Psoralen reverses glutathione-S-transferase π-mediated multidrug resistance in breast cancer stem cells

doi:10.3969/j.issn.2095-4344.2017.13.007

Abstract

Abstract:

BACKGROUND: Breast cancer stem cells not only lead to the occurrence of breast cancer, but also may cause breast cancer metastasis and recurrence. The relationship between stem cells and cell resistance is also gaining increasing attentions, and the focus on the stem cell treatment may result in unexpected results.
OBJECTIVE: To explore the reversal effect of psoralen on glutathione-S-transferase π (GST-π) in human breast cancer MCF-7/ADR cells and its mechanism.
METHODS: MCF-7/ADR cells were cultured and enriched in serum-free medium to obtain breast cancer stem cells. RT-PCR and western blot were used to detect the expression of GST-π at the levels of gene and protein in the MCF-7/ADR cells after treatment with 0, 4, 8, 12, 16 mg/L psoralen. To observe the activation of nuclear factor-κB, western blot was used. The expression of GST-π was detected by RT-PCR in 18 μmol/L SN50 group and 8 mg/L psoralen group. Cell counting kit-8 assay was used to detect the effect of doxorubicin on cell proliferation.
RESULTS AND CONCLUSION: Compared with the control group, psoralen reduced the expression of GST-π at the mRNA and protein levels, and significantly inhibited the activation of nuclear factor-κB. It was suggested that psoralen could reverse the multidrug resistance of human breast cancer MCF-7/ADR stem cells by decreasing the expression level of GST-π. The mechanism may be achieved by inhibiting the nuclear factor-κB signal pathway.

中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程

Key words: Psoralens, Breast Neoplasms, NF-kappa B, Tissue Engineering

CLC Number:

R394.2

Hua Yi-tong, Wang Xiao-hong, Xu Cheng-feng, Cheng Kai, Sun Hong-guang, Zhang Ying-zhe, Liu Jian, Yang Zhen-lin. Psoralen reverses glutathione-S-transferase π-mediated multidrug resistance in breast cancer stem cells[J]. Chinese Journal of Tissue Engineering Research, 2017, 21(13): 2003-2008.

Figures/Tables 1

2.1 无血清培养条件下可有效富集乳腺癌干细胞在无血清培养基中，细胞聚集生长趋势比较明显，同时也会有一部分细胞凋亡。相关文献报道，聚集成球细胞干细胞比例明显增高[11]。作者所在实验室前期实验也证实这点，多次检测结果表明成球细胞保持干细胞特性[9]。此次实验显示MCF-7/ADR在完全培养基中贴壁生长，在无血清培养条件下出现聚集成球，贴壁不紧密现象，且有悬浮生长趋势(图1)。 2.2 补骨脂素对MCF-7/ADR细胞GST-π mRNA表达水平的影响 0，4，8，12，16 mg/L补骨脂素处理MCF-7/ADR细胞48 h后，与对照组(0 mg/L)相比，各种质量浓度补骨脂素组MCF-7/ADR细胞中GST-π mRNA表达水平下调(P < 0.05)，但不同质量浓度补骨脂素组GST-π mRNA表达水平差异无显著性意义(P > 0.05；图2)。 2.3 补骨脂素对MCF-7/ADR干细胞中GST-π蛋白表达水平的影响 Western Blot 检测结果表明，MCF-7/ADR细胞存在GST-π蛋白水平的高表达。0，4，8，12，16 mg/L补骨脂素处理MCF-7/ADR细胞后，GST-π蛋白表达水平明显下降(P < 0.05)，且不同质量浓度补骨脂素组GST-π蛋白表达水平差异无显著性意义(P > 0.05；图3)。 2.4 补骨脂素对核因子κB活化状态的影响选取8 mg/L补骨脂素处理细胞后，Western Blot 检测结果发现补骨脂素能明显抑制核因子κB从胞质向胞核内转移，只有活化后的核因子κB才能进入细胞核发挥作用[12]，说明补骨脂素能够阻止其向核内转移，明显减弱核因子κB的活化状态(P < 0.05；图4)。 2.5 核因子κB特异性抑制剂对补骨脂素作用的影响 RT-PCR技术检测结果表明，补骨脂素和SN50都能降低GST-π的表达，补骨脂素抑制作用不如SN50明显(P < 0.05)。两者合用对GST-π的抑制作用最强(图5)。 2.6 补骨脂素对逆转耐药性的影响 CCK-8结果显示，不同药物处理后的微球体，阿霉素对其细胞增殖的影响也不同。结果显示，对照组MCF-7/ADR的IC50为(90.37±5.78) mg/L，补骨脂素组MCF-7/ADR的IC50为(79.96±2.93) mg/L，SN50组MCF-7/ADR的IC50为(59.17±3.97) mg/L，补骨脂素+SN50组MCF-7/ADR的IC50为(50.32±3.01) mg/L。可见MCF-7/ADR细胞微球体对阿霉素的IC50明显高于其他处理组细胞，进一步证明补骨脂素能逆转细胞耐药。"

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