摘要
目的探讨敲低蛋白质精氨酸甲基转移酶6(PRMT6)基因对胃癌细胞BGC823增殖的影响及机制。方法收集30例胃癌患者肿瘤组织和癌旁组织,免疫组织化学染色(IHC)检测PRMT6的蛋白表达;Western blot检测正常胃黏膜细胞和多种胃癌细胞株中PRMT6的表达水平;siRNA干扰胃癌细胞PRMT6的表达后,Western blot与qRT-PCR验证转染效果;MTT实验和集落克隆实验检测胃癌细胞增殖能力;Western blot与qRT-PCR检测下调PRMT6的表达对其下游细胞周期相关基因CDKN1A、CDKN1B、CDKN1C和CCND1表达的影响。结果免疫组化结果显示胃癌组织中的PRMT6的表达量显著高于癌旁组织,差异具有统计学意义(P<0.01);Western blot结果显示多株胃癌细胞中,BGC823细胞的PRMT6表达水平较高;siRNA显著降低细胞中PRMT6蛋白及mRNA的表达;MTT和集落克隆结果显示敲低PRMT6表达显著抑制胃癌细胞的增殖能力;敲低PRMT6表达后,qRT-PCR结果显示CDKN1A、CDKN1B和CDKN1C的mRNA表达水平与对照组相比无显著差异,Western blot与qRT-PCR结果显示其下游基因CCND1表达水平降低。结论PRMT6在胃癌组织中高表达,且可以促进胃癌细胞的增殖,机制可能与其调控CCND1的表达有关。
Objective To investigate the effect and its mechanism of the downregulation of protein arginine methyltransferase 6(PRMT6)gene on the proliferation of gastric cancer BGC823 cell.Methods The protein expression of PRMT6 was detected by immunohistochemical staining(IHC)in tumor and paracancer tissues of 30 patients with gastric cancer.Western blot was used to detect the expression levels of PRMT6 in normal gastric mucosa cells and various gastric cancer cell lines.After siRNA interfered with PRMT6 expression in gastric cancer cells,the transfection effect was verified by Western blot and qRT-PCR.MTT assay and colony cloning assay were used to detect the proliferation of gastric cancer cells.Western blot and qRT-PCR were used to detect the effect of downregulated PRMT6 expression on the expression of downstream cell cycle-related genes CDKN1A,CDKN1B,CDKN1C and CCND1.Results Immunohistochemical results showed that the expression level of PRMT6 in gastric cancer tissues was significantly higher than that in adjacent tissues,and the difference was statistically significant(P<0.01).Western blot results showed that PRMT6 expression level was higher in gastric cancer cell lines BGC823.SiRNA significantly reduced the expression of PRMT6 protein and mRNA in cells.MTT and colony cloning results showed that the downregulation of PRMT6 expression significantly inhibited the proliferation of gastric cancer cells.After knockdown PRMT6 expression,qRT-PCR results showed that there was no significant difference in mRNA expression levels of CDKN1A,CDKN1B and CDKN1C compared with the control group.Western blot and qRT-PCR results showed that the expression level of CCND1 downstream gene was decreased.Conclusion PRMT6 is highly expressed in gastric cancer tissues and can promote the proliferation of gastric cancer cells,which may be related to the regulation of CCND1 expression.
作者
杜军
陈玉忠
梁晚晴
韩冰清
李雷
马家驰
DU Jun(Department of Surgical Oncology,The First Affiliated Hospital of Bengbu Medical College,Bengbu 233000,China)
出处
《牡丹江医学院学报》
2022年第5期1-4,18,共5页
Journal of Mudanjiang Medical University
基金
安徽高校自然科学重点项目(KJ2019A0396)
蚌埠医学院自然科学重点项目(2020byzd081)。
