Downregulation of EVI1 Expression Inhibits Cell Proliferation and Induces Apoptosis in Hilar Cholangiocarcinoma via the PTEN/AKT Signalling Pathway

Abstract

Aims: Hilar cholangiocarcinoma (HCCA) is a tumour with high malignancy, low surgical resection potential, and a poor prognosis. Ecotropic Viral Integration site 1 (EVI1) is a transcriptional regulator that has been proven to be associated with tumourigenesis and progression in many human solid tumours. However, the expression of EVI1 and its role in HCCA progression remain unclear. The aim of this study was to clarify the association between EVI1 expression and clinical outcomes in patients with HCCA. Methods: The expression of EVI1 in HCCA tissue samples and cell lines was examined by quantitative real-time PCR (qRT-PCR), Western blotting, and immunohistochemistry (IHC). Kaplan-Meier analysis was used for survival analysis. A log-rank test was performed for univariate analysis of survival, and a Cox regression model was utilized for multivariate analysis of survival. Cell proliferation was measured by cell counting kit-8 (CCK-8), colony formation, and 5-ethynyl-2'-deoxyuridine (EdU) assays. The cell cycle was evaluated by flow cytometry. Cell apoptosis was detected by flow cytometry and a terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick-end labelling (TUNEL) assay. In vivo tumour growth was observed for xenografts in nude mice. Results: EVI1 expression was upregulated in HCCA tissue samples and correlated with a poor prognosis. In clinical specimens, the expression of EVI1 correlated with tumour histological grade and tumour size. Knocking down EVI1 expression reduced HCCA cell proliferation, blocked cell cycle progression, and promoted apoptosis in vitro and in vivo. Furthermore, we found that EVI1 could regulate the AKT signalling pathway by regulating PTEN levels in HCCA. Conclusion: Our data revealed that EVI1 played important roles in HCCA tumourigenesis and development. Our findings suggest that EVI1 may be a potentially useful therapeutic target in HCCA.

siRNA-dependent Gene Silencing of EVI1 Decreased Cell Proliferation in MKN45 Cells

“…Next, we found that EVI1 depletion inhibited cell proliferation by inducing apoptosis in MKN45 cells. This result is consistent with various cancer studies that showed EVI1 promoted cell growth by faster advancement through cell cycle and suppression of apoptosis [20,21]. In an experiment with EVI1 in hematopoietic cells in mice, EVI1 overexpression was accompanied by increased cdk2 activity, heightened levels of hyperphosphorylated Rb protein, decreased p27 protein levels, and shortened G1 phase [22].…”

“…In Hilar Cholangiocarcinoma (HCCA), EVI1 repression reduced cell proliferation, promoted apoptosis, and blocked cell cycle progression. EVI1 could also regulate PTEN levels in the AKT signaling pathway of HCCA [20]. Since the genes related to cell cycle and apoptosis were not investigated in this paper, the detailed molecular mechanism of EVI1 with cell cycle and apoptosis regulating genes should be further analyzed in the future.…”

SRPK1/2 and PP1α exert opposite functions by modulating SRSF1-guided MKNK2 alternative splicing in colon adenocarcinoma

“…The proliferation assay colony formation assay were conducted as previously described [ 13 , 14 ]. Proliferation capacity was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assays at designated time points (6 h, 24 h, 48 h, 72 h, 96 h).…”

“…Tumor size was measured every 5 days using the following formula: V = (length×width 2 )/2. The mice were sacrificed on the 21st day after injection, and the xenograft tumors were removed and weighed as described before [ 13 ]. The Animal Welfare Committee of Shandong University approved all procedures involving animals.…”

EVI1 in Leukemia and Solid Tumors

“…These results indicate that the interaction between the PTEN/AKT/mTOR signaling pathway and the EVI1-polycomb complexes could be potential therapeutic targets for leukemia with activated EVI1 [ 30 ]. Knockdown of EVI1 also represses cell proliferation and promotes apoptosis via the PTEN/AKT signaling pathway in hilarcholangiocarcinoma [ 31 ].…”