Li H G, Zhao L H, Lu A, Liu J B, Su Z J, Wang X B, Gao Y J
Department of Thyroid Surgery,Henan Provincial People's Hospital, Zhengzhou, 450003, China.
Department of Disinfection Supply Center,Fuwai Central China Cardiovascular Hospital, Zhengzhou, 450003, China.
Zhonghua Yi Xue Za Zhi. 2021 Nov 2;101(40):3329-3337. doi: 10.3760/cma.j.cn112137-20210207-00379.
To explore the effect and possible mechanism of circ_0023990 on the radiosensitivity of thyroid cancer cells. qRT-PCR was used to detect the expression of circ_0023990 in the cancer tissues of 55 patients with thyroid cancer and thyroid cancer cell lines (TPC-1, KTC-1, FTC-133 and CAL-62), and the relationship between the expression of circ_0023990 in cancer tissues and the clinical characteristics of the patients were analyzed. Thyroid cancer cells TPC-1 and KTC-1 were divided into sh-circ_0023990 group, sh-NC group, sh-circ_0023990+anti-miR-873-5p group, sh-circ_0023990+anti-miR-NC group, miR-873-5p group, miR-NC group, miR-873-5p+pcDNA-ANXA2 group and miR-873-5p+pcDNA group, and then clone formation experiment was used to detect cell radiosensitivity. After each group of cells was irradiated with 4Gy radiation, the expression of γH2AX protein in the cells was detected by Western Blot. The dual luciferase reporter gene experiment verified the targeting relationship between circ_0023990 and miR-873-5p or miR-873-5p and ANXA2. The expression of circ_0023990 in thyroid cancer tissues was higher than that in normal tissues (2.15±0.09 vs. 0.97±0.05, <0.05), and its expression was closely related to tumor size, lymph node metastasis and TNM staging of patients with thyroid cancer (<0.05). The expression of circ_0023990 in thyroid cancer cell lines (TPC-1, KTC-1, FTC-133 and CAL-62) were higher than that of normal thyroid cells HTori-3 (3.16±0.38, 2.63±0.28, 1.82±0.24, 1.71±0.22 vs. 1.00±0.10, all <0.05). The survival scores of TPC-1 and KTC-1 cells in the sh-circ_0023990 group were significantly lower than those in the sh-NC group (<0.05), and the sensitization ratios were 2.482, 1.643; The survival scores of TPC-1 and KTC-1 cells in the sh-circ_0023990+anti-miR-873-5p group were higher than those in the sh-circ_0023990+anti-miR-NC group (<0.05), and the sensitization ratios were 0.305, 0.441, respectively. The survival scores of TPC-1 and KTC-1 cells in the miR-873-5p group were lower than those in the miR-NC group (<0.05), and the sensitization ratios were 2.044, 1.653 respectively. The survival scores of TPC-1 and KTC-1 cells in the miR-873-5p+pcDNA-ANXA2 group was higher than that in the miR-873-5p+pcDNA group (<0.05), and the sensitization ratios were 0.496, 0.686, respectively. The expression of γH2AX protein in TPC-1 and KTC-1 cells of the 4 Gy+sh-circ_0023990 group were higher than that in the 4 Gy+sh-NC group (2.68±0.27 vs. 1.87±0.25, 2.46±0.19 vs. 1.77±0.14; all <0.05), but the expression of γH2AX protein in TPC-1 and KTC-1 cells of the 4 Gy+sh-circ_0023990+anti-miR-873-5p group were lower than that in the 4 Gy+sh-circ_0023990+anti-miR-NC group (1.13±0.09 vs. 1.69±0.09, 1.11±0.08 vs. 1.60±0.08; both <0.05). The expression of γH2AX protein in TPC-1 and KTC-1 cells in the 4 Gy+miR-873-5p group were higher than that in the 4 Gy+miR-NC group (2.35±0.16 vs. 1.84±0.14, 2.26±0.12 vs. 1.77±0.13; both <0.05), but the expression of γH2AX protein in TPC-1 and KTC-1 cells of the 4 Gy+miR-873-5p+pcDNA-ANXA2 group were lower than that in the 4 Gy+miR-873-5p+pcDNA group (1.96±0.12 vs. 2.41±0.12, 1.92±0.07 vs. 2.28±0.12; both <0.05). circ_0023990 targeted the negative regulation of miR-873-5p, and ANXA2 was the target gene of miR-873-5p. circ_0023990 was highly expressed in thyroid cancer tissues and cell lines, and it may promote the radiotherapy resistance of thyroid cancer cells in vivo through regulating miR-873-5p/ANXA2 axis.
