Circ_UBE2C promotes proliferation and glycolysis of lung cancer cells by regulating miR-107/HK2 axis

Ji NIE; Chen LIU; Dandan PU; Mei CHA; Yunhui ZHANG

doi:10.16016/j.2097-0927.202311127

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Circ_UBE2C promotes proliferation and glycolysis of lung cancer cells by regulating miR-107/HK2 axis

Basic Medicine | 更新时间：2026-01-09

- Circ_UBE2C promotes proliferation and glycolysis of lung cancer cells by regulating miR-107/HK2 axis
- Journal of Army Medical University Vol. 46, Issue 15, Pages: 1729-1739(2024)
- 作者机构：
  
  1.650032 昆明，云南省第一人民医院，昆明理工大学附属医院呼吸与危重症医学科
  2.650093 昆明，昆明理工大学医学院
- 作者简介：
- 基金信息：
  
  the Open Project of Yunnan Provincial Clinical Medical Center(2021LCZXXF-HX09);the Research Fund for Doctoral Program of Yunnan Provincial First People's Hospital(KHBS-2022-030)
- DOI：10.16016/j.2097-0927.202311127
  CLC： R394.3;R730.23;R734.2
- Received：27 November 2023，
  
  Revised：2024-01-22，
  
  Published：15 August 2024
- 稿件说明：
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Ji NIE, Chen LIU, Dandan PU, et al. Circ_UBE2C promotes proliferation and glycolysis of lung cancer cells by regulating miR-107/HK2 axis[J]. Journal of Army Medical University, 2024, 46(15): 1729-1739.
DOI：

Ji NIE, Chen LIU, Dandan PU, et al. Circ_UBE2C promotes proliferation and glycolysis of lung cancer cells by regulating miR-107/HK2 axis[J]. Journal of Army Medical University, 2024, 46(15): 1729-1739. DOI： 10.16016/j.2097-0927.202311127.

摘要

目的

探讨circ_UBE2C对肺癌细胞增殖和糖酵解的影响及其作用机制。

方法

基于TCGA数据库分析circ_UBE2C、miR-107和己糖激酶2(hexokinase 2，HK2)在肺癌组织和正常肺组织中的表达情况。采用Kaplan-Meier法绘制生存曲线，分析circ_UBE2C/miR-107/HK2表达量和肺癌患者生存期的相关性。常规培养人正常肺上皮细胞(BEAS-2B)和人肺癌细胞株(A549、NCI-H460和NCI-H1299)，采用qRT-PCR检测circ_UBE2C和miR-107的表达量，Western blot检测HK2和PCNA蛋白的表达量。将A549和NCI-H1299细胞分为空白组(NC)、circ_UBE2C敲减组(si-circ)、HK2敲减组(si-HK2)、同时敲减miR-107+HK2组(miR-inhibitor+si-HK2)和同时敲减circ_UBE2C+miR-107组(si-circ+miR-inhibitor)。CCK-8和克隆形成实验分析细胞增殖活力。葡萄糖摄取比色测定试剂盒、乳酸检测试剂盒和ATP含量测定试剂盒分析A549和NCI-H1299细胞葡萄糖摄取量、乳酸生成量和ATP产生量。Seahorse XF糖酵解应激测试试剂盒和Seahorse XF细胞线粒体应激测试试剂盒检测细胞外酸化率(extracellular acidification ratio，ECAR)和细胞耗氧率(oxygen consumption ratio，OCR)。双荧光素酶报告基因实验评估circ_UBE2C和miR-107、miR-107和HK2的靶向关系。

结果

与正常组织相比，肺癌组织中circ_UBE2C和HK2表达上调，miR-107表达下调(

0.05)。与BEAS-2B细胞相比，A549和NCI-H1299细胞中circ_UBE2C和HK2表达量升高，miR-107表达下调(

0.05)。生存分析显示，circ_UBE2C和HK2高表达或miR-107低表达肺癌患者的生存期较短(

0.05)。相比于NC组，si-circ或si-HK2组A549和NCI-H1299细胞增殖活力、葡萄糖摄取量、乳酸生成量、ATP产生量、ECAR和OCR均显著减少(

0.05)。双荧光素酶报告基因证实，circ_UBE2C靶向负调控miR-107，且HK2是miR-107的靶基因。与si-HK2组相比，miR-inhibitor+si-HK2组和si-circ+miR-inhibitor组中A549和NCI-H1299细胞增殖活力和糖酵解水平显著增加。

结论

敲减circ_UBE2C可显著抑制肺癌细胞增殖和糖酵解水平，其机制是通过靶向上调miR-107抑制HK2表达。

Abstract

Objective

To investigate the effects of circ_UBE2C on the proliferation and glycolysis of lung cancer cells and its mechanism of action.

Methods

The expression of circ_UBE2C

miR-107

and hexokinase 2 (HK2) in lung cancer tissues and normal lung tissues were analyzed based on the The Cancer Genome Atlas (TCGA) database. Kaplan-Meier survival curve was plotted to analyze the correlation between circ_UBE2C/miR-107/HK2 expression and survival of the lung cancer patients. qRT-PCR was used to detect the expression of circ_UBE2C and miR-107

and Western blotting was employed to measure the expression of HK2 and PCNA in human normal lung epithelial cells (BEAS-2B) and human lung cancer cell lines (A549

NCI-H460

and NCI-H1299). Then A549 and NCI-H1299 cells were divided into the blank group (NC)

circ_UBE2C knockdown group (si-circ)

HK2 knockdown group (si-HK2)

simultaneous knockdown of miR-107+HK2 group (miR-inhibitor+si-HK2)

and simultaneous knockdown of circ_UBE2C+miR-107 group (si-circ+miR-inhibitor). CCK-8 assay and colony formation assay were employed to measure the proliferation of above cell groups. The glucose uptake

lactate generation

and ATP production in the cells were measured by glucose uptake colorimetric assay kit

lactic acid detection kit

and ATP content determination kit

respectively. Seahorse XF glycolytic stress test and Seahorse XF cellular mitochondrial stress test were respectively performed to detect the extracellular acidification ratio (ECAR) and cellular oxygen consumption ratio (OCR). The targeting relationship between the circ_UBE2C and miR-107

and miR-107 and HK2 was validated by dual-luciferase reporter gene assay.

Results

Compared with normal lung tissues

the expression of circ_UBE2C and HK2 was up-regulated

while that of miR-107 was down-regulated in lung cancer tissues (

0.05). The expression of circ_UBE2C and HK2 was elevated

and that of miR-107 was reduced in A549 and NCI-H1299 cells when compared with BEAS-2B cells (

0.05). Survival analysis showed that the patients with high expression of circ_UBE2C and HK2 or low expression of miR-107 had shorter survival (

0.05). Compared with the NC group

both si-circ and si-HK2 resulted in significantly reduced proliferation

glucose uptake

lactate generation

ATP production

and ECAR and OCR values in A549 and NCI-H1299 cells (

0.05). Dual luciferase reporter gene assay confirmed that circ_UBE2C could directly bind to and negatively regulate miR-107 expression. HK2 was then identified as a downstream target of miR-107. Compared with the si-HK2 group

the proliferative ability and glycolysis level of A549 and NCI-H1299 cells were significantly increased in the miR-inhibitor+si-HK2 group and the si-circ+miR-inhibitor group.

Conclusion

Knockdown of circ_UBE2C significantly suppresses the proliferation and glycolysis of lung cancer cells

via

targeting up-regulation of miR-107 and then exerting inhibitory effect on HK2 expression.

关键词

Keywords

references

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