miR-215抑制骨肉瘤细胞生长及其分子机制
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摘要
目的:研究miR-215对骨肉瘤细胞143B生长的影响及其分子机制。方法:通过转染miR-215 agomir稳定升高143B细胞内miR-215表达水平,CCK-8实验、平板克隆形成实验以及流式细胞术检测143B细胞的增殖状况以及周期分布。利用生物信息学分析miR-215潜在的靶基因,并通过双荧光素酶报告基因实验、qPCR实验、Western blot实验验证miR-215靶基因。结果:在143B细胞内将miR-215表达水平上调数倍。CCK-8实验、平板克隆形成实验表明miR-215能够抑制143B细胞生长,流式细胞术实验表明miR-215能够延缓143B细胞周期进程。生物信息学、qPCR实验和Western blot实验表明转化生长因子β受体1(transforming growth factorβreceptor 1,TGFβR1)是miR-215潜在的靶基因。结论:miR-215能够抑制骨肉瘤细胞生长,并且该抑制作用可能通过miR-215靶向抑制TGFβR1实现,为骨肉瘤早期诊断和靶向治疗提供了理论依据。
Objective:To study the effects of miR-215 on the growth of 143 B and its molecular mechanism.Methods:miR-215 agomir was transfected to upregulate miR-215 expression level,and agomir NC group and blank group were regarded as control group.qPCR was performed to examine the expression level of miR-215 in 143 B cells.CCK-8,colony formation assay and FCM were used to study the effects of miR-215 on the growth of 143 B cells.TGFβR1 was predicted to be a potential target gene with bioinformatics and confirmed by means of dual luciferase reporter gene assay,qPCR and Western blot.Results:miR-215 agomir was efficiently expressed in 143 B cells,and CCK-8 assay,colony formation assay and FCM showed that upregulation of miR-215 can inhibit the proliferation of 143 B cells.Dual luciferase reporter gene assay,qPCR and Western blot confirmed that transforming growth factor β receptor 1(TGFβR1) was a target gene of miR-215.These results tentatively illuminated the molecular mechanism by which miR-215 inhibited growth of 143 B cells.Conclusion:miR-215 could inhibit growth of 143 B cells,which may be explained by TGFβR1 being a target gene of miR-215.
Objective:To study the effects of miR-215 on the growth of 143 B and its molecular mechanism.Methods:miR-215 agomir was transfected to upregulate miR-215 expression level,and agomir NC group and blank group were regarded as control group.qPCR was performed to examine the expression level of miR-215 in 143 B cells.CCK-8,colony formation assay and FCM were used to study the effects of miR-215 on the growth of 143 B cells.TGFβR1 was predicted to be a potential target gene with bioinformatics and confirmed by means of dual luciferase reporter gene assay,qPCR and Western blot.Results:miR-215 agomir was efficiently expressed in 143 B cells,and CCK-8 assay,colony formation assay and FCM showed that upregulation of miR-215 can inhibit the proliferation of 143 B cells.Dual luciferase reporter gene assay,qPCR and Western blot confirmed that transforming growth factor β receptor 1(TGFβR1) was a target gene of miR-215.These results tentatively illuminated the molecular mechanism by which miR-215 inhibited growth of 143 B cells.Conclusion:miR-215 could inhibit growth of 143 B cells,which may be explained by TGFβR1 being a target gene of miR-215.
引文
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[3] Zhou SW,Su BB,Zhou Y,et al.Aberrant miR-215 expression is associated with clinical outcome in breast cancer patients[J].Med Oncol,2014,31(11):259.
[4] Cai X,Peng D,Wei H,et al.miR-215 suppresses proliferation and migration of non-small cell lung cancer cells[J].Oncol Lett,2017,13(4):2349-2353.
[5] Yongqing Tong,Bei Liu,Hongyun Zheng,et al.MiR-215,an activator of the CTNNBIP1/β-catenin pathway,is a marker of poor prognosis in human glioma[J].Oncotarget,2015,6(28):25024-25033.
[6] Li S,Gao J,Gu J,et al.MicroRNA-215 inhibits relapse of colorectal cancer patients following radical surgery[J].Med Oncol,2013,30(2):549.
