丹参酮ⅡA对黑素瘤细胞侵袭迁移能力的抑制作用及机制

Inhibitory effects of tanshinone ⅡA on migration and invasion of melanoma cells and their mechanisms

目的 探讨丹参酮ⅡA对黑素瘤A375细胞侵袭转移的作用及趋化因子受体7（CXCR7）基因与蛋白表达的影响。方法 体外培养黑素瘤A375细胞，经1、2、4 mg/L丹参酮ⅡA处理48 h后，细胞划痕实验和Transwell细胞体外侵袭实验分别测定细胞迁移和侵袭能力，实时荧光定量PCR和Western印迹分别检测A375细胞CXCR7 mRNA和蛋白的表达水平。结果 1、2、4 mg/L丹参酮ⅡA处理A375细胞48 h后，Transwell细胞体外侵袭实验显示，每高倍（× 200）视野下迁移穿过聚碳酸酯膜的细胞数分别为71.00 ± 4.00、51.00 ± 2.00、37.00 ± 3.61，而细胞划痕实验显示，细胞迁移数分别为301 ± 3.00、253.00 ± 3.61、126.00 ± 7.00，均显著低于对照组（105.33 ± 6.51、332.00 ± 6.24，均P 〈 0.05），且丹参酮ⅡA对A375细胞侵袭、迁移能力的抑制作用随丹参酮ⅡA浓度的升高而增强（均P 〈 0.05）。实时荧光定量PCR及Western印迹显示，1、2、4 mg/L丹参酮ⅡA组CXCR7 mRNA的相对表达量分别为0.63 ± 0.04、0.44 ± 0.02、0.31 ± 0.01，蛋白表达水平分别为0.573 ± 0.015、0.416 ± 0.011、0.260 ± 0.055，均显著低于对照组（1.00 ± 0.02、0.9000 ± 0.010，均P 〈 0.05），且丹参酮ⅡA对A375细胞CXCR7 mRNA和蛋白表达的抑制能力随丹参酮ⅡA浓度的升高而增强（均P 〈 0.05）。结论 丹参酮ⅡA可通过下调CXCR7表达抑制黑素瘤A375细胞的侵袭迁移能力。

Objective To evaluate effects of tanshinone ⅡA on the invasion and migration of melanoma A375 cells, as well as on the mRNA and protein of CXC chemokine receptor type 7 （CXCR7）. Methods In vitro cultured A375 cells were divided into 4 groups to be treated with tanshinone ⅡA at different concentrations of 1, 2 and 4 mg/L, and 0.1% dimethyl sulfoxide （DMSO） （control group） for 48 hours, respectively. Wound scratch assay and Transwell invasion assay were conducted to estimate the migratory and invasive abilities of A375 cells, respectively, and real-time fluorescence-based quantitative PCR （qRT-PCR） and Western blot analysis to determine the mRNA and protein of CXCR7 in A375 cells, respectively. Results After 48-hour treatment, the 1-, 2- and 4-mg/L tanshinone ⅡA groups showed significantly decreased number of A375 cells crossing the polycarbonate membrane per high-power field （× 200） （71.00 ± 4.00, 51.00 ± 2.00 and 37.00 ± 3.61, respectively） in Transwell invasion assay, as well as decreased number of A375 cells migrating to the scratch zone （301 ± 3.00, 253.00 ± 3.61 and 126.00 ± 7.00, respectively） in wound scratch assay, compared with the control group （105.33 ± 6.51, 332.00 ± 6.24, respectively, all P 〈 0.05）. Additionally, qRT-PCR and Western blot analysis revealed that the mRNA and protein of CXCR7 was significantly lower in the 1-, 2- and 4-mg/L tanshinone ⅡA groups than in the control groups （CXCR7 mRNA： 0.63 ± 0.04, 0.44 ± 0.02 and 0.31 ± 0.01 vs. 1.00 ± 0.02; CXCR7 protein： 0.573 ± 0.015, 0.416 ± 0.011 and 0.260 ± 0.055 vs. 0.9000 ± 0.010; all P 〈 0.05）. Moreover, inhibitory effects of tanshinone ⅡA on the migration and invasion of A375 cells, as well as on the mRNA and protein of CXCR7, were significantly enhanced with the increase of tanshinone ⅡA concentrations （all P 〈 0.05）. Conclusion Tanshinone ⅡA can inhibit the migratory and invasive abilities of melanoma A375 cells by down-regulating CXCR7 .