下调受体酪氨酸激酶样孤儿受体1表达对黑色素瘤B16F10生物学特性的影响

Effects of down-regulating expression of receptor tyrosine kinase like orphan receptor 1 on biological characteristics for melanoma B16F10 cell

目的探讨下调黑色素瘤(melanoma)B16F10细胞中受体酪氨酸激酶样孤儿受体1(receptor tyrosine kinase like orphan receptor 1,ROR1)分子表达对其生物学特性的影响,为治疗黑色素瘤筛选候选靶标奠定基础。方法①以慢病毒载体介导shRNA下调B16F10细胞中ROR1的表达,并检测其下调效果;②CCK8法检测B16F10,Scrambled和Lv-shROR1-B16F10等3组细胞的增殖能力,再通过划痕实验和Transwell实验检测3组细胞的迁移能力,最后用流式细胞仪检测3组细胞的细胞周期;③将3组细胞接种到小鼠体内,观察小鼠的致瘤能力,并检测其致瘤效果。结果①经慢病毒感染后得到Lv-shROR1-B16F10和Scrambled两株细胞;②下调ROR1能够抑制细胞的增殖能力和细胞的迁移能力(P<0.05),使细胞停滞在G0~G1(P<0.05);③下调ROR1能抑制B16F10细胞的致瘤能力,瘤组织中ROR1的表达水平也降低(P<0.05)。结论ROR1可参与黑色素瘤细胞的增殖、转移以及细胞的致瘤性,是一个有效治疗黑色素瘤的候选靶标。

Objective To investigate the effects of down-regulating the expression of receptor tyrosine kinase like orphan receptor 1(ROR1)on melanoma cell’s biological characteristics,and to provide with a base by screening candidate target in treatment of melanoma.Methods①Lentiviral vectors were used to mediate shRNA to down-regulate the expression of ROR1 in B16 F10 cells,and in detection of the effect from down-regulation.②The CCK8 method was used to detect the proliferation ability of B16 F10,Scramble and Lv-shROR1-B16 F10 cells,then the migration ability of the three group cells was tested by the scratch test and the Transwell experiment,and finally the cell cycle of the three groups of cells was detected by flow cytometry.③After inoculating three groups of cells into mice,respectively,the cellular tumorigenic ability was observed and evaluated.Results①After lentivirus infection,namely Lv-shROR1-B16 F10 and Scramble,two infected group cells were obtained.②Down-regulating ROR1 expression inhibited the cellular proliferation ability(P<0.05)and cellular migration ability(P<0.05),and cell cycle process was arrested in the G0—G1 phases(P<0.05).③Down-regulation of ROR1 inhibited the tumorigenic ability of B16 F10 cells,and the expression level of ROR1 in the tumor tissue was also decreased(P<0.05).Conclusion ROR1 is involved in the melanoma cell proliferation,metastasis,and tumorigenicity,suggesting the ROR1 is one of candidate targets for treatment of melanoma.