Expression change of miRNA-210 in plasma of patients with esophageal squamous cell carcinoma undergoing radiotherapy and its effect on esophageal squamous cancer Eca109 cells in vitro
10.3760/cma.j.cn115355-20190320-00091
- VernacularTitle:miRNA-210在食管鳞状细胞癌放疗患者血浆中表达的变化及其体外对食管鳞状细胞癌Eca109细胞的影响
- Author:
Chenglin LI
1
;
Yadi WANG
;
Fengfeng ZHOU
Author Information
1. 山东省临沂市人民医院肿瘤科 276000
- From:
Cancer Research and Clinic
2020;32(7):467-473
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the expression change of blood plasma miRNA-210 (miR-210) in blood plasma for patients with esophageal squamous cell carcinoma (ESCC) before and after radiotherapy and its effect on cell proliferation, cycle, apoptosis of ESCC cells in vitro.Methods:The blood specimens from 22 patients with newly diagnosed ESCC (ESCC group) before and after radical radiotherapy between December 2013 and March 2015 in Linyi People's Hospital of Shandong Province as well as 15 healthy controls (the healthy control group) were collected. miRNA-21 (miR-21) was treated as the positive controls. The real-time polymerase chain reaction (RT-PCR) was used to detect the expression level of miR-210 and miR-21 in blood plasma before and after radiotherapy for patients, and the expression of miR-21 in ESCC Eca109 cells at normoxia and hypoxia time. The cell proliferation, cycle and apoptosis of Eca109 cells in untransfected group (the blank control group), miR-21 transfected mimics negative control RNA group (negative control group) and miR-210 transfected mimics RNA group (miR-210 group) were detected by using EdU cell proliferation assay and flow cytometry.Results:A total of 59 plasma samples from ESCC group and the healthy control group were collected. The relative expression level [median ( P25, P75)] of blood plasma miR-210 and miR-21 in ESCC group before radiotherapy was higher than that in the healthy control group, and the difference was statistically significant [4.04×10 -4 (2.06×10 -4, 6.68×10 -4) vs. 0.54×10 -4 (0.28×10 -4, 0.77×10 -4), 397.07×10 -4 (181.77×10 -4, 742.93×10 -4) vs. 127.43×10 -4 (21.97×10 -4, 184.65×10 -4); U value was 37.0, 49.0, respectively, all P < 0.01]. The expression level of miR-210 and miR-21 before radiotherapy in ESCC group was not related with tumor location and differentiation degree (all P > 0.05). After the radical radiotherapy one week later, the relative expression level of miR-210 and miR-21 in blood plasma of ESCC group was 65.33×10 -4 (22.15×10 -4, 160.87×10 -4), 437.23×10 -4 (327.18×10 -4, 749.09×10 -4), respectively, which was higher compared with that before radiotherapy ( U value was 32.0, 154.0, respectively, both P < 0.05), and the increase of miR-210 was more significant. The expression level of miR-210 after hypoxic cultured Eca109 cells for 12 h was increased compared with the normal oxygen with the peak value 12 h later, and the difference was statistically significant ( P < 0.01). EdU cell proliferation assay showed that the Eca109 cell proliferative activity after transfection of 24 h and 48 h in miR-210 group was decreased compared with the negative control group and the blank control group, and the difference was statistically significant (all P < 0.01). Flow cytometry analysis showed that the proportion of G 2/M phase of Eca109 cells in miR-210 group after transfection of 24 h was increased compared with the negative control group and the blank control group, and the difference was statistically significant ( P < 0.05). After transfection of 48 h, the increased proportion in G 2/M phase was more obvious ( P <0.01); there was no statistically difference in the apoptotic cell proportion among three groups after transfection of 24 h and 48 h (all P > 0.05). Conclusions:miR-210 is highly expressed in the blood plasma of ESCC patients, especially the significant increase in the expression level of miR-210 in blood plasma after radical radiotherapy. miR-210 is highly expressed in hypoxic ESCC Eca109 cells. The overexpression of miR-210 can inhibit cell proliferation and its mechanism may be related with cell arrest in G 2/M phase.
