Smad 6调控STAT3活性促进胶质瘤细胞的增殖

Smad 6 promotes gliomagenesis via regulating STAT3 transcriptional activity

目的研究Smad 6对人脑胶质瘤细胞增殖能力的影响及其机制。方法采用Western blot检测原代胶质瘤细胞核中Smad 6的表达水平。构建慢病毒介导的细胞核稳定过表达和敲减Smad 6的胶质瘤细胞株;CCK8细胞增殖和Ed U掺入增殖实验检测Smad 6对胶质瘤细胞增殖能力的影响;干细胞成球实验检测Smad 6对胶质瘤干细胞样细胞自我更新能力的影响。裸鼠皮下荷瘤模型检测Smad 6对胶质瘤细胞体内成瘤能力的影响;荧光素酶报告基因和Western blot检测Smad6对STAT3靶基因的调控,分析Smad 6对信号转导和转录激活因子3(STAT3)活性的调节作用。结果 Western blot和细胞增殖实验显示,原代胶质瘤细胞核中Smad 6表达水平与细胞增殖能力相关。成功构建Smad 6细胞核稳定过表达和敲减胶质瘤细胞株;核Smad 6过表达显著促进胶质瘤细胞的增殖和干细胞样细胞的自我更新能力,而Smad 6敲减则显著抑制胶质瘤细胞的增殖和干细胞样细胞的自我更新能力。细胞核Smad 6促进胶质瘤细胞中STAT3的转录调控活性及其靶基因的表达。结论胶质瘤细胞核中Smad 6通过调控STAT3的转录调控活性,促进胶质瘤细胞的增殖和干细胞样细胞的自我更新能力。

Objective To investigate the effects of Smad 6 on gliomagenesis and its mechanism. Methods The expression of Smad 6 in primary glioma cells was detected by Western blot. Stable glioma cell lines with nuclear-Smad 6 overexpression or knockdown mediated by lentivirus were established. Cell proliferation was examined by CCK8 and Ed U experiments. The effect of Smad 6 on the self-renew of glioma stem-like cells was tested by tumor-sphere formation assay. Effect of Smad 6 on tumorgenesis in vivo was tested in nude mice. Luciferase reporter assay and Western blot analysis were used to examine STAT3 transcriptional activity and its targeting proteins expression. Results Western blot and cell growth assay confirmed that Smad 6 expression was associated with cell growth of patient-derived glioma cells. Smad 6 overexpression significantly promoted cell proliferation and tumor-sphere formation. Reporter assay and Western blot assay revealed that Smad 6 enhanced the activity of STAT3. Conclusion Smad 6 that expressed in nuclei of glioma cells promotes cell proliferation and self-renew of tumor stem-like cells via enhancing the transcriptional activity of STAT3.