实验动物与比较医学 ›› 2020, Vol. 40 ›› Issue (5): 403-.DOI: 10.3969/j.issn.1674-5817.2020.05.007

• 论著 • 上一篇    下一篇

草氨酸钠对胆管癌细胞自噬与凋亡的影响

何大伟1,何敖林1,周明慧1,于亚平2,张明华2   

  1. 1. 昆山市第一人民医院临床实验研究中心,昆山 215300;
    2. 昆山市第一人民医院肝胆外科,昆山 215300
  • 收稿日期:2019-12-06 出版日期:2020-10-15 发布日期:2020-11-19
  • 作者简介:何大伟(1987—), 男, 助理研究员, 研究方向: 细胞自噬与肿瘤的发生发展。 E-mail: 824171022@qq.com
  • 基金资助:
    昆山市科技计划项目(KS1537)

Effects of Sodium Oxalate on Autophagy and Apoptosis of Cholangiocarcinoma Cells#br#

HE Dawei1, HE Aolin1, ZHOU Minghui1, YU Yaping2, ZHANG Minghua2   

  1. 1. Clinical Experimental Research Center,  2. Department of Hepatobiliary surgery First People’s Hospital of Kunshan, Kunshan 215300, Jiangsu Province, China
  • Received:2019-12-06 Online:2020-10-15 Published:2020-11-19

摘要: 目的   观察乳酸脱氢酶A(lactate dehydrogenase A,LDHA)抑制剂草氨酸钠对人胆管癌细胞株Hucct-1自噬与凋亡的影响,并初步探讨其相关机制。方法    用不同浓度的草氨酸钠处理Hucct-1细胞,或将LDHA小干扰RNA 转染Hucct-1细胞后,CCK8法检测细胞增殖,流式细胞术检测细胞凋亡,划痕愈合实验检测细胞迁移,实时荧光定量PCR和蛋白质印迹法分别检测自噬相关mRNA和蛋白的表达水平,乳酸检测试剂盒检测细胞内乳酸的含量。结果   草氨酸钠抑制Hucct-1细胞增殖(P<0.01),并呈剂量依赖性,其半数抑制浓度为50 mmol/L。用50 mmol/L草氨酸钠干预Hucct-1细胞或LDHA小干扰RNA转染Hucct-1细胞后,LDHA mRNA和蛋白表达水平均降低(均P<0.01),细胞内乳酸的含量减少(P<0.01),细胞凋亡率升高(P<0.01),细胞迁移距离缩短(P<0.01),细胞中自噬相关基因7(autophagy-related gene 7,ATG7)、Beclin-1和微管相关蛋白1轻链3Ⅱ(microtubule-associated protein 1 light chain 3Ⅱ,LC-3Ⅱ)表达水平升高(均P<0.01),P62蛋白表达水平降低(P<0.01)。结论   草氨酸钠通过抑制Hucct-1细胞中LDHA表达来促进细胞自噬,诱导细胞凋亡。



关键词: 草氨酸钠, 胆管癌, 乳酸脱氢酶A, 细胞自噬, 细胞凋亡

Abstract:

Objective    To observe the effects of sodium oxalate, an inhibitor of lactate dehydrogenase A (LDHA), on autophagy and apoptosis of cholangiocarcinoma cell line Hucct-1, and to explore the related mechanisms. Methods    Hucct-1 cells were treated with different concentrations of sodium oxalate. And LDHA siRNA was transfected into Hucct-1 cells by LipofectAMINE 2000 and Opti-MEM medium. Then the cell proliferation was detected by CCK8 assay, the apoptosis was detected by flow cytometry, and the cell migration was detected by scratch healing test. Meanwhile, the expressions of autophagy-related mRNAs and proteins were detected respectively by RT-PCR and Western blotting. And the lactic acid detection kit was used to detect lactate content in cells. Results    Sodium oxalate inhibited the proliferation of Hucct-1 cells in a dose-dependent manner, its  half-inhibitory concentration increased was 50 mmol/L. After Hucct-1 cells were treated with 50 mmol/L sodium oxalate or transfected with LDHA siRNA, the mRNA and protein expression levels of LDHA were decreased (both P<0.01), and the intracellular lactic acid content was decreased (P<0.01). In the meantime, the apoptosis rate was increased (P<0.01), and the cell migration distance was decreased (P<0.01). And the expression levels of autophagy-related gene 7 (ATG7), Beclin-1 and microtubule-associated protein 1 light chain 3Ⅱ (LC-3Ⅱ) were increased (all P<0.01). In addition, the expression of P62 protein was significantly decreased (P<0.01). Conclusion    Sodium oxalate can promote autophagy and induce apoptosis by inhibiting the expression of LDHA in Hucct-1 cells.


Key words: Sodium oxalate, Cholangiocarcinoma, Lactate dehydrogenase A, Autophagy, Apoptosis     

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