c-Myc调控PIK3R3抑制黑色素瘤B16-F10细胞的迁移和侵袭

发布时间：2018-03-27 08:19

本文选题：miR-a　切入点：c-Myc　出处：《中国免疫学杂志》2017年01期

【摘要】：目的:探讨c-Myc调控PIK3R3对黑色素瘤B16-F10细胞的迁移和侵袭的影响。方法:运用免疫组化检测PIK3R3在正常皮肤组织和黑色素瘤组织的表达;检测慢病毒(LV3-c-Myc和LV3-PIK3R3)对c-Myc和PIK3R3基因的沉默效率;使用Ed U增殖实验检测沉默c-Myc和PIK3R3后黑色素瘤细胞株B16-F10的增殖情况;Transwell侵袭实验检测c-Myc和PIK3R3的表达对黑色素瘤细胞B16-F10的侵袭能力的影响;划痕实验检测c-Myc和PIK3R3的表达对黑色素瘤细胞B16-F10迁移能力的影响;q PCR检测沉默c-Myc和PIK3R3后miR-199a的表达;q PCR检测转染miR-199a mimics和miR-199a inhibitor后c-Myc和PIK3R3的表达;双荧光素酶报告基因系统检测miR-199a对c-Myc和PIK3R3转录活性的影响。结果:和正常皮肤组织比较,PIK3R3在黑色素瘤组织中表达明显增加;沉默c-Myc和PIK3R3基因后,黑色素瘤细胞株B16-F10的增殖、侵袭和转移能力明显降低;沉默c-Myc和PIK3R3基因后,miR-199a表达上调;转染miR-199a mimics后,c-Myc和PIK3R3表达降低;转染miR-199a inhibitor后,c-Myc和PIK3R3表达上调;双荧光素酶报告基因系统检测结果显示,miR-199a可以直接调控c-Myc和PIK3R3的转录活性。结论:c-Myc可以通过miR-199a调控PIK3R3的表达,从而促进黑色素瘤细胞的增殖、侵袭和迁移能力。
[Abstract]:Objective: to investigate the effect of c-Myc on the migration and invasion of B16-F10 cells in melanoma. Methods: the expression of PIK3R3 in normal skin and melanoma tissues was detected by immunohistochemistry, and the silencing efficiency of c-Myc and PIK3R3 genes by lentivirus LV3-c-Myc and LV3-PIK3R3) was detected. The proliferation of melanoma cell line B16-F10 after c-Myc and PIK3R3 silencing was detected by Ed U proliferation assay. The effect of c-Myc and PIK3R3 expression on the invasion ability of B16-F10 was detected by transwell invasion assay. The effect of c-Myc and PIK3R3 expression on B16-F10 migration ability of melanoma cells was detected by scratch test. The expression of miR-199a after the silencing of c-Myc and PIK3R3 was detected by PCR. The expression of c-Myc and PIK3R3 after transfection of miR-199a mimics and miR-199a inhibitor was detected by miR-199a mimics and miR-199a inhibitor. Double luciferase reporter gene system was used to detect the effect of miR-199a on c-Myc and PIK3R3 transcriptional activities. Results: compared with normal skin tissues, the expression of PIK3R3 in melanoma tissues was significantly increased, and the proliferation of B16-F10 cells was observed after c-Myc and PIK3R3 gene silencing. The ability of invasion and metastasis was significantly decreased, the expression of miR-199a was up-regulated after silencing c-Myc and PIK3R3 gene, the expression of c-Myc and PIK3R3 was decreased after transfection of miR-199a mimics, and the expression of c-Myc and PIK3R3 was up-regulated after transfection of miR-199a inhibitor. The results of double luciferase reporter gene system showed that miR-199a could directly regulate the transcriptional activity of c-Myc and PIK3R3. Conclusion: c-Myc can regulate the expression of PIK3R3 through miR-199a, thus promoting the proliferation, invasion and migration of melanoma cells.
【作者单位】：四川省医学科学院四川省人民医院皮肤外科;四川省医学科学院四川省人民医院肾脏内科;
【基金】：四川省卫计委科研课题(No.140084)
【分类号】：R739.5

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