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Fig. 8 | Journal of Experimental & Clinical Cancer Research

Fig. 8

From: Radiation induces NORAD expression to promote ESCC radiotherapy resistance via EEPD1/ATR/Chk1 signalling and by inhibiting pri-miR-199a1 processing and the exosomal transfer of miR-199a-5p

Fig. 8

a The starBase database predicted the complementary sequences between miR-199a-5p and NORAD. b EEPD1 expression in radioresistant ESCC cells (KYSE-150R and TE-1R) and their parental cells (KYSE-150 and TE-1) was measured using western blotting. The effects of NORAD knockdown on EEPD1 expression in KYSE-150R and TE-1R cells were measured using western blotting. c EEPD1 expression in KYSE-150 and TE-1 cells transduced with miR-199a-5p mimics or miR-199a-5p inhibitors, respectively, was measured using western blotting. d Dual-luciferase reporter assay conducted by cotransfecting the wild-type or mutant 3’UTR EEPD1 mRNA with the miR-199a mimic or negative control into KYSE-150 cells. e Relative importance of genes for segregating patients with radioresistant ESCC from radiosensitive patients calculated using the random forest algorithm. The importance of each gene was ranked by its IncMSC values and IncNodePurity values. f ROC curve for segregating patients with radioresistant ESCC from radiosensitive patients based on EEPD1 expression. g The effect of EEPD1 on radiotherapy outcomes in ESCC xenografts. Subcutaneous ESCC xenografts were established with SH-EEPD1 cells and SH-NC cells. Tumours were exposed to 8 Gy of X-ray radiation. The tumours were harvested and imaged at 3 weeks after radiotherapy delivery; tumour volumes were calculated and analysed. h KYSE-150 and TE-1 cells were transduced with an I-SceI expression plasmid to induce DSBs, and homologous recombination repair in EEPD1 knockdown cells and control cells was assayed using flow cytometry and is presented as the percentage of GFP-positive cells. i Levels of ATM, ATR, Chk1 and phosphorylated ATM, ATR, and Chk1 were analysed in NORAD knockdown cells and control cells treated with or without 8 Gy of X-ray using western blotting. j Levels of phosphorylation of ATR and Chk1 were analysed in EEPD1 knockdown cells and control cells treated with or without 8 Gy of X-rays using western blotting. k Levels of phosphorylation of ATR and Chk1 were analysed using western blotting in KYSE-150-SH-NORAD cells with or without EEPD1 overexpression after 8 Gy of MV X-ray treatment

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