靶向沉默IL-6对乳腺癌细胞侵袭和迁移能力的影响
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摘要
目的:观察白细胞介素6(IL-6)与人乳腺癌MCF-7细胞生长、侵袭和迁移的关系及其作用机制。方法:合成IL-6小干扰RNA(si RNA),并转染至MCF-7细胞中。实验分为空白对照组、阴性对照组和IL-6 si RNA组。通过MTT实验观察沉默IL-6基因表达对细胞活力的影响,Transwell小室法检测细胞侵袭和迁移能力的变化,采用q PCR和Western blot法检测转染后IL-6表达水平的变化,同时采用Western blot法观察上皮-间充质转化(EMT)相关蛋白、信号转导及转录激活因子3(STAT3)、磷酸化STAT3(p-STAT3)、基质金属蛋白酶2(MMP-2)和基质金属蛋白酶9(MMP-9)的蛋白水平。结果:与对照组相比,转染IL-6 si RNA的MCF-7细胞中IL-6的表达量显著下降(P <0. 05)。沉默IL-6表达显著抑制MCF-7细胞的活力(P <0. 05),并降低其侵袭和迁移能力(P <0. 05),显著抑制N-cadherin和vimentin的表达(P <0. 05),细胞中STAT3无明显变化,p-STAT3、MMP-2和MMP-9的蛋白水平显著降低(P <0. 05)。结论:沉默IL-6表达可能通过抑制EMT,并降低STAT3的磷酸化水平进而抑制MMP-2和MMP-9的分泌,从而减弱乳腺癌MCF-7细胞的活力,并降低其侵袭和迁移能力。
AIM: To investigate the relationship between interleukin-6( IL-6) and viability,invasion and migration of human breast cancer MCF-7 cells and its mechanism. METHODS: IL-6 small interfering RNA( si RNA) was synthesized and transfected into the MCF-7 cells. The cells were divided into blank control group,negative control group and IL-6 si RNA group. The viability of the MCF-7 cells was measured by MTT assay,and the invasion and migration abilities were analyzed by Transwell assay. The expression of IL-6 at m RNA and protein levels was examined by q PCR and Western blot. The protein levels of epithelial-mesenchymal transition( EMT)-related proteins,signal transducer and activator of transcription 3( STAT3),p-STAT3,matrix metalloproteinase-2( MMP-2) and MMP-9 were determined by Western blot. RESULTS: Compared with control groups,IL-6 si RNA decreased the expression of IL-6 significantly and reduced the viability of MCF-7 cells( P < 0. 05). Silencing of IL-6 expression significantly decreased the invasion and migration abilities of MCF-7 cells( P < 0. 05),inhibited the expression of EMT-related proteins( P < 0. 05),and reduced the protein levels of p-STAT3,MMP-2 and MMP-9( P < 0. 05). CONCLUSION: Silencing of IL-6 expression inhibits the viability of MCF-7 cells and reduces the invasion and migration abilities by inhibiting the expression of EMT-related proteins,decreasing the phosphorylation of STAT3 and thus inhibiting the secretion of MMP-2 and MMP-9.
AIM: To investigate the relationship between interleukin-6( IL-6) and viability,invasion and migration of human breast cancer MCF-7 cells and its mechanism. METHODS: IL-6 small interfering RNA( si RNA) was synthesized and transfected into the MCF-7 cells. The cells were divided into blank control group,negative control group and IL-6 si RNA group. The viability of the MCF-7 cells was measured by MTT assay,and the invasion and migration abilities were analyzed by Transwell assay. The expression of IL-6 at m RNA and protein levels was examined by q PCR and Western blot. The protein levels of epithelial-mesenchymal transition( EMT)-related proteins,signal transducer and activator of transcription 3( STAT3),p-STAT3,matrix metalloproteinase-2( MMP-2) and MMP-9 were determined by Western blot. RESULTS: Compared with control groups,IL-6 si RNA decreased the expression of IL-6 significantly and reduced the viability of MCF-7 cells( P < 0. 05). Silencing of IL-6 expression significantly decreased the invasion and migration abilities of MCF-7 cells( P < 0. 05),inhibited the expression of EMT-related proteins( P < 0. 05),and reduced the protein levels of p-STAT3,MMP-2 and MMP-9( P < 0. 05). CONCLUSION: Silencing of IL-6 expression inhibits the viability of MCF-7 cells and reduces the invasion and migration abilities by inhibiting the expression of EMT-related proteins,decreasing the phosphorylation of STAT3 and thus inhibiting the secretion of MMP-2 and MMP-9.
引文
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[2]Inoue M,Nakagomi H,Nakada H,et al.Specific sites of metastases in invasive lobular carcinoma:a retrospective cohort study of metastatic breast cancer[J].Breast Cancer,2017,24(5):667-672.
[3]De Santis CE,Ma J,Goding Sauer A,et al.Breast cancer statistics,2017,racial disparity in mortality by state[J].CA Cancer J Clin,2017,67(6):439-448.
