[The effect and mechanism of 2-methoxyestradiol on human laryngeal papilloma cell line]

Lin Chuang Er Bi Yan Hou Tou Jing Wai Ke Za Zhi. 2019 Oct;33(10):945-950. doi: 10.13201/j.issn.1001-1781.2019.10.011.
[Article in Chinese]

Abstract
in English, Chinese

Objective:To explore the signal pathway that mediates the effect of 2-methoxyestradiol（2ME2） on human laryngeal papilloma cell line, in terms of cell proliferation and neovascularization. Method:HIF-1α expression of human laryngeal papilloma cell line（Hs840. T） was interfered using siRNA, and the cells were then processed by 2ME2 in two concentrations. RT-PCR and ELISA were performed to detect the difference of HIF-1α in cells with normal or lower HIF-1α mRNA level, with ELISA test of excretory VEGF level and CCK8 test of cell viability. Result:The IC50of 2ME2 in Hs840. T was 0.309 μmol/L in terms of the inhibition effect of cell proliferation（P<0.01）. Baseline level of intracellular HIF-1α was detectable, and procession of Hs840. T cells by 2ME2 of 0.4 μmol/L inhibited the transcription and expression of HIF-1α by（76.8±2.0）% and（68.6±3.5）% [vs blank group（100.0±2.7）% and（100.0±6.9）%, P<0.01]. VEGF excretion decreased to（50.8±2.1） and（28.1±4.0）% [vs blank group（100.0±3.1）%, P<0.01]after procession by 2ME2 of 0.2 μmol/L and 0.4 μmol/L. After the successful interference of HIF-1α by siRNA, the inhibition effect on cell proliferation by 2ME2 of 0.4 μmol/L decreased to（51.5±3.8）% [vs control group（65.7±1.7）%, P<0.01]. siRNA interference of HIF-1α lead to a decrease of HIF-α mRNA and protein level to（16.3±0.9）% and（7.4±0.8）% [vs cells not interfered（76.8±2.0）% vs（68.6±3.5）%, P<0.01]. Secretory VEGF dropped to（41.0±2.9）% [vs cells not interfered（28.1±4.0）%, P<0.05]. Conclusion:2ME2 has a significant inhibitory effect on human laryngeal cell line. The inhibition of cell proliferation was mediated by a lower level of HIF-1α and therefore lower VEGF. 2ME2 might serve as a novel potential therapy for patients of recurrent respiratory papillomatosis.

目的：探究2-甲氧基雌二醇（2ME2）在人喉乳头状瘤细胞株中对细胞增殖和新生血管生成的抑制效应，及介导这2种效应的信号通路。 方法：采用siRNA敲低喉乳头状瘤Hs840.T细胞株的HIF-1α表达，以高（0.4 μmol/L）、低（0.2 μmol/L）浓度的2ME2处理Hs840.T细胞，在低氧诱导因子-1α（HIF-1α）敲低前后，分别使用CCK8法检测细胞增殖抑制率，RT-PCR法检测HIF-1α mRNA水平，ELISA法检测HIF-1α蛋白和分泌性血管内皮生长因子（VEGF）水平。 结果：Hs840.T细胞对于2ME2的半数抑制浓度为0.309 μmol/L，2ME2对Hs840.T细胞的增殖有抑制作用（P<0.01）。Hs840.T细胞中检测到HIF-1α的基础表达，用0.4 μmol/L的2ME2处理细胞株，可使细胞株中HIF-1α的mRNA和蛋白水平分别降至（76.8±2.0）%和（68.6±3.5）%[空白组为（100.0±2.7）%及（100.0±6.9）%，P<0.01]。用0.2 μmol/L及0.4 μmol/L的2ME2处理细胞株后，可使细胞株中VEGF分泌量分别降为（50.8±2.1）%及（28.1±4.0）%[空白组为（100.0±3.1）%，P<0.01]。siRNA成功敲低Hs840.T细胞株中HIF-1α后，以0.4 μmol/L的2ME2处理细胞株，发现细胞增殖抑制作用降低为（51.5±3.8）%[HIF-1α未敲低组为（65.7±1.7）%，P<0.01]。同时，HIF-1α的mRNA和蛋白水平分别降至（16.3±0.9）%和（7.4±0.8）%[HIF-1α未敲低组为（76.8±2.0）%和（68.6±3.5）%，P<0.01]，VEGF分泌量降为（41.0±2.9）%[HIF-1α未敲低组为（28.1±4.0）%，P<0.05]。 结论：2ME2对人喉乳头状瘤细胞有明显的抑制作用。2ME2通过抑制HIF-1α基因的转录，从而抑制VEGF分泌，实现对人喉乳头状瘤细胞生长的抑制。2ME2可能为复发性喉乳头状瘤患者的治疗提供新思路。.

Keywords: 2-methoxyestradiol; hypoxia-inducible factor-1α; proliferation; recurrent respiratory papillomatosis.

MeSH terms

  • 2-Methoxyestradiol / metabolism*
  • Cell Line, Tumor
  • Estradiol
  • Humans
  • Hypoxia-Inducible Factor 1, alpha Subunit / metabolism
  • Laryngeal Neoplasms
  • Papilloma
  • Vascular Endothelial Growth Factor A / metabolism

Substances

  • Hypoxia-Inducible Factor 1, alpha Subunit
  • Vascular Endothelial Growth Factor A
  • Estradiol
  • 2-Methoxyestradiol