RNA干扰CysLT2R对LTC4介导的人乳腺癌细胞生物学行为的影响
Effects of cysteinyl leukotriene receptor Ⅱ on the biological behavior of MCF-7 cells
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摘要:目的 观察MCF-7细胞低表达半胱氨酰白三烯Ⅱ型受体(CysLT2R)对半胱氨酰白三烯(LTC4)介导的细胞生长、侵袭能力、细胞周期和凋亡等生物学行为的影响。方法 针对CysLT2R的已知cDNA序列, 设计并体外转录合成4条特异性shRNA, 用脂质体介导转染入MCF-7细胞48 h, 同时设阴性对照组(1条非特异性序列转染)、正常MCF-7对照组(未转染)。Western blot检测干扰后CysLT2R蛋白表达水平下调的变化。应用噻唑蓝比色(MTT)法检测50 nM LTC4作用下细胞株生长能力; 应用流式细胞仪检测50 nM LTC4作用下细胞周期和凋亡变化; 应用趋化小室检测50 nM LTC4作用下MCF-7细胞侵袭运动活性。结果 与正常MCF-7细胞和阴性对照组比较, MCF-7细胞CysLT2R低表达组的细胞生长增殖无明显变化(P > 0.05), 干扰后, MCF-7细胞表现出侵袭活性明显增强(P < 0.05), 细胞周期和凋亡无明显变化。结论 CysLT2R在人乳腺癌MCF-7细胞的侵袭过程中起着一定的作用, CysLT2R有可能是一种新的乳腺肿瘤抑制基因。Abstract:Objective The objective of this research is to study the effects of cysteinyl leukotriene receptor Ⅱ(CysLT2R) on the biological behavior of the MCF-7 human breast carcinoma cell line facilitated by proinflammatory leukotriene C4(LTC4).Methods Four small interfering RNAs(shRNA) targeting CysLT2R were transfected into the MCF-7 human breast carcinoma cell line.Western blot assay was used to determine the down-regulation of the protein.The cell cycle changes and apoptosis in the presence of LTC4 were detected using flow cytometry.The proliferation of cells under the impact of 50 nM LTC4 was analyzed using methyl thiazolyl tetrazolium assay.Cell invasion assays were performed on breast cancer cells in the presence of LTC4.Results No significant change in the cell cycle and apoptosis among the MCF-7 cells(transfected with CysLT2R-shRNA) were observed.The low-expression of CysLT2R had no significant effect on the proliferation of MCF-7 cells.The LTC4-induced cell invasion was evidently increased by the silencing of CysLT2R(P < 0.05).Conclusion Our results show the potential role of CysLT2R in the process of breast cancer cell invasion.CysLT2R may be a new breast carcino ma-suppressor gene.
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