核糖核苷酸还原酶M2下调抑制人宫颈癌细胞的增殖、迁移和侵袭  

RRM2 Silencing Inhibits Proliferation,Migration and Invasion of Human Cervical Cancer Cells

作　　者：李雨颖 蔡锦威 林江涛 陈旭明 邱惠萍[1] LI Yu-Ying;CAI Jin-Wei;LIN Jiang-Tao;CHEN Xu-Ming;QIU Hui-Ping(Department of Medicine,Quzhou College of Technology,Quzhou 324000,Zhejiang,China;Quzhou Kecheng Hospital,Quzhou 324000,Zhejiang,China;Zhejiang University School of Medicine,Hangzhou 310058,China)

机构地区：[1]衢州职业技术学院医学院,浙江衢州324000 [2]衢州市柯城区人民医院肿瘤内科,浙江衢州324000 [3]浙江大学医学院,杭州310058

出　　处：《中国生物化学与分子生物学报》2020年第9期1121-1128,共8页Chinese Journal of Biochemistry and Molecular Biology

基　　金：衢州职业技术学院科研项目基金(No.QZYZ1710);衢州市科技计划项目(No.2017G14);浙江省教育厅科研项目(No.Y201941458);衢职院病毒致瘤研究科技创新团队项目资助。

摘　　要：核糖核苷酸还原酶M2(ribonucleotide reductase, RRM2)在多种癌症组织中高表达,这与肿瘤细胞的侵袭、转移及肿瘤血管形成等生物学行为密切相关。目前,关于以RRM2为靶点治疗宫颈癌的研究并不多,其对宫颈癌细胞的增殖、迁移和侵袭能力的影响及机制也尚未明确。本研究采用RNA干扰技术下调RRM2,以人宫颈癌细胞株CaSki和HeLa细胞为研究对象,实验分为Ctrl-siRNA组和RRM2-siRNA组。通过RT-PCR和Western印迹检测RRM2-siRNA的干扰作用,采用CCK-8和EDU实验评价细胞的增殖和DNA合成的情况,通过划痕实验和Transwell小室侵袭实验观察细胞的迁移和侵袭能力,并用Western印迹检测NF-κB和基质金属蛋白酶-9(matrix metalloproteinase-9, MMP-9)蛋白质表达水平。结果显示,在CaSki和HeLa细胞中,RRM2-siRNA组RRM2的mRNA和蛋白质表达较Ctrl-siRNA组均下降(P<0.05),表明RRM2-siRNA能有效抑制RRM2的表达。与Ctrl-siRNA组相比,RRM2-siRNA组在转染24、48、72 h后,细胞增殖率下降(P<0.05)。且DNA合成率与细胞划痕愈合程度,宫颈癌细胞侵袭数量也显著下降(P<0.05)。结果表明,RRM2下调后,抑制DNA合成与细胞增殖、迁移和侵袭过程。此外,RRM2-siRNA组NF-κB和MMP-9蛋白质水平较Ctrl-siRNA组降低。本研究提示,下调RRM2抑制宫颈癌细胞的增殖、迁移和侵袭,该作用可能通过NF-κB和MMP-9介导,这为宫颈癌中以RRM2作为靶点的治疗提供实验依据。It is reported that overexpression of ribonucleotide reductase M2(RRM2)in a wide variety of cancer tissues is closely related to the biological behavior including invasion and metastasis of tumor cells and tumor angiogenesis.To date,studies on RRM2 as a target to treat cervical cancer and its role in cervical cancer are still scanty.This study investigates the role of RRM2 on proliferation,migration and invasion of cervical cancer cell lines CaSki and HeLa.The cells were divided into transfected group(transfected with RRM2-siRNA)and negative control group(transfected with Ctrl-siRNA).The interference effect of RRM2-siRNA was detected by RT-PCR and Western blot.The influence of DNA synthesis and cell proliferation were tested by EDU incorporation assay and CCK-8 assay,respectively.The ability of migration and invasion were observed by cell wound healing assay and cell Transwell assay.At the same time,Western blot was used to measure the expression of NF-κB and MMP-9.The results showed that RRM2 was knocked down successfully in CaSki and HeLa cells(P<0.05).Compared with Ctrl-siRNA group,the proliferation of CaSki and HeLa cells were inhibited significantly at 24,48 and 72 hours in RRM2-siRNA group(P<0.05).Moreover,the DNA synthesis of CaSki and HeLa cells was decreased obviously in RRM2-siRNA group(P<0.05).In addition,the capacity of migration and invasion in CaSki and HeLa cells was restrained apparently after silencing RRM2(P<0.05).Meanwhile,the expression levels of NF-κB and MMP-9 proteins were dropped evidently in RRM2-siRNA group(P<0.05).In conclusion,the study suggests that knock down of RRM2 suppresses cervical carcinoma proliferation,invasion and metastasis,which may be mediated by NF-κB and MMP-9.These results could be meaningful to precision treatment of cervical carcinoma with RR inhibitors.

关 键 词：核糖核苷酸还原酶M2 宫颈癌 基质金属蛋白酶-9 NF-ΚB RNA干扰 

分 类 号：R730.23[医药卫生—肿瘤]