探讨circ_0023990对甲状腺癌细胞放射敏感性的影响及可能机制。采用qRT-PCR检测55例甲状腺癌患者癌组织及甲状腺癌细胞系（TPC-1、KTC-1、FTC-133和CAL-62）中circ_0023990的表达，并分析癌组织中circ_0023990表达与患者临床特征的关系。将甲状腺癌细胞TPC-1和KTC-1分为sh-circ_0023990组、sh-NC组、sh-circ_0023990+anti-miR-873-5p组、sh-circ_0023990+anti-miR-NC组、miR-873-5p组、miR-NC组、miR-873-5p+pcDNA-ANXA2组和miR-873-5p+pcDNA组，然后采用克隆形成实验检测细胞放射敏感性。每组细胞经4Gy辐射后，采用蛋白质免疫印迹法检测细胞中γH2AX蛋白的表达。双荧光素酶报告基因实验验证circ_0023990与miR-873-5p或miR-873-5p与ANXA2之间的靶向关系。circ_0023990在甲状腺癌组织中的表达高于正常组织（2.15±0.09 vs. 0.97±0.05，<0.05），其表达与甲状腺癌患者的肿瘤大小、淋巴结转移及TNM分期密切相关（<0.05）。circ_0023990在甲状腺癌细胞系（TPC-1、KTC-1、FTC-133和CAL-62）中的表达高于正常甲状腺细胞HTori-3（3.16±0.38、2.63±0.28、1.82±0.24、1.71±0.22 vs. 1.00±0.10，均<0.05）。sh-circ_0023990组中TPC-1和KTC-1细胞的存活分数显著低于sh-NC组（<0.05），敏化率分别为2.482、1.643；sh-circ_0023990+anti-miR-873-5p组中TPC-1和KTC-1细胞的存活分数高于sh-circ_0023990+anti-miR-NC组（<0.05），敏化率分别为0.305、0.441。miR-873-5p组中TPC-1和KTC-1细胞的存活分数低于miR-NC组（<0.05），敏化率分别为2.044、1.653。miR-873-5p+pcDNA-ANXA2组中TPC-1和KTC-1细胞的存活分数高于miR-873-5p+pcDNA组（<0.05），敏化率分别为0.496、0.686。4Gy+sh-circ_0023990组中TPC-1和KTC-1细胞中γH2AX蛋白的表达高于4Gy+sh-NC组（2.68±0.27 vs. 1.87±0.25，2.46±0.19 vs. 1.77±0.14；均<0.05），但4Gy+sh-circ_0023990+anti-miR-873-5p组中TPC-1和KTC-1细胞中γH2AX蛋白的表达低于4Gy+sh-circ_0023990+anti-miR-NC组（1.13±0.09 vs. 1.69±0.09，1.11±0.08 vs. 1.60±0.08；均<0.05）。4Gy+miR-873-5p组中TPC-1和KTC-1细胞中γH2AX蛋白的表达高于4Gy+miR-NC组（2.35±0.16 vs. 1.84±0.14，2.26±0.12 vs. 1.77±0.13；均<0.05），但4Gy+miR-873-5p+pcDNA-ANXA2组中TPC-1和KTC-1细胞中γH2AX蛋白的表达低于4Gy+miR-873-5p+pcDNA组（1.96±0.12 vs. 2.41±0.12，1.92±0.07 vs. 2.28±0.12；均<0.05）。circ_0023990靶向负调控miR-873-5p，ANXA2是miR-873-5p的靶基因。circ_0023990在甲状腺癌组织和细胞系中高表达，其可能通过调控miR-873-5p/ANXA2轴促进甲状腺癌细胞的放疗抵抗。
