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SRC抑制胶质母细胞瘤的增殖、迁移、侵袭和干性维持的分子机制

SRC Regulates Glioma Proliferation, Migration, Invasion and Cancer Stem-like Cell Self-renewal

  • 摘要: 目的 研究SRC在胶质母细胞瘤发生发展中的作用,并初步探讨可能的分子机制。方法 采用生物信息学的方法分析SRC在胶质母细胞瘤中的表达变化;利用shRNA下调胶质瘤母细胞系U87MG中SRC的表达,通过RT-PCR和免疫印迹法验证其抑制效率,并筛选出稳定干涉的细胞株;采用WST-1法、划痕愈合实验和Transwell迁移实验检测SRC shRNA干涉后细胞增殖、迁移和侵袭能力的变化;利用干细胞培养液筛选出SRC shRNA稳定干涉的胶质瘤干细胞,观察SRC shRNA对肿瘤干细胞干性的影响;利用细胞免疫荧光法观察干性基因SOX2的表达变化。结果 在胶质母细胞瘤标本中SRC的表达水平高于对照组,筛选到两条有效的SRC shRNA序列;通过shRNA下调SRC的表达后可以显著抑制胶质瘤母细胞U87MG的增殖、迁移、侵袭和肿瘤干细胞干性维持,并且可以明显抑制SOX2的表达。结论 SRC通过调控胶质母细胞瘤的增殖、迁移、侵袭和干性维持影响其发生发展,其对干性维持的作用可能是通过影响SOX2的表达实现的。

     

    Abstract: Objective To investigate the effects of SRC on the proliferation, migration and stemness of glioblastoma multiforme(GBM), and to explore its possible mechanism. Methods The Oncomine database was used to detect the SRC expression in GBM. shRNA was applied to specifically suppress the expression of SRC in U87MG cells; and the inhibition rate was detected by RT-PCR and Western blot, then the stable cell lines were obtained . The effects of SRC shRNA intervention on cell proliferation, migration and invasion were investigated by WST-1, wound-healing test and Transwell migration and invasion test, respectively. In order to examine the effects of SRC on glioma self-renewal, we used stem cell cultures to screen SRC shRNA glioma stem cells. U87MG cells were cultured in serum free medium(SFM) and tumor sphere formation was observed. The expression of stemness-related gene SOX2 was detected by immunofluorescence staining. Results SRC expression was increased in glioblastoma specimens compared with the controls. Knockdown of SRC inhibited the migration, invasion, proliferation and cancer stem cell stemness of U87MG cells. Furthermore, down-regulation of SRC expression inhibited the proliferation, migration, invasion and selfrenewal capacity of U87 glioma stem-like cells, and the expression level of the GSC marker SOX2 was decreased. Conclusion SRC regulates the GBM cell proliferation, migration and glioma stem-like cell selfrenewal, and it regulates the stemness through influencing the expression of SOX2.

     

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