miR-381-3p对口腔癌细胞增殖、侵袭和迁移能力的调控作用及其机制OACSTPCD
Regulatory effects and mechanism of miR-381-3p on proliferation,invasion,and migration of oral cancer cells
目的 观察微小RNA(miR)-381-3p对口腔癌细胞增殖、侵袭、迁移能力的调控作用,并基于甲基转移酶SETDB1基因调控探讨相关机制.方法 采用RT-PCR法检测口腔癌细胞系CAL-27和正常口腔黏膜上皮细胞系ATCC中的miR-381-3p.培养CAL-27细胞,分为模拟物组、阴性对照组和未转染组,模拟物组和阴性对照组分别转染miR-381-3p模拟物和miR-381-3p阴性对照,未转染组不进行转染;采用CCK-8法检测细胞增殖能力,Transwell小室实验检测细胞侵袭能力,划痕愈合实验检测细胞迁移能力.采用TargetScan(http://www.targetscan.org/vert_80/)预测miR-381-3p与SETDB1结合位点,应用双荧光素酶报告基因分析进行验证;采用RT-PCR法检测CAL-27细胞和ATCC细胞中的SETDB1 mRNA,Western blotting法检测SETDB1蛋白;将CAL-27细胞分为模拟物组、阴性对照组和未转染组,模拟物组、阴性对照组分别转染miR-381-3p模拟物和阴性对照,未转染组不进行转染,采用RT-PCR法检测SETDB1 mRNA.结果 CAL-27细胞中miR-381-3p表达低于ATCC细胞(P<0.05).模拟物组在培养12、24、36、48 h时细胞增殖能力低于阴性对照组和未转染组(P均<0.05).模拟物组细胞迁移距离、穿膜细胞数均低于阴性对照组和未转染组(P均<0.05).SETDB1存在连续的、可以与miR-381-3p互补的核苷酸序列;共转染SETDB1-WT和miR-381-3p质粒的CAL-27细胞相对荧光素酶活性低于其他组细胞(P均<0.05);CAL-27细胞中SETDB1 mRNA和蛋白表达高于ATCC细胞(P均<0.05);模拟物组SETDB1 mRNA表达低于阴性对照组和未转染组(P均<0.05).结论 miR-381-3p能够抑制口腔癌细胞的增殖、侵袭和迁移能力,其作用机制可能与调节SETDB1表达有关.
Objective To observe the regulatory effects of miR-381-3p on the proliferation,invasion,and migration abilities of oral cancer cells,and to explore the relevant mechanism based on methyltransferases SETDB1 gene regulation.Methods The miR-381-3p in the oral cancer cell line CAL-27 and the normal oral mucosal epithelial cell line ATCC was detected by RT-PCR.CAL-27 cells were cultured and divided into the mimic,negative control and untransfected groups,and cells in the mimic and negative control groups were transfected with miR-381-3p mimic and miR-381-3p negative con-trol,respectively,and cells in the untransfected group were not transfected.cell proliferation ability was measured by CCK-8 assay,Transwell chamber assay was used to measure the cell invasion,and cell migration ability was determined by Scratch-healing assay.TargetScan(http://www.targetscan.org/vert_80/)was used to predict the binding sites be-tween miR-381-3p and SETDB1,and we tested this with the dual luciferase report gene analysis.The SETDB1 mRNA was detected by RT-PCR in CAL-27 cells and ATCC cells and SETDB1 protein was detected by Western blotting.CAL-27 cells were divided into the mimic group,negative control group and untransfected group,cells in the mimic and negative control groups were transfected with miR-381-3p mimic and miR-381-3p negative control,and cells in the untransfected group were not transfected.SETDB1 mRNA in each group was detected by RT-PCR.Results The expression of miR-381-3p was lower in CAL-27 cells than in ATCC cells(P<0.05).The cell proliferation abilities of miR-381-3p in the mim-ic group were lower than those of the miR-381-3p negative control group and untransfected group at 12,24,36 and 48 h(all P<0.05).The number of transmembrane cells and cell migration distance in miR-381-3p mimic group were lower than those in the miR-381-3p negative control group and untransfected group(all P<0.05).SETB1 gene had a continuous nucleotide sequence that could complement miR-381-3p.Relative luciferase activity of CAL-27 cells co-transfected with SETDB1-WT and miR-381-3p was lower than that of cells in the other groups(all P<0.05).The expression levels of SET-DB1 mRNA and protein in the CAL-27 cells were higher than those in the ATCC cells(both P<0.05);the expression of SETDB1 mRNA was lower in the mimic group than in the negative control group and the untransfected group(all P<0.05).Conclusion MiR-381-3p can inhibit the proliferation,invasion and migration of oral cancer cells,and its mechanism may be related to regulation of SETDB1 expression.
郝妍;白相宇;霍峰;陈喜波
承德医学院附属医院口腔科,河北承德 067000
临床医学
微小RNA-381-3p口腔癌SETDB1基因细胞增殖细胞侵袭细胞迁移
microRNA-381-3poral carcinomaSETDB1 genecell proliferationcell invasioncell migration
《山东医药》 2024 (020)
6-10 / 5
河北省医学科学研究课题计划项目(20210143).
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