[4]Ghasemi H.Roles of IL-6 in ocular inflammation:a review[J].Ocul Immunol Inflamm,2018,26(1):37-50.
[5]Wu X,Tao P,Zhou Q,et al.IL-6 secreted by cancer-associated fibroblasts promotes epithelial-mesenchymal transition and metastasis of gastric cancer via JAK2/STAT3signaling pathway[J].Oncotarget,2017,8(13):20741-20750.
[6]Qin X,Yan M,Wang X,et al.Cancer-associated fibroblast-derived IL-6 promotes head and neck cancer progression via the osteopontin-NF-kappa B signaling pathway[J].Theranostics,2018,8(4):921-940.
[7]Sahana KR,Akila P,Prashant V,et al.Quantitation of vascular endothelial growth factor and interleukin-6 in different stages of breast cancer[J].Rep Biochem Mol Biol,2017,6(1):33-39.
[8]Todoric J,Antonucci L,Karin M.Targeting inflammation in cancer prevention and therapy[J].Cancer Prev Res(Phila),2016,9(12):895-905.
[9]Park MH,Hong JT.Roles of NF-κB in cancer and inflammatory diseases and their therapeutic approaches[J].Cells,2016,5(2):E15.
[10]Sun Z,Meng Y,Liu G,et al.Effect of interleukin-1βand tumor necrosis factorαgene silencing on mouse gastric cancer cell proliferation and migration[J].Oncol Lett,2016,11(4):2559-2565.
[11]Iyengar NM,Zhou XK,Gucalp A,et al.Systemic correlates of white adipose tissue inflammation in early-stage breast cancer[J].Clin Cancer Res,2016,22(9):2283-2289.
[12]Wu YS,Chung I,Wong WF,et al.Paracrine IL-6 signaling mediates the effects of pancreatic stellate cells on epithelial-mesenchymal transition via Stat3/Nrf2 pathway in pancreatic cancer cells[J].Biochim Biophys Acta,2017,1861(2):296-306.
[13]Culig Z,Pencik J,Merkel O,et al.Breaking a paradigm:IL-6/STAT3 signaling suppresses metastatic prostate cancer upon ARF expression[J].Mol Cell Oncol,2016,3(2):e1090048.
[14]Li E,Zhang J.Essential role of SH3GL1 in interleukin-6(IL-6)-and vascular endothelial growth factor(VEGF)-triggered p130cas-mediated proliferation and migration of osteosarcoma cells[J].Hum Cell,2017,30(4):300-310.
[15]Lee SO,Yang X,Duan S,et al.IL-6 promotes growth and epithelial-mesenchymal transition of CD133+cells of non-small cell lung cancer[J].Oncotarget,2016,7(6):6626-6638.
[16]Bharti R,Dey G,Ojha PK,et al.Diacerein-mediated inhibition of IL-6/IL-6R signaling induces apoptotic effects on breast cancer[J].Oncogene,2016,35(30):3965-3975.
[17]曹蕾,李磊,胡明珠,等.STC2促进人肝癌细胞Hep G2增殖和EMT相关的迁移[J].中国病理生理杂志,2017,33(6):1000-1005.
[18]Li J,Riedt T,Goossens S,et al.The EMT transcription factor Zeb2 controls adult murine hematopoietic differentiation by regulating cytokine signaling[J].Blood,2017,129(4):460-472.
[19]潘祉谕,达静静,董蓉,等.Snail1/IGF-1信号通路介导高糖诱导的肾小管上皮细胞EMT[J].中国病理生理杂志,2017,33(9):1662-1668.
[20]李逦,王纯,卢宏达.马钱子碱通过抑制IL-6/STAT3信号通路诱导结肠癌SW480细胞凋亡[J].中国病理生理杂志,2016,32(6):998-1003.
[21]Baek SH,Ko JH,Lee H,et al.Resveratrol inhibits STAT3 signaling pathway through the induction of SOCS-1:Role in apoptosis induction and radiosensitization in head and neck tumor cells[J].Phytomedicine,2016,23(5):566-577.
[22]Wang Y,Liang T,Wang Y,et al.Long non-coding RNAAK093407 promotes proliferation and inhibits apoptosis of human osteosarcoma cells via STAT3 activation[J].Am JCancer Res,2017,7(4):892-902.
[23]Kim C,Baek SH,Um JY,et al.Resveratrol attenuates constitutive STAT3 and STAT5 activation through induction of PTPεand SHP-2 tyrosine phosphatases and potentiates sorafenib-induced apoptosis in renal cell carcinoma[J].BMC Nephrol,2016,17:19.
[24]Zergoun AA,Zebboudj A,Sellam SL,et al.IL-6/NOS2inflammatory signals regulate MMP-9 and MMP-2 activity and disease outcome in nasopharyngeal carcinoma patients[J].Tumour Biol,2016,37(3):3505-3514.
[25]Zou M,Zhang X,Xu C.IL6-induced metastasis modulators p-STAT3,MMP-2 and MMP-9 are targets of 3,3’-diindolylmethane in ovarian cancer cells[J].Cell Oncol(Dordr),2016,39(1):47-57.