SIRT1对葡萄膜黑色素瘤细胞增殖的表观遗传调控

doi:10.3760/cma.j.cn115909-20210702-00259

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Abstract Objective: To investigate the effects of the silent information regulator protein1 (SIRT1) on the proliferation of uveal melanoma cells. Methods: In this experimental research, the expression of SIRT1 was detected in uveal melanoma cells (M23, SP6.5) and uveal melanocytes through Western blot and realtime quantitative PCR analysis. M23 and SP6.5 were treated with EX527 to inhibit SIRT1 activity and SIRT1 siRNA transfection to inhibit SIRT1 expression. DMSO treatment and siNC transfection were used in the control group. MTS, flow cytometry and colony formation assays were used to evaluate cell viability, cell cycle and colony formation ability. A mouse tumor-genesis experiment tested the ability of tumor formation in vivo. Proliferation-associated proteins were detected by Western blot analysis. Data were analyzed by independent t test. Results: SIRT1 was highly expressed in M23 and SP6.5 at mRNA (t=17.08, P<0.001; t=13.24, P<0.001) and protein (t=6.26, P=0.008) levels. MTS assay showed that the viability of M23 and SP6.5 was inhibited by EX527 and depended on the concentrations; siSIRT1 transfection also inhibited viability significantly (t=5.94, P<0.001; t=10.73, P<0.001). Flow cytometry revealed that EX527 treatment resulted in G1 phase arrest in M23 and SP6.5 (t=5.10, P=0.047; t=7.57, P=0.002); the percentage of cells in the G1 stage were markedly high in the siSIRT1 transfection group compared to the NC group (t=6.41, P=0.003; t=6.10, P=0.004). Compared to the control groups, the clone numbers of M23 and SP6.5 cells in the EX527 group (t=5.04, P=0.001; t=5.93, P<0.001) and siSIRT1 transfected group (t=11.44, P<0.001; t=8.24, P<0.001) were obviously decreased. In addition, siSIRT1 transfection markedly suppressed the tumor-genesis in vivo (t=5.50, P<0.001). Western blot analysis showed that the expression levels of E2F1 (t=12.10, P=0.003; t=5.96, P=0.004), CYCLIND2 (t=36.28, P<0.001; t=16.58, P<0.001), p-RB (t=17.55, P<0.001; t=21.34, P<0.001), as well as p-AKT were significantly decreased while P21 (t=4.63, P=0.010; t=7.90, P=0.001) was increased in EX527-treated cells. In siSIRT1 transfected cells, E2F1 (t=4.60, P=0.01; t=4.89, P=0.008), CYCLIND2 (t=5.13, P=0.007; t=5.63, P=0.005), p-RB (t=4.45, P=0.011; t=6.53, P=0.003), and p-AKT (t=9.61, P<0.001; t=6.44, P=0.003) decreased but P21 (t=5.88, P=0.004; t=10.51, P<0.001) increased significantly. Conclusions: Inhibition of SIRT1 activity or expression suppresses UM cell proliferation, suggesting SIRT1 is a promising target for the treatment of this malignancy.

Key words： Silent information regulator protein 1 EX527 uveal melanoma cells cell proliferation

Received: 02 July 2021

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Fund:National Natural Science Foundation of China (81900818); Foundation of Wenzhou Science & Technology Plan Project (Y20170761)

Corresponding Authors: Dongsheng Yan, State Key Laboratory of Ophthalmology, Optometry and Vision Science; Eye Hospital, Wenzhou Medical University, Wenzhou 325027, China (Email: dnaprotein@hotmail. com)

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Yong Lin,Li Li,Qi Liu等. The Epigenetic Regulation of SIRT1 on the Proliferation of Uveal Melanoma Cells[J]. Chinese Journal of Optometry Ophthalmology and Visual science, 2022, 24(5): 329-336.

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https://www.cjoovs.com/EN/10.3760/cma.j.cn115909-20210702-00259 OR https://www.cjoovs.com/EN/Y2022/V24/I5/329

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