奇果菌素调控miR-646/ADAM17通路对白血病细胞增殖、凋亡的影响及机制  

Experimental study on the effects of grifolin on the proliferation and apoptosis of leukemic cell by regulating miR-646/ADAM17 pathway

作　　者：努尔阿米娜·依明尼亚孜[1] 帕力旦·艾沙 阿依姆妮萨·阿卜杜热合曼[1] Nur Amina Iminyazi;Palidan Esha;Aimnesa Abdul Rehmann(Department of Hematology,The First People’s Hospital of Kashi District,Xinjiang, Kashi 844000,China)

机构地区：[1]新疆维吾尔自治区喀什地区第一人民医院血液科,844000

出　　处：《河北医药》2020年第24期3702-3707,共6页Hebei Medical Journal

基　　金：新疆维吾尔自治区自然科学基金(青年项目)(编号:2018D01C017)。

摘　　要：目的探讨奇果菌素(Gri)对白血病细胞增殖、凋亡的影响及其作用机制。方法体外培养白血病K562细胞,经不同浓度的Gri处理细胞后,采用甲基噻唑基四唑(MTT)法测定Gri对细胞增殖的抑制作用;流式细胞术检测细胞凋亡;实时荧光定量PCR(qRT-PCR)与蛋白免疫印迹法(Western blot)检测Gri对微小RNA-646(miR-646)与解聚素-金属蛋白酶17(ADAM17)表达的影响;双荧光素酶报告基因检测miR-646与ADAM17的靶向作用关系;观察miR-646过表达或干扰ADAM17表达对细胞增殖及凋亡的影响,以及抑制miR-646表达后细胞抑制率及凋亡率的变化;Western blot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Gri呈剂量依赖性抑制K562细胞增殖(P<0.05),细胞凋亡率随着药物浓度的增加而显著增高(P<0.05),促进miR-646、p21、Bax表达(P<0.05),抑制ADAM17、CyclinD1、Bcl-2表达(P<0.05);双荧光素酶报告实验证实miR-646可负性调控ADAM17的表达;miR-646过表达或干扰ADAM17表达后可显著抑制细胞增殖,促进细胞凋亡,抑制miR-646表达可逆转Gri对细胞增殖、凋亡的作用。结论奇果菌素可通过调控miR-646/ADAM17通路抑制细胞增殖及促进其凋亡从而发挥抗白血病作用。Objective To investigate the effects of grifolin(Gri)on the proliferation and apoptosis of leukemic cell by regulating miR-646/ADAM17 pathway,and to explore its action mechanism.Methods The leukemia K562 cells were cultured in vitro,and the cells were treated with different concentrations of Gri,and the inhibitory effects of Gri on cell proliferation were detected by methylthiazolyl tetrazolium(MTT),and cell a poptosis was detected by flow cytometry.Real-time quantitative PCR(qRT-PCR)and protein immunosorbent assay(Western Blot)were used to detect the effects of Gri on the expression levels of microRNA-646(miR-646)and disintegrin-metalloproteinase 17(ADAM17).The dual luciferase reporter gene was used to detect the targeting correlation between miR-646 and ADAM17.Moreover the effects of miR-646 overexpression or interference with ADAM17 expression on cell proliferation and apoptosis were observed,and the changes of cell inhibition rate and apoptosis rate after miR-646 expression was inhibited.Western Blot was used to detect the expression levels of cyclin 1(CyclinD1),B lymphocyte tumor-2(Bcl-2),p21,and B lymphoma-2 related protein(Bax).Results Gri inhibited the proliferation of K562 cells in a dose-dependent manner(P<0.05),and the apoptosis rate was increased significantly with the increase of drug concentration(P<0.05),which promoted the expressions of miR-646,p21 and Bax(P<0.05),however,the expression levels of ADAM17,CyclinD1 and Bcl-2 were inhibited(P<0.05).Dual luciferase reporter experiments confirmed that miR-646 could negatively regulate the expression of ADAM17.In addition the overexpression of miR-646 or interference with the expression of ADAM17 could significantly inhibit the cell proliferation,promote apoptosis,and inhibit the expression of miR-646,which could reverse the effects of Gri on cell proliferation and apoptosis.Conclusion Grifolin can exert anti-leukemia effects by inhibiting cell proliferation and promoting apoptosis through regulating miR-646/ADAM17 pathway.

关 键 词：奇果菌素 白血病 miR-646 ADAM17 增殖 凋亡 

分 类 号：R733.7[医药卫生—肿瘤]