SF1a-PRLR和ΔS2 SF1a-PRLR对乳腺癌MCF-7细胞增殖和microRNA表达的影响

doi:10.3969/j.issn.1004-616x.2020.03.009

摘要/Abstract

摘要： 目的：探讨过表达SF1a-PRLR及其变体ΔS2 SF1a-PRLR基因对人乳腺癌MCF-7细胞增殖和microRNA表达的影响。方法：利用同源重组技术将SF1a-PRLR和ΔS2 SF1a-PRLR cDNA分别重组至慢病毒，再将携带不同基因的重组病毒，即空病毒、携带SF1a-PRLR cDNA和ΔS2 SF1a-PRLR cDNA的慢病毒分别转染MCF-7细胞，经嘌呤霉素多次筛选得到稳定转染细胞株MCF7-con（对照组）、MCF7-SF1a-PRLR（SF1a组）、MCF7-ΔS2 SF1a-PRLR（ΔS2 SF1a组），将3组稳定转染细胞株培养后进行细胞增殖实验，提取总RNA进行小分子RNA高通量测序。结果：增殖实验结果显示，培养48 h时，对照组、SF1a组和ΔS2 SF1a组的D（492）值分别为1.85±0.29、2.24±0.26、2.57±0.23，3组之间两两比较，差异均有统计学意义（P均 < 0.05）。测序结果显示，各组间microRNA表达均有显著差异，SF1a组与对照组比较，差异表达的microRNAs共有176个（32个表达上调，144个表达下调）；ΔS2 SF1a组与对照组相比，差异表达的microRNAs共206个（52个表达上调，154个表达下调）；ΔS2 SF1a组与SF1a组比较，差异表达的microRNA共5个（miR-4454、miR-215-5p、miR-797、miR-622表达上调，miR-210-5p表达下调），其中，miR-210-5p在对照组、SF1a组、ΔS2 SF1a组中的相对表达水平分别为66.18、31.67、13.07，两两比较均具有显著差异（P均 < 0.05）。结论：过表达SF1a-PRLR或ΔS2 SF1a-PRLR均能促进人乳腺癌MCF-7细胞的体外增殖且显著影响人乳腺癌MCF-7细胞的microRNA表达，两者对大部分microRNA表达的影响作用相似，仅对miR-4454等5个microRNAs的表达具有显著影响。

关键词: 乳腺癌, 催乳素短受体1a, 丢失S2的催乳素短受体1a, 微小核糖核酸, 差异表达

Abstract: OBJECTIVE: To investigate the effects of SF1a-PRLR and its variant ΔS2 SF1a-PRLR over-expression on proliferation and microRNA expression in human breast cancer cells (MCF-7). METHODS: SF1a-PRLR and ΔS2 SF1a-PRLR cDNAs were recombined separately into lentivirus by using homologous recombination strategy,and the viruses carrying different genes (including SF1a-PRLR,or ΔS2 SF1a-PRLR cDNA) or not carrying any target gene (control) were transfected into MCF-7 cells,respectively. The stably transfected cells including MCF7-con (control group),MCF7-SF1a-PRLR (SF1a),and MCF7-ΔS2 SF1a-PRLR (ΔS2 SF1a) were obtained through multiple screening with puromycin. Total RNAs were extracted using Trizol reagent,and high-throughput sequencing of small RNAs were performed. In addition, separate sets of the cells were tested using the MTS cell proliferation assay. RESULTS: Cell proliferation assay results show that at 48 h,the D(492) values of the control,SF1a and ΔS2 SF1a groups were 1.85±0.29,2.24±0.26 and 2.57±0.23,with significant differences among them (all P < 0.05). Furthermore, sequencing data show that there were significant differences on microRNA expression among the groups. SF1a induced 176 microRNAs expression changes(32 were up-regulated and 144 were down-regulated) and ΔS2 SF1a induced 206 microRNAs expression changes (52 were up-regulated and 154 were down-regulated). For comparison,4 microRNAs (miR-4454,miR-215-5p,miR-622 and miR-797) were up-regulated and 1 (miR-210-5p) was down-regulated from ΔS2 SF1a-PRLR over-expression. The relative expressions of miR-210-5p in the control,SF1a and ΔS2 SF1a groups were 66.18、31.67 and 13.07,with significant differences among the three groups (all P < 0.05). CONCLUSION: Both SF1a-PRLR and ΔS2 SF1a-PRLR over-expression promoted proliferation of breast cancer cells (MCF-7),and significantly affected the expression of microRNAs in a similar way for most of the detected genes. Several microRNAs such as miR-4454,etc. are affected in a different mode by SF1a PRLR and ΔS2 SF1a-PRLR.

Key words: breast cancer, SF1a-PRLR, ΔS2 SF1a-PRLR, microRNA, differential expression

中图分类号:

张惠娟, 黄大元, 张洁, 唐海欧, 谢安心, 谭敦勇. SF1a-PRLR和ΔS2 SF1a-PRLR对乳腺癌MCF-7细胞增殖和microRNA表达的影响[J]. 癌变·畸变·突变, 2020, 32(3): 203-208.

ZHANG Huijuan, HUANG Dayuan, ZHANG Jie, TANG Haiou, XIE Anxin, TAN Dunyong. Effects of SF1a-PRLR and ΔS2 SF1a-PRLR on proliferation and microRNA expressionin MCF-7 breast cancer cell[J]. Carcinogenesis, Teratogenesis & Mutagenesis, 2020, 32(3): 203-208.

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