基础研究(Basic Research)

巨噬细胞移动抑制因子促进有氧糖酵解与直肠癌细胞耐药性的关系

Published at: 2017年第26卷第10期

黄荣 1 , 汪泓 1 , 陈保华 1 , 樊明湖 1
1 中国人民解放军第一八四医院 普通外科,江西 鹰潭 335000
通讯作者 泓 汪 Email: a13667010409@163.com
DOI: 10.3978/j.issn.1005-6947.2017.10.007
基金:

摘要

目的:探讨巨噬细胞移动抑制因子(MIF)在结直肠癌细胞耐药中的作用和机制。方法:用5-氟尿嘧啶(5-FU)药物浓度持续递增法诱导人结直肠癌细胞株LoVo细胞构建5-FU耐药的人结肠癌LoVo/5-FU细胞。检测指标包括细胞对5-FU的敏感性(IC50)、细胞MIF蛋白表达、细胞葡萄糖摄取水平、细胞中乳酸脱氢酶(LDH)活性、细胞培养基上清中乳酸水平,分别用CCK-8法、Western blot、2-NBDG法、微孔法、试剂盒法检测。比较LoVo/5-FU细胞与亲本LoVo细胞间上述指标的差异,并检测用siRNA或慢病毒技术干扰或过表达MIF蛋白,或用PFKFB3抑制剂PFK-15抑制有氧糖酵解后,LoVo/5-FU细胞上述指标的变化。结果:成功构建LoVo/5-FU细胞,该细胞的MIF蛋白表达、对5-FU的IC50、葡萄糖摄取、LDH活性和乳酸生成水平都较其亲本LoVo细胞明显升高(均P<0.05);在LoVo/5-FU细胞上,siRNA干扰MIF后表现为MIF蛋白表达、对5-FU的IC50、葡萄糖摄取、LDH活性和乳酸生成水平均明显减少,而MIF蛋白过表达后表现为上述指标的明显升高(均P<0.05);用PFK-15抑制有氧糖酵解后,LoVo/5-FU细胞对5-FU的IC50、葡萄糖摄取、乳酸水平均明显降低(均P<0.05),但LDH活性无明显变化(P>0.05),PFK-15对MIF过表达的LoVo/5-FU细胞也有同样作用(均P<0.05)。结论:MIF通过上调LoVo细胞有氧糖酵解,诱导其对5-FU耐药能力的增加。


Relationship of macrophage migration inhibition factor enhancing aerobic glycolysis to drug resistance of colorectal cancer cells

Abstract

Objective: To investigation the action and mechanism of macrophage migration inhibition factor (MIF) in drug resistance of colorectal cancer. Methods: The 5-fluorouracil (5-FU)-resistant human colon cancer LoVo/5-FU cells were established by stepwise exposure of human colon cancer LoVo cells to increasing concentrations of 5-FU. The studied parameters included the sensitivity of cells to 5-FU (IC50), MIF protein expression, glucose uptake ability of cells, lactic dehydrogenase (LDH) activity of cells and lactate production from cultured cells supernatant, which were detected by CCK-8 assay, Western blot analysis, 2-NBDG method, microporous assay and kit assay, respectively. The differences in above parameters between LoVo/5-FU cells and their parent LoVo cells were compared, and changes in these parameters in LoVo/5-FU cells before and after MIF interference and overexpression by siRNA and lentivirus transfection, or inhibition of aerobic glycolysis by PFKFB3 inhibitor PFK-15 were examined. Results: The LoVo/5-FU cells were successfully constructed, which showed significantly increased MIF protein expression, IC50 to 5-FU, glucose uptake, LDH activity and lactate production level compared with their parent LoVo cells (all P<0.05). In LoVo/5-FU cells, the IC50 to 5-FU, glucose uptake, LDH activity and lactate production level were significantly decreased after MIF interference by siRNA, while those above parameters were significantly increased after MIF overexpression (all P<0.05). After inhibition of aerobic glycolysis by PFK-15, the IC50 to 5-FU, glucose uptake and lactate production level in LoVo/5-FU cells were significantly decreased (all P<0.05), but the LDH activity showed no significant change (P>0.05), and the same effects were exerted by PFK-15 in LoVo/5-FU cells with MIF overexpression (all P<0.05). Conclusion: MIF enhanced the resistance of LoVo cells to 5-FU by increasing the aerobic glycolysis.


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引用

引用本文: 荣 黄, 泓 汪, 保华 陈, 明湖 樊. 巨噬细胞移动抑制因子促进有氧糖酵解与直肠癌细胞耐药性的关系[J]. 中国普通外科杂志, 2017, 26(10): 1265-1271.
Cite this article as: HUANG Rong, WANG Hong, CHEN Baohua, FAN Minghu . Relationship of macrophage migration inhibition factor enhancing aerobic glycolysis to drug resistance of colorectal cancer cells[J]. Chin J Gen Surg, 2017, 26(10): 1265-1271.