ABSTRACT
Objective:To investigate the effect of ubiquitin binding enzyme 2T (UBE2T) on the radiosensitivity of lung adenocarcinoma and unravel its possible mechanism.Methods:A total of 45 patients pathologically diagnosed with different stages of lung adenocarcinoma and treated with radiotherapy in the Second Affiliated Hospital of Zunyi Medical University from March, 2019 to December, 2021 were enrolled, and the efficacy was evaluated according to response evaluation criteria in solid tumors (RECIST1.1). All patients were divided into radiosensitive group ( n=25) and radioresistant group ( n=20). Radiosensitive group was complete remission (CR)+partial remission (PR), and radioresistant group was stable disease (SD) + progression disease (PD). Immunohistochemistry (IHC) was used to calculate the score based on the staining intensity and the number of positive cells. Chi-square test was combined to analyze the correlation between the expression level of UBE2T in paraffin specimens of lung adenocarcinoma patients and the radiosensitivity of patients. Lentivirus UBE2T-interfered (UBE2Tsh) A549 and UBE2T-overexpressed SPC-A-1 lung adenocarcinoma cells and their respective controls were constructed for irradiation and colony formation assay. The survivor fraction curve was fitted by single-hit multi-target model. The DNA double-strand break (DSB) marker γH2AX foci were detected by immunofluorescence (IF). The expression levels of UBE2T, γH 2AX and Rad51 proteins were detected by Western blot. Cell cycle and apoptosis rate of A549 were determined by flow cytometry. Binary variables were statistically analyzed by Fisher's exact probability method and measurement data were assessed by t-test. Results:High-expression level of UBE2T was correlated with the radiosensitivity of lung adenocarcinoma patients ( P<0.05). UBE2Tsh improved the radiosensitivity of A549 lung adenocarcinoma cells, and the sensitizing enhancement ratio (SER) was 1.795. UBE2T overexpression decreased the radiosensitivity of SPC-A-1 lung adenocarcinoma cells with an SER of 0.293. γH2AX foci number per cell were significantly increased in UBE2Tsh A549 cells after irradiation ( P<0.01) . Compared with the control group, the expression level of γH2AX protein was up-regulated ( P<0.01)and that of Rad51 protein was down-regulated in UBE2Tsh A549 cells after radiation ( P<0.001). Compared with the control group, the expression level of γH2AX protein was down-regulated ( P<0.05) and that of Rad51 protein was up-regulated in UBE2T overexpressed SPC-A-1 cells ( P<0.001). The proportion of UBE2Tsh A549 cells in G 2 phase was decreased ( P<0.01) and cell apoptosis was increased ( P<0.001). Conclusions:UBE2T might promote the radioresistance of lung adenocarcinoma cells by enhancing DNA DSB repair induced by radiotherapy, inducing cell cycle G 2 phase arrest, and reducing cell apoptosis.
ABSTRACT
Objective: To investigate the expression and clinical significances of ubiquitin conjugating enzyme E2 T (UBE2T) in hepatocellular carcinoma (HCC), and explore the role of UBE2T gene expression in migration and invasion of HCC cells. Methods: The expression of UBE2T mRNA in 54 pairs of HCC tissues and the matched noncancerous liver tissues was analyzed by real-time fluorescent quantitative PCR. The expression level of UBE2T protein in 233 cases of HCC tissues was detected by immunohistochemistry, then the relationship between UBE2T expression and clinicopathological features of HCC patients was analyzed. The recombinant lentiviral vector pWPXL-UBE2T contained UBE2T gene sequences was constructed and further transfected into Hep3B and SMMC-7721 cells. In the other hand, the recombinant lentiviral vector GV248-shUBE2T-1 and GV248-shUBE2T-2 targeted UBE2T gene were transfected into MHCC-LM3 and SK-Hep1 cells. The forced expression and knockdown of UBE2T gene were validated by real-time fluorescent quantitative PCR and Western blotting. Afterwards, the effects of UBE2T gene overexpression and silencing on migratory and invasive abilities of HCC cells were detected by Transwell chamber assays, respectively. Results: The UBE2T mRNA level in HCC tissues was significantly higher than that in the matched noncancerous liver tissues (P < 0.000 1). The expression of UBE2T protein was correlated with intrahepatic metastasis of HCC (P = 0.004) in 233 patients with HCC. Compared with the control group, the forced expression of UBE2T gene markedly promoted both migration and invasion of Hep3B and SMMC-7721 cells (both P < 0.01); while the knockdown of UBE2T gene obviously inhibited the migration and invasion of MHCC-LM3 and SK-Hep1 cells as compared with the negative control group (both P < 0.05). Conclusion: High expression level of UBE2T in HCC tissues is correlated with the presence of intrahepatic metastasis, and the overexpression of UBE2T gene can promote the migration and invasion of HCC cells.