Wang Qing-Xuan, Lv Lin, Ye Dan-Rong, Sun Yi-Han, Pan Xin-Xin, Bhandari Adheesh, Zhang Xiao-Hua, Wang Ou-Chen, Liu Hai-Guang
Department of Thyroid and Breast Surgery, The First Affiliated Hospital of Wenzhou Medical University, Wenzhou, Zhejiang Province.
Department of Oncology, Jinhua Municipal Central Hospital, Jinhua Hospital of Zhejiang University, Jinhua, China.
Ann Clin Lab Sci. 2019 Sep;49(4):481-487.
Chemotherapy resistance reduces the effectiveness of chemotherapeutic drugs greatly, resulting in treatment failure. Therefore, exploring chemoresistance-related genes and the corresponding mechanism is extremely important. The central role of CD44v6 in colorectal cancer has been previously reported. However, the effects of CD44v6 gene knockdown on the chemosensitivity of colorectal cancer cells are not conclusive.
A stable CD44v6 knockdown cell model in HT29 cells (HT29-KD) was established via lentiviral transduction. A quantitative real-time polymerase chain reaction (PCR) was carried out to confirm the knockdown efficiency. The chemosensitivity of cells to 5-fluorouracil (5-FU) was determined by a cell counting kit (CCK)-8 assay. Cell apoptosis and the cell cycle were assessed by flow cytometry.
The CD44v6 knockdown cell model was successfully constructed by using lentiviral transduction. Upon treatment with 5-FU, the inhibitory rate for cell activity of HT29-KD cells was significantly higher than that of the control group (HT29-NC). CD44v6 gene knockdown did not significantly affect HT-29 cell proliferation, according to the CCK-8 assay and cell cycle analysis. The cell apoptosis assay revealed that CD44v6 gene knockdown promoted HT-29 cell apoptosis. Without 5-FU treatment, there was no significant difference in terms of the relative expression level of the autophagy-related gene BECN1 between the two groups. However, with 5-FU treatment, the relative expression level of BECN1 in HT29-KD cells was much lower than that in HT29-NC cells.
Our study confirms that CD44v6 gene knockdown can enhance chemosensitivity in HT29 cells by promoting apoptosis and inhibiting autophagy, thus affirming the effects of CD44v6 on the chemosensitivity of colorectal cancer.
化疗耐药性极大地降低了化疗药物的有效性，导致治疗失败。因此，探索与化疗耐药相关的基因及其相应机制极为重要。先前已有报道CD44v6在结直肠癌中起核心作用。然而，CD44v6基因敲低对结直肠癌细胞化疗敏感性的影响尚无定论。
通过慢病毒转导在HT29细胞中建立稳定的CD44v6敲低细胞模型（HT29-KD）。进行定量实时聚合酶链反应（PCR）以确认敲低效率。通过细胞计数试剂盒（CCK）-8法测定细胞对5-氟尿嘧啶（5-FU）的化疗敏感性。通过流式细胞术评估细胞凋亡和细胞周期。
利用慢病毒转导成功构建了CD44v6敲低细胞模型。用5-FU处理后，HT29-KD细胞的细胞活性抑制率显著高于对照组（HT29-NC）。根据CCK-8法检测和细胞周期分析，CD44v6基因敲低对HT-29细胞增殖无显著影响。细胞凋亡检测显示，CD44v6基因敲低促进了HT-29细胞凋亡。在未用5-FU处理时，两组自噬相关基因BECN1的相对表达水平无显著差异。然而，在用5-FU处理后，HT29-KD细胞中BECN1的相对表达水平远低于HT29-NC细胞。
我们的研究证实，CD44v6基因敲低可通过促进凋亡和抑制自噬增强HT29细胞的化疗敏感性，从而肯定了CD44v6对结直肠癌化疗敏感性的影响。
