21 ± 0.07) and ATM(0.12 ± 0.03, P < 0.05) proteins were significantly lower in the PmiR-210 cells than in control cells, but no differences were found in the levels of the corresponding mR NAs in the two cell types(P > 0.05 for all). Exogenous mi R-210 expression decreased the diameter of pmi R-210 cell spheres(vs control cells, 0.60 ± 0.14, P < 0.05).CONCLUSION: Mi R-210 expression is negatively correlated with the pathological type and the local survivalrate after radiotherapy, and high expression of miR-210 may reverse the radioresistance of ESCC stem-like cells.展开更多
基金Supported by National Natural Science Foundation of China,No.30972962
文摘AIM: To investigate the expression of miR-210 and the role it plays in the cell cycle to regulate radioresistance in oesophageal squamous cell carcinoma(ESCC). METHODS: Mi R-210 expression was evaluated in 37 pairs of ESCC tissues and matched para-tumorous normal oesophageal tissues from surgical patients who had not received neoadjuvant therapy, and in the cells of two novel radioresistant cell lines, TE-1R and Eca-109 R, using quantitative reverse transcription-polymerase chain reaction(q RT-PCR). The transient up-regulation of mi R-210 expression in TE-1R and Eca-109 R cells was studied using liposomes and was confirmed using qR T-PCR. The rate of cell survival after a series of radio-treatment doses was evaluated using the cloneformation assay. Flow cytometry was used to detect the changes to the cell cycle patterns due to radiation treatment. RT-PCR and Western blot were used to detect the expression of ataxia telangiectasia mutated(ATM) and DNA dependent protein kinase(DNA-PKcs) after irradiation, and the cell sphere formation assay was used to evaluate the proliferative ability of the cancer stem-like cells.RESULTS: The level of mi R-210 expression was significantly decreased, by 21.3% to 97.2%, with the average being 39.2% ± 16.1%, in the ESCC tissues of most patients(81.1%, 30 of 37 vs patients with high mi R-210 expression, P < 0.05). A low level of expression of miR-210 was correlated with a poorly differentiated pathological type(P < 0.01) but was not correlated with the T-stage or lymph node infiltration(both P > 0.05). Early local recurrences(< 18 mo, n = 19) after radiotherapy were significantly related with low miR-210 expression(n = 13, P < 0.05). The level of mi R-210 was decreased by approximately 73%(vs TE-1, 0.27 ± 0.10, P < 0.01) in the established radioresistant TE-IR cell line and by 52%(vs Eca-109, 0.48 ± 0.17, P < 0.05) in the corresponding Eca-109 R line. Transient transfection with a mi R-210 precursor increased the level of mi R-210 expression, leading to a significant increase in cell survival after radiotherapy(P < 0.05). Twenty-four hours after radiation, the proportion of pmiR-210 cells in S phase was increased(vs control cells, 30.4% ± 0.4%, and vs untreated TE-1R cells, 23.3% ± 0.7%, P < 0.05 for both). The levels of DNA-PKcs(0.21 ± 0.07) and ATM(0.12 ± 0.03, P < 0.05) proteins were significantly lower in the PmiR-210 cells than in control cells, but no differences were found in the levels of the corresponding mR NAs in the two cell types(P > 0.05 for all). Exogenous mi R-210 expression decreased the diameter of pmi R-210 cell spheres(vs control cells, 0.60 ± 0.14, P < 0.05).CONCLUSION: Mi R-210 expression is negatively correlated with the pathological type and the local survivalrate after radiotherapy, and high expression of miR-210 may reverse the radioresistance of ESCC stem-like cells.
