中华皮肤科杂志 ›› 2015, Vol. 48 ›› Issue (12): 856-860.

• 论著 • 上一篇    下一篇

RNA干扰趋化因子受体CXCR7基因对黑素瘤M14细胞侵袭和迁移的影响

刘排1,田蔚蔚1,李小静2,李志锋2,燕霞3,孙建方4   

  1. 1. 江西省皮肤病专科医院
    2. 河北邯郸 河北工程大学附属医院
    3. 河北工程大学附属医院
    4. 南京 中国医学科学院北京协和医学院皮肤病研究所
  • 收稿日期:2015-02-05 修回日期:2015-09-21 出版日期:2015-12-15 发布日期:2015-12-01
  • 通讯作者: 孙建方 E-mail:fangmin5758@aliyun.com

Effects of RNA interference targeting the chemokine receptor 7 gene on the invasion and migration of the human melanoma cell line M14

  • Received:2015-02-05 Revised:2015-09-21 Online:2015-12-15 Published:2015-12-01

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摘要：

目的 探讨靶向沉默趋化因子受体CXCR7基因对黑素瘤细胞株M14侵袭和迁移能力的影响。方法 采用Western印迹实验检测黑素瘤M14和A375细胞CXCR7蛋白的表达，选取高表达CXCR7蛋白的M14细胞作为研究对象。将M14细胞分为实验组、阴性对照组和空白对照组：实验组转染CXCR7-siRNA，阴性对照组转染阴性对照siRNA，空白对照组不做任何处理。采用实时定量PCR和蛋白质印迹法分别检测CXCR7 mRNA和蛋白在M14细胞中的表达水平， Transwell小室法及细胞划痕法检测干扰前后细胞侵袭及迁移能力的变化。 结果 实验组M14细胞CXCR7 mRNA及蛋白相对表达量分别为0.412 ± 0.023、0.144 ± 0.005，明显低于阴性对照组（1.211 ± 0.117、1）以及空白对照组（1.000 ± 0.102、1.016 ± 0.004），差异均有统计学意义（F值分别为30.068、11 485.5，P值分别为0.001、0.000）。实验组M14细胞穿过聚碳酸酯膜的细胞数为（20.617 ± 1.503）个，明显少于阴性转染对照组［（42.000 ± 6.018）个］和空白对照组［（43.627 ± 2.152）个］，3组间比较差异有统计学意义（F = 32.416，P = 0.001）。划痕实验中，实验组每高倍视野（× 200）下细胞迁移数为15.00 ± 1.10，明显少于阴性对照组（44.90 ± 2.20）和空白对照组（45.30 ± 2.30），3组间比较差异有统计学意义（F = 2 411.945，P = 0.000）。 结论 靶向沉默CXCR7能显著抑制黑素瘤M14细胞的侵袭和迁移，有望为皮肤黑素瘤提供潜在的治疗靶点。

Abstract:

Liu Pai *, Tian Weiwei, Li Xiaojing, Li Zhifeng, Yan Xia, Sun Jianfang. *Department of Dermatology, Jiangxi Province Dermatosis Special Hospital, Nanchang 330001, China Corresponding authors: Sun Jianfang, Email: fangmin5758@aliyun.com; Li Xiaojing, Email: zlmdsh@126.com 【Abstract】 Objective To explore the effects of targeted silencing of the chemokine receptor 7 （CXCR7） gene on the invasion and migration of the melanoma cell line M14. Methods Western-blot analysis was performed to determine the protein expression of CXCR7 in melanoma cell lines M14 and A375, and CXCR7-overexpressing M14 cells were used in this study. Cultured M14 cells were divided into three groups: experimental group transfected with a small interfering RNA （siRNA） targeting CXCR7 （CXCR7-siRNA）, negative control group transfected with a negative control siRNA, blank control group receiving no treatment. Real-time quantitative PCR and Western-blot analysis were conducted to determine the mRNA and protein expressions of CXCR7 respectively in M14 cells, Transwell chambers were used to evaluate the invasive activity of M14 cells, and wound healing assay to estimate the migratory activity of M14 cells. Results The experimental group showed significantly decreased mRNA and protein expressions of CXCR7 compared with the negative control group and blank control group （CXCR7 mRNA: 0.412 ± 0.023 vs. 1.211 ± 0.117 and 1.000 ± 0.102, F = 30.068, P = 0.001; CXCR7 protein: 0.144 ± 0.005 vs. 1 and 1.016 ± 0.004, F =11 485.5, P = 0.000）. The number of M14 cells crossing the polycarbonate membrane per high-power field （× 200） was significantly smaller in the experimental group than in the negative control group and blank control group （20.617 ± 1.503 vs. 42.000 ± 6.018 and 43.627 ± 2.152, F = 32.416, P = 0.001）. Similarly, the number of migrating M14 cells in wound healing assay was significantly decreased in the experimental group compared with the negative control group and blank control group （15.00 ± 1.10 vs. 44.90 ± 2.20 and 45.30 ± 2.30, F = 2 411.945, P = 0.000）. Conclusion Targeted silencing of the CXCR7 gene can significantly inhibit the invasion and migration of M14 cells in vitro, which may provide a potential target for the treatment of cutaneous melanoma.