Journal of Shanghai Jiao Tong University (Medical Science) >

2023 , Vol. 43 >Issue 9: 1071 - 1079

DOI: https://doi.org/10.3969/j.issn.1674-8115.2023.09.001

Basic research

Effect of BRCA1 R1325K mutation on proliferation and apoptosis of gallbladder cancer cells

  • Jingxiao YANG ,
  • Ziyao JIA ,
  • Wenguang WU ,
  • Xiangsong WU ,
  • Fei ZHANG ,
  • Huaifeng LI ,
  • Yidi ZHU ,
  • Maolan LI
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  • 1.Department of Biliary-Pancreatic Surgery, Renji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200127, China
    2.Department of General Surgery, Xinhua Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200092, China
    3.Shanghai Key Laboratory of Biliary Tract Disease Research, Shanghai 200092, China
Li Maolan, E-mail: limaolan6@163.com.

Received date: 2023-01-03

  Accepted date: 2023-08-29

  Online published: 2023-09-28

Supported by

National Key Research and Development Program of China(2021YFE0203300);National Natural Science Foundation of China(82073206);Shanghai Municipal Health Commission Health Industry Clinical Research Special Project(20224Z0014);Shuguang Program of Shanghai Education Development Foundation and Shanghai Municipal Education Commission(20SG14);Basic Research Project of Science and Technology Commission of Shanghai Municipality(20JC1419100);Clinical Science and Technology Innovation Project of Shanghai Shenkang Hospital Development Center(SHDC12019110);Shanghai Clinical Medical Research Center for Digestive Diseases(19MC1910200);“Two-hundred Talents” Program of Shanghai Jiao Tong University School of Medicine(20181808)

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Abstract

Objective ·To investigate the effects of breast cancer susceptibility gene 1 (BRCA1) R1325K mutation [arginine (R) to lysine (K) mutation at amino acid 1325] on the proliferation and apoptosis of gallbladder cancer cell lines GBC-SD and NOZ. Methods ·BRCA1 wild-type overexpression lentivirus, BRCA1 R1325K mutation overexpression lentivirus, and negative control lentivirus were used to construct the stable transgenic strains of gallbladder carcinoma, cell lines GBC-SD and NOZ. The cells were divided into the control group without the target gene, the BRCA1 wild-type group, and the BRCA1 R1325K mutation group. The expression of target protein was verified by Western blotting. The BRCA1 R1325K mutant gallbladder cancer cells were treated with 20 μmol/L Olaparib, a BRCA1 mutation inhibitor. Gallbladder cancer cell lines were divided into the control group, the BRCA1 wild-type group, the BRCA1 R1325K mutation group, and the BRCA1 R1325K mutation+Olaparib group according to the target gene expression and whether or not the inhibitor was added. The effect of BRCA1 R1325K mutation on proliferation and clonogenesis ability of gallbladder cancer cell lines GBC-SD and NOZ was observed by CCK8 assay and clonogenesis assay, respectively. The effect of BRCA1 R1325K mutation on apoptosis of gallbladder cancer cell lines GBC-SD and NOZ was observed by TUNEL assay. The expressions of apoptosis-related proteins, cleaved PARP, Bcl-2 and Bax, were detected by Western blotting. The inhibitor Olaparib was used to treat the BRCA1 R1325K mutant gallbladder cancer cell lines GBC-SD and NOZ. The phenotypic changes (promoting proliferation, enhancing clonogenesis and inhibiting apoptosis) induced by BRCA1 R1325K mutation were tested in the presence of Olaparib to determine whether the changes could be reversed by the inhibitor. Results ·The results of CCK8 assay and clonogenesis assay showed that BRCA1 R1325K mutation could promote the proliferation of gallbladder cancer cell lines GBC-SD and NOZ, and improve their clonal formation ability, compared with the control group and the BRCA1 wild-type group. Olaparib inhibited the proliferation of gallbladder cancer cell lines overexpressing BRCA1 R1325K mutation (P<0.05). Through TUNEL and Western blotting, it was found that overexpression of wild-type BRCA1 could induce the apoptosis of gallbladder cancer cell lines GBC-SD and NOZ, compared with the control group. Compared with the control group and the BRCA1 wild-type group, the BRCA1 R1325K mutation group had anti-apoptotic effect, in which the expression of apoptosis-inhibiting protein Bcl-2 increased and the expression of pro-apoptotic protein Bax decreased (P<0.05). Conclusion ·BRCA1 R1325K mutation can promote the proliferation of GBC-SD and NOZ cell lines and inhibit their apoptosis.

Key words： gallbladder cancer; BRCA1; mutation; proliferation; apoptosis

Cite this article

Jingxiao YANG , Ziyao JIA , Wenguang WU , Xiangsong WU , Fei ZHANG , Huaifeng LI , Yidi ZHU , Maolan LI . Effect of BRCA1 R1325K mutation on proliferation and apoptosis of gallbladder cancer cells[J]. Journal of Shanghai Jiao Tong University (Medical Science), 2023 , 43(9) : 1071 -1079 . DOI: 10.3969/j.issn.1674-8115.2023.09.001

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