Autophagic Cell Death Induced by GANT61 Inhibits Proliferation of Human Breast Cancer Cell Line MCF-7
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摘要:目的
观察Hedgehog通路抑制剂GANT61对人乳腺癌MCF-7细胞的抑制作用。
方法GANT61作用MCF-7细胞后，通过流式细胞术检测细胞死亡；提取处理细胞总RNA和蛋白。分别采用Real-time PCR和Western blot检测Hedgehog通路SHH、Gli1、Gli2及自噬标志物LC3-Ⅱ表达；处理细胞固定后通过电子显微镜观察GANT61诱导的自噬体形态；自噬抑制剂3-MA处理细胞或自噬相关基因atg5 siRNA转染细胞后再用GANT61作用，流式细胞术检测细胞死亡数量。
结果GANT61能明显诱导MCF-7死亡，降低Hedgehog通路SHH、Gli1、Gli2表达水平，提高LC3-Ⅱ蛋白表达，并可诱导MCF-7细胞产生自噬体。3-MA及atg5 siRNA可减弱GANT61诱导的细胞死亡。
结论GANT61通过诱导自噬性细胞死亡机制发挥抗乳腺癌细胞活性。
Abstract:ObjectiveTo investigate the inhibitory effect of GANT61 on human breast cancer cell line MCF-7.
MethodsCell death was measured by flow cytometry after GANT61 treatment; the total RNA and protein were extracted from the treated cells; the mRNA and protein expression levels of SHH, Gli1, Gli2 and LC3-Ⅱ were detected by Real-time PCR and Western blot, respectively. Autophagosomes formation was observed under transmission electronic microscopy (TEM). After that, 3-MA and atg5 siRNA were used to illustrate the relationship between autophagy and cell death.
ResultsGANT61 induced significantly the cell death of MCF-7 cells. The expressions of SHH, Gli1 and Gli2 were decreased; however, autophagy marker LC3-II expression was increased after GANT61 treatment. Meanwhile, autophagosomes could be observed under TEM. 3-MA and atg5 siRNA could inhibit GANT61-induced cell death.
ConclusionGANT61 inhibits the proliferation of human breast cancer cell line MCF-7 through autophagic cell death.
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Key words:
- GANT61 /
- Autophagic cell death /
- Breast cancer /
- MCF-7 cell
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图 1 不同浓度GANT61诱导MCF-7细胞死亡
Figure 1 MCF-7 cells death induced by different concentration of GANT61
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图 2 不同浓度GANT61处理后MCF-7细胞SHH、Gli1、Gli2表达
Figure 2 SHH, Gli1 and Gli2 expression in MCF-7 cells treated with different concentration of GANT61
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图 4 3-MA (A)和atg5(B) siRNA抑制GANT61诱导的细胞死亡
Figure 4 3-MA (A) and atg5(B) siRNA inhibited GANT61-induced cell death
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