右美托咪定调控miR-526b-3p/S100A4表达影响胰腺癌细胞增殖及凋亡的机制研究

Mechanism of Dexmedetomidine in Influencing the Proliferation and Apoptosis of Pancreatic Cancer Cells by Regulating miR-526b-3p/S100A4 Expression

目的:探讨右美托咪定(Dex)对胰腺癌细胞增殖、凋亡的影响及其可能作用机制。方法:应用MTT法检测Dex对人胰腺癌Capan-1细胞增殖的抑制作用;采用流式细胞术检测细胞凋亡率;实时荧光定量聚合酶链反应(qRT-PCR)与蛋白免疫印迹法检测Dex对微小RNA-526b-3p(miR-526b-3p)和钙结合蛋白A4(S100A4)表达的影响;检测miR-526b-3p过表达与干扰S100A4表达对Capan-1细胞增殖及凋亡的影响;双荧光素酶报告实验验证miR-526b-3p与S100A4的靶向作用关系;抑制miR-526b-3p表达验证Dex对Capan-1细胞增殖和凋亡的作用机制;Western blot法检测细胞周期蛋白1(CyclinD1)、B淋巴细胞瘤-2(Bcl-2)、p21、B淋巴细胞瘤-2相关蛋白(Bax)蛋白表达。结果:用不同浓度的Dex处理后,细胞抑制率、细胞凋亡率与miR-526b-3p、p21、Bax的表达水平显著高于对照组(P<0.05),S100A4、CyclinD1、Bcl-2的表达水平显著降低(P<0.05),且呈明显的浓度依赖性(P<0.05);双荧光素酶报告实验与Western blot实验证实miR-526b-3p与S100A4存在靶向结合关系,且miR-526b-3p可负性调控S100A4的表达;miR-526b-3p过表达或干扰S100A4表达后细胞抑制率、细胞凋亡率与p21、Bax的表达水平显著升高(P<0.05),CyclinD1、Bcl-2的表达水平显著降低(P<0.05);抑制miR-526b-3p表达可逆转Dex对Capan-1细胞增殖与凋亡的作用。结论:Dex可通过调控miR-526b-3p/S100A4表达抑制胰腺癌细胞增殖并诱导细胞凋亡。

Objective: To investigate the effect of dexmedetomidine( Dex) on the proliferation and apoptosis of pancreatic cancer cells and its possible mechanism. Methods: The inhibitory effect of Dex on the proliferation of human pancreatic cancer Capan-1 cells was detected by MTT assay. The apoptosis rate was measured by flow cytometry. The effects of Dex on the expressions of miR-526b-3p and S100A4 were detected by qRT-PCR and Western blot. The effect of miR-526b-3p overexpression and the interference with S100A4 expression on the proliferation and apoptosis of Capan-1 cells was examined. The dual luciferase reporter assay validated the targeting relationship between miR-526b-3p and S100A4. The inhibition of miR-526b-3p expression validated the action mechanism of Dex on the proliferation and apoptosis of Capan-1 cells. The expressions of CyclinD1,Bcl-2,p21 and Bax protein were detected by Western blot. Results: After the treatment with different concentrations of Dex,the cell inhibition rate,apoptosis rate and expression levels of miR-526b-3p,p21 and Bax were significantly higher than those of the control group( P<0.05),and the expression levels of S100A4,CyclinD1 and Bcl-2 were significantly decreased( P<0.05),and showed a significant concentration-dependence( P<0.05). The dual luciferase reporter assay and Western blot assay confirmed the targeted binding relationship between miR-526b-3p and S100A4,and miR-526b-3p negatively regulated the expression of S100A4. After the overexpression of miR-526b-3p or the interference with S100A4 expression,the cell inhibition rate,apoptosis rate and expression levels of p21 and Bax were significantly increased( P< 0.05),and the expression levels of CyclinD1 and Bcl-2 were significantly decreased( P<0.05). The inhibition of miR-526b-3p expression reversed the effect of Dex on the proliferation and apoptosis of Capan-1 cells. Conclusion: Dex inhibits the proliferation of pancreatic cancer cells and induces the apoptosis by regulating miR-526b-3p/S100A4 expression.