[7] Mihriban Karaayvaz,Timothy Pal,Bo Song,et al.Prognostic significance of miR-215 in colon cancer[J].Clin Colorectal Cancer,2011,10(4):340-347.
[8] Liang H,Li Y,Luo RY,et al.MicroRNA-215 is a potential prognostic marker for cervical cancer[J].J Huazhong Univ Sci Technolog Med Sci,2014,34(2):207-212.
[9] Antonio Agostini,Marta Brunetti,Ben Davidson,et al.The microRNA miR-192/215 family is upregulated in mucinous ovarian carcinomas[J].Sci Rep,2018,8:11069.
[10] Boni V,Bitarte N,Cristobal I,et al.miR-192/miR-215 influence5-fluorouracil resistance through cell cycle-mediated mechanisms complementary to its post-transcriptional thymidilate synthase regulation[J].Mol Cancer Ter,2010,9(8):2265-2275.
[11] Nakaoka T,Saito Y,Shimamoto Y,et al.Cluster microRNAs miR-194 and miR-215 suppress the tumorigenicity of intestinal tumor organoids[J].Cancer Sci,2017,108(4):678-684.
[12] Wang L,Wang YM,Xu S,et al.MicroRNA-215 is upregulated bytreatment with adriamycin and leads to the chemoresistance of hepatocellular carcinoma cells and tissues[J].Mol Med Rep,2015,12(4):5274-5280.
[13] Ge G,Zhang W,Niu L,et al.MiR-215 functions as a tumor suppressor in epithelial ovarian cancer through regulation of the X-chromosome-linked inhibitor of apoptosis[J].Oncol Rep,2016,35(3):1816-1822.
[14] Ye M,Zhang J,Zhang J,et al.Curcumin promotes apoptosis by activating the p53-miR-192-5p/215-XIAP pathway in non-small cell lung cancer[J].Cancer Let,2015,357(1):196-205.
[15] Yao Y,Shen H,Zhou Y,et al.MicroRNA-215 suppresses the proliferation,migration and invasion of non-small cell lung carcinoma cells through the downregulation of matrix metalloproteinase-16 expression[J].Exp Ter Med,2018,15(4):3239-3246.
[16] Martin JW,Squire JA,Zielenska M,et al.The genetics of osteosarcoma[J].Sarcoma,2012,2012:627254.
[17] Siegel RL,Miller KD,Jemal A,et al.Cancer statistics,2017[J].CA Cancer J Clin,2017,67(1):7-30.
[18] Wang YX,Zhang TJ,Yang DQ,et al.Reduced miR-215 expression predicts poor prognosis in patients with acute myeloid leukemia[J].Jpn J Clin Oncol,2016,46(4):350-356.
[19] Song B,Wang Y,Titmus MA,et al.Molecular mechanism of chemoresistance by miR-215 in osteosarcoma and colon cancer cells[J].Mol Cancer,2010,9:96.
[20] Chen Z,Liu K,Li L,et al.miR-215 promotes cell migration and invasion of gastric cancer by targeting retinoblastoma tumor suppressor gene 1[J].Pathol Res Pract,2017,213(8):889-894.
[21] Li N,Zhang QY,Zou JL,et al.miR-215 promotes malignant progression of gastric cancer by targeting RUNX1[J].Oncotarget,2016,7(4):4817-4828.
[22] Zang Y,Wang T,Pan J,et al.miR-215 promotes cell migration and invasion of gastric cancer cell lines by targeting FOXO1[J].Neoplasma,2017,64(4):579-587.
[23] Li QW,Zhou T,Wang F,et al.MicroRNA-215 functions as a tumor suppressor and directly targets ZEB2 in human pancreatic cancer[J].Genet Mol Res,2015,14(4):16133-16145.
[24] Li Y,Liu G,Li X,et al.Long non-coding RNA SBF2-AS1 promotes hepatocellular carcinoma progression through regulation of miR-140-5p-TGFBR1 pathway[J].Biochem Biophys Res Commun,2018,503(4):2826-2832.
[25] Zhou B,Guo W,Sun C,et al.Linc00462 promotes pancreatic cancer invasiveness through the miR-665/TGFBR1-TGFBR2/SMAD2/3 pathway[J].Cell Death Dis,2018,9(6):706.