Research Paper

Figure 4. LncRNA CASC15 inhibits the sensitivity of ESCC to radiotherapy by decreasing miR-140-5p and upregulating MMP7. (A) RT-qPCR to detect the expression of lncRNA CASC15, miR-140-5p, and MMP7 in the radio-sensitive and radio-resistant ESCC patients (*p < 0.05 vs. the sensitive group). (B) Western blot analysis to measure the protein expression of MMP7 in the radio-sensitive and radio-resistant ESCC patients (*p < 0.05 vs. the sensitive group). (C) RT-qPCR to determine the expression of lncRNA CASC15 in ESCC naive cell line TE-1 and radiation resistant cell line TE-1R (*p < 0.05 vs. ESCC parental cell line TE-1). (D) RT-qPCR to test the transfection efficiency in EC109 cells. (E) CCK-8 assay to evaluate cell viability of EC109 cells. (F) Flow cytometry to assess cell apoptosis of EC109 cells. (G) Western blot analysis to determine the protein level of Fas, Ki-67 and cleaved Caspase 3 in EC109 cells. (H) Clone formation ability of EC109 cells after X-ray radiation. In panel (C–G), *p < 0.05 vs. EC109 cells transfected with 4 Gy + NC mimic. #p < 0.05 vs. EC109 cells transfected with 4 Gy + sh-NC. &p < 0.05 vs. EC109 cells transfected with 4 Gy + oe-NC + sh-NC; @p < 0.05 vs. EC109 cells transfected with 4 Gy + oe-CASC15 + sh-NC. The measurement data were presented as the mean ± standard deviation. Data in panel (A, B) were analyzed by unpaired t test, and data in panel (C, E, F) were analyzed by one-way analysis of variance, followed by Tukey’s post hoc test. Data in panel D were analyzed by repeated-measures analysis of variance, followed by Tukey’s post hoc test, and data in panel (E) were analyzed by two-way analysis of variance, followed by Tukey’s post hoc test; the experiment was repeated 3 times. 4 Gy + NC mimic, EC109 cells transfected with negative control of mimic and treated with 4 Gy irradiation; 4 Gy + miR-140-5p mimic, EC109 cells transfected with miR-140-5p mimic and treated with 4 Gy irradiation; 4 Gy + sh-NC, EC109 cells transfected with negative control plasmids of short hairpin RNA and treated with 4 Gy irradiation; 4 Gy + sh-MMP7, EC109 cells transfected with short hairpin RNA against matrix metallopeptidase 7 and treated with 4 Gy irradiation; 4 Gy + oe-NC + sh-NC, EC109 cells transfected with negative control plasmids of overexpression vector and short hairpin RNA and treated with 4 Gy irradiation; 4 Gy + oe-CASC15 + sh-NC, EC109 cells transfected with plasmids overexpressing lncRNA CASC15 and negative control of short hairpin RNA and treated with 4 Gy irradiation; 4 Gy + oe-CASC15 + sh-MMP7, EC109 cells transfected with plasmids overexpressing lncRNA CASC15 and short hairpin RNA against matrix metallopeptidase 7 and treated with 4 Gy irradiation.