Objective: To explore the possible mechanism of radiotherapy regulating the expression of PD-L1 in esophageal carcinoma. Methods: Three esophageal cancer cell lines (Eca109, Kyse150, TE1) were irradiated with different doses of X-rays, and 6 Gy+ AG490 group was set. The mRNA expression of PD-L1 was detected by real-time quantitative polymerase chain reaction (RT-qPCR). The protein expressions of PD-L1, STAT3, p-STAT3 were detected by western blotting and the protein level of IL-6 was detected by ELISA. Results: The mRNA expressions of PD-L1 in Eca109, Kyse150 and TE1 were 2.86±0.30, 960.01±21.27 and 106.78±6.67, higher than 1.07±0.15 in normal esophageal cell line HET-1A (P<0.01). The protein expressions of PD-L1 in Eca109, Kyse150 and TE1 were 0.091±0.036, 1.533±0.079 and 0.914±0.035, higher than 0.063±0.01 in normal esophageal cell line HET-1A (P<0.01). After 48 hours of 6 Gy irradiation, the protein expression levels of PD-L1 in Eca109, Kyse150 and TE1 were 0.135±0.007, 1.66±0.06 and 1.32±0.06, higher than 0.09±0.01, 1.21±0.05 and 0.93±0.03 of the 0 Gy group (P<0.01), while the protein expression levels of p-STAT3 in Eca109, Kyse150 and TE1 were 1.44±0.26, 0.75±0.04 and 1.92±0.17, higher than 0.18±0.05, 0.48±0.02 and 0.36±0.06 of the 0 Gy group (P<0.01). IL-6 protein expression increased significantly after different doses of irradiation (P<0.01). After the IL-6/STAT3 signaling pathway was blocked by the specific inhibitor AG490, the expressions of PD-L1 of Eca109, Kyse150 and TE1 in the 6 Gy+ AG490 groups were 0.11±0.03, 1.07±0.08 and 0.96±0.11, without significant differences of 0.09±0.01, 0.96±0.05 and 0.85±0.09 of the 0 Gy group (P>0.05), while the protein expressions of p-STAT3 were 0.76±0.11, 0.59±0.06 and 0.96±0.12, without significant differences of 0.67±0.08, 0.54±0.06 and 0.84±0.11 of the 0 Gy group (P>0.05). Conclusion: Radiotherapy may regulate the expression of PD-L1 in esophageal cancer cells through IL-6 / STAT3 signaling pathway.
目的： 探讨放疗调控食管癌细胞程序性死亡蛋白配体1(PD-L1)表达的可能机制。 方法： 以不同剂量X线照射食管癌细胞株Eca109、Kyse150和TE1，同时设6 Gy照射+AG490组。采用实时荧光定量聚合酶链反应检测细胞中PD-L1 mRNA的表达，采用Western blot检测细胞中PD-L1、信号转导和转录激活因子3(STAT3)和磷酸化细胞信号传导和转录活化因子3(p-STAT3)的蛋白表达，采用酶联免疫吸附试验检测细胞中白细胞介素6(IL-6)的蛋白表达。 结果： 食管癌细胞Eca109、Kyse150和TE1中，PD-L1 mRNA的表达量分别为2.86±0.30、960.01±21.27和106.78±6.67，均高于正常食管细胞株HET-1A(1.07±0.15，均P<0.01)；PD-L1蛋白的表达量分别为0.091±0.036、1.533±0.079和0.914±0.035，亦均高于正常食管细胞株HET-1A(0.063±0.01，均P<0.01)。经6 Gy照射后，Eca109、Kyse150和TE1细胞中PD-L1蛋白的表达量峰值分别为0.135±0.007、1.66±0.06和1.32±0.06，均高于0 h组(分别为0.09±0.01、1.21±0.05和0.93±0.03，均P<0.01)；p-STAT3蛋白的表达量峰值分别为1.44±0.26、0.75±0.04和1.92±0.17，均高于0 h组(分别为0.18±0.05、0.48±0.02和0.36±0.06，均P<0.01)。不同剂量照射后，Eca109、Kyse150和TE1细胞中IL-6蛋白表达均显著增加(均P<0.01)。IL-6/STAT3信号通路被特异性抑制剂AG490阻断后，6 Gy放射+AG490组Eca109、Kyse150和TE1细胞中PD-L1蛋白的表达水平分别为0.11±0.03、1.07±0.08和0.96±0.11，与相应细胞株的0 Gy照射组(分别为0.09±0.01、0.96±0.05和0.85±0.09)比较，差异均无统计学意义(均P>0.05)；p-STAT3蛋白的表达水平分别为0.76±0.11、0.59±0.06和0.96±0.12，与相应细胞株的0 Gy照射组(分别为0.67±0.08、0.54±0.06和0.84±0.11)比较，差异亦均无统计学意义(均P>0.05)。 结论： 放疗可能通过IL-6/STAT3信号通路调控食管癌细胞的PD-L1表达。.
Keywords: Esophageal carcinoma; IL-6/STAT3 pathway; Programmed cell death ligand-1; Radiotherapy.