槐角黄酮调控miR-199a-3p/p38MAPK/NF-κB对乳腺癌细胞增殖、迁移、侵袭和炎症因子分泌的影响

Effect of miR-199a-3p/p38MAPK/NF-κB regulated by sophorae fructus flavonoids on cell proliferation, migration, invasion and secretion of inflammatory factors in breast cancer

  • 摘要:
    目的 探讨槐角黄酮对乳腺癌细胞增殖、迁移、侵袭和炎症因子分泌的影响。
    方法 将乳腺癌细胞BT549分为正常对照(NC)组、低剂量组、中剂量组、高剂量组、miR-NC组、miR-199a-3p组、高剂量+anti-miR-NC组、高剂量+anti-miR-199a-3p组。采用CCK-8、克隆形成实验评估细胞增殖; 采用Transwell法评估细胞迁移和侵袭。采用实时荧光定量聚合酶链反应(RT-qPCR)检测miR-199a-3p表达。采用酶联免疫吸附测定(ELISA)法检测细胞培养液上清中肿瘤坏死因子-α(TNF-α)、白细胞介素-6(IL-6)以及白细胞介素-1β(IL-1β)水平。采用蛋白免疫印迹(Western blot)检测p-p38MAPK和p-p65蛋白表达。
    结果 与NC组比较,低、中、高剂量组BT549细胞活力、克隆形成数、迁移数、侵袭数降低, miR-199a-3p表达升高,细胞培养液上清中TNF-α、IL-6以及IL-1β水平降低,差异均有统计学意义(P < 0.05)。与miR-NC组比较, miR-199a-3p组BT549细胞活力、克隆形成数、迁移数、侵袭数降低,细胞培养液上清中TNF-α、IL-6以及IL-1β水平降低,差异均有统计学意义(P < 0.05)。与高剂量+anti-miR-NC组比较,高剂量+anti-miR-199a-3p组BT549细胞活力、克隆形成数、迁移数、侵袭数升高,细胞培养液上清中TNF-α、IL-6以及IL-1β水平升高,差异均有统计学意义(P < 0.05)。与NC组比较,高剂量组BT549细胞p-p38MAPK和p-p65蛋白表达降低,差异有统计学意义(P < 0.05)。与高剂量+anti-miR-NC组比较,高剂量+anti-miR-199a-3p组BT549细胞p-p38MAPK和p-p65蛋白表达升高,差异有统计学意义(P < 0.05)。
    结论 槐角黄酮可抑制乳腺癌细胞增殖、迁移、侵袭和炎症反应,其机制可能是通过上调miR-199a-3p表达、抑制p38MAPK/NF-κB通路实现的。

     

    Abstract:
    Objective To investigate the effect of sophorae fructus flavonoids on cell proliferation, migration, invasion and secretion of inflammatory factors in breast cancer.
    Methods The breast cancer cells BT549 were divided into normal control (NC) group, low-dose group, medium-dose group, high-dose group, miR-NC group, miR-199a-3p group, high-dose+anti-miR-NC group, and high-dose+anti-miR-199a-3p group. CCK-8 and clone formation assays were used to evaluate cell proliferation; Transwell method was performed to evaluate cell migration and invasion. Real-time fluorescence quantitative polymerase chain reaction (RT-qPCR) was used to detect the expression of miR-199a-3p. EILSA method was used to detect the levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-1β (IL-1β) in the cell culture supernatant. Expression levels of p-p38MAPK and p-p65 proteins were detected by Western blot.
    Results Compared with the NC group, the cell viability, clone formation number, migration number and invasion number of BT549 cells in the low-, medium- and high-dose groups were significantly reduced, the expression of miR-199a-3p was significantly increased, and the levels of TNF-α, IL-6 and IL-1β in the cell culture supernatant were significantly reduced (P < 0.05). Compared with the miR-NC group, the cell viability, clone formation number, migration number and invasion number of BT549 cell in the miR-199a-3p group were significantly reduced, and the levels of TNF-α, IL-6 and IL-1β in the cell culture supernatant were significantly reduced as well (P < 0.05). Compared with the high-dose+anti-miR-NC group, the cell viability, clone formation number, migration number and invasion number of BT549 in the high-dose+anti-miR-199a-3p group were significantly increased, and the levels of TNF-α, IL-6 and IL-1β in the cell culture supernatant were significantly increased as well (P < 0.05). Compared with the NC group, the expression levels of p-p38MAPK and p-p65 proteins of BT549 cells in the high-dose group were significantly reduced (P < 0.05). Compared with the high-dose+anti-miR-NC group, the expression levels of p-p38MAPK and p-p65 proteins of BT549 cells in high-dose+anti-miR-199a-3p group were significantly increased (P < 0.05).
    Conclusion Sophorae fructus flavonoids can inhibit cell proliferation, migration, invasion and inflammation reaction in breast cancer, and its mechanism may be achieved by up-regulating the expression of miR-199a-3p and inhibiting p38MAPK/NF-κB pathway.

     

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