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. 2018 May;22(5):2569-2579.
doi: 10.1111/jcmm.13499. Epub 2018 Mar 8.

Effect of CLIC1 gene silencing on proliferation, migration, invasion and apoptosis of human gallbladder cancer cells

Yue-Ming He  1 , Zhong-Lin Zhang  1 , Quan-Yan Liu  1 , Yu-Sha Xiao  1 , Lei Wei  1 , Chen Xi  2 , Xiang Nan  2
Affiliations

Effect of CLIC1 gene silencing on proliferation, migration, invasion and apoptosis of human gallbladder cancer cells

Yue-Ming He et al. J Cell Mol Med. 2018 May.

Retraction in

  • Retraction.
    [No authors listed] [No authors listed] J Cell Mol Med. 2021 May;25(9):4522. doi: 10.1111/jcmm.16517. J Cell Mol Med. 2021. PMID: 33949111 Free PMC article. No abstract available.

Abstract

This study aimed to explore the effects of CLIC1 gene silencing on proliferation, migration, invasion and apoptosis of human gallbladder cancer (GBC). GBC and normal gallbladder tissues were extracted for the detection of mRNA and protein expressions of CLIC1. GBC-SD and NOZ cells in the logarithmic growth phase were selected to conduct the experiment. Three different siRNA recombined expression vectors were established using CLIC1 as a target at different sites. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) and Western blotting were, respectively, used to detect the CLIC1 mRNA and protein expressions. MTT assay was performed to detect the cell proliferation. Flow cytometry was applied to measure the cell apoptosis and cell cycle distribution. The variations of cell migration and invasion were evaluated using Transwell assay. GBC tissues showed higher CLIC1 mRNA and protein expressions than normal gallbladder tissues. The CLIC1 mRNA and protein expressions in the CLIC1 siRNA group were significantly lower than those in the NC and blank groups. Compared with the NC and blank groups, the CLIC1 siRNA group showed a significant decrease in cell proliferation but an obvious increase in apoptosis rate in GBC cells. Besides, in the CLIC1 siRNA group, cell percentage in G0/G1 and G2/M phase was gradually increased but decreased in S phases. The migration and invasion abilities in GBC cells were significantly lower than those in the NC and blank groups. Our study demonstrates that CLIC1 gene silencing could promote apoptosis and inhibit proliferation migration and invasion of GBC cells.

Keywords: CLIC1 gene silencing; GBC-SD cell; apoptosis; gallbladder cancer; invasion; migration; proliferation.

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Figures

Figure 1
Figure 1
CLIC1 mRNA and protein expressions in GBC and normal gallbladder tissues. (A) RT‐qPCR of CLIC1 mRNA expression; (B) statistical results of CLIC1 protein expression; (C) Western blotting of CLIC1 protein expression; *compared with normal gallbladder tissues, P < 0.05.
Figure 2
Figure 2
CLIC1 expressions in GBC‐SD, EH‐GB1, NOZ and SGC‐996 cell lines. *compared with GBC‐SD and NOZ cell lines, P < 0.05.
Figure 3
Figure 3
mRNA expression of CLIC1 at different time‐points in the CLIC1 siRNA, NC and blank groups. (A) CLIC1 mRNA expression in GBC‐SD cells at 24, 48 and 72 hrs after transfection; (B) CLIC1 mRNA expression in NOZ cells at 24, 48 and 72 hrs after transfection; *compared with the NC and blank groups, P < 0.05.
Figure 4
Figure 4
Protein expression of CLIC1 at different time‐points in the CLIC1 siRNA, NC and blank groups. (A) Western blotting of CLIC1 protein expression in GBC‐SD cells; (B) statistical results of CLIC1 protein expressions in GBC‐SD cells; (C) Western blotting of CLIC1 protein expression in NOZ cells; (D) statistical results of CLIC1 protein expressions in NOZ cells; *compared with the NC and blank groups, P < 0.05.
Figure 5
Figure 5
Transfection efficiency and growth state of GBC‐SD and NOZ cells in the CLIC1 siRNA, NC and blank groups under a fluorescent inverted phase contrast microscope.
Figure 6
Figure 6
Cell proliferations in the CLIC1 siRNA, NC and blank groups by MTT assay. (A) cell proliferation rate in GBC‐SD cells; (B) cell proliferation rate in NOZ cells; *compared with the NC and blank groups, P < 0.05.
Figure 7
Figure 7
Cell apoptosis at 48 hrs after transfection in the CLIC1 siRNA, NC and blank groups by flow cytometry. (A, C) cell apoptosis of GBC‐SD and NOZ in the CLIC1 siRNA, NC and blank groups by flow cytometry; (B, D) the bar chart of GBC‐SD and NOZ cells apoptosis rate in the CLIC1 siRNA, NC and blank groups; *compared with NC and blank groups, P < 0.05.
Figure 8
Figure 8
Cell cycle distributions at 48 hrs after transfection in the CLIC1 siRNA, NC and blank groups by flow cytometry. (A, C) flow cytometry results of GBC‐SD and NOZ cell migration (200×); (B, D) statistical results of GBC‐SD and NOZ cell cycle; *compared with the NC and blank groups, P < 0.05.
Figure 9
Figure 9
Cell migrations at 48 hrs after transfection in the CLIC1 siRNA, NC and blank groups by Transwell assay. (A, C) results of GBC‐SD and NOZ cell migration under microscope (200×); (B, D) statistical results of GBC‐SD and NOZ cell migration; *compared with the NC and blank groups, P < 0.05.
Figure 10
Figure 10
Cell invasions at 48 hrs after transfection in the CLIC1 siRNA, NC and blank groups by Transwell assay. (A, C) results of GBC‐SD and NOZ cell invasion in three groups under a microscope (200×); (B, D) statistical results of GBC‐SD and NOZ cell invasion in three groups; *compared with the NC and blank groups, P < 0.05.
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