Establishment of cell model of breast cancer-associated fibroblasts’ energy metabolism reprogramming induced by hypoxia

Tang Shifu; Zhou Mingli; Sun Yifan; Wang Linchun; Liu Chunming; Yang Li; Tang Xi; Zhang Hailong; Gao Chao; Huang Guangming; Qu Haiyang; Du Yan’e; Xu Liyun; Wen Siyang; Lang Lei; Liu Manran

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Establishment of cell model of breast cancer-associated fibroblasts’ energy metabolism reprogramming induced by hypoxia

更新时间：2023-12-18

- Establishment of cell model of breast cancer-associated fibroblasts’ energy metabolism reprogramming induced by hypoxia
- Journal of Chongqing Medical University Vol. 40, Issue 8, Pages: 1084-1088(2015)
- 作者机构：
  
  1. 广西中医药大学第三附属医院检验科,柳州,545001
  2. 重庆医科大学临床检验诊断学教育部重点实验室,重庆,400016
  3. 四川省达州市中西医结合医院消化内科,达州,635000
  4. 重庆医科大学脂糖代谢性疾病重庆市重点实验室脂质研究中心,重庆,400016
- 作者简介：
- 基金信息：
- DOI：
  CLC：
- Published：2015，
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Tang Shifu, Zhou Mingli, Sun Yifan, et al. Establishment of cell model of breast cancer-associated fibroblasts’ energy metabolism reprogramming induced by hypoxia. [J]. Journal of Chongqing Medical University 40(8):1084-1088(2015)
DOI：

Tang Shifu, Zhou Mingli, Sun Yifan, et al. Establishment of cell model of breast cancer-associated fibroblasts’ energy metabolism reprogramming induced by hypoxia. [J]. Journal of Chongqing Medical University 40(8):1084-1088(2015) DOI：

摘要

目的：构建低氧诱导的乳腺癌癌相关成纤维细胞（cancer-associated fibroblasts，CAF）能量代谢重塑（energy metabolism reprogramming，EMR）细胞模型，为其后续分子机制和生物学功能研究奠定基础。方法：以成对的永生化的乳腺癌癌旁组织来源正常成纤维细胞（normal fibroblasts，NF）和CAF为处理对象，根据[O2]不同，NF和CAF细胞各分为常氧处理组和低氧处理组。不同氧浓度分别处理NF和CAF细胞0、1、3、6、12 h和24 h。葡萄糖消耗实验和乳酸生成实验检测细胞糖酵解水平；线粒体活性检测实验评估细胞氧化磷酸化（oxidative phosphorylation，OP）活性；Western blot实验检测细胞能量代谢相关蛋白包括单羧酸转运子4（monocarboxylate transporters 4，MCT4）、细胞色素氧化酶Ⅳ亚基（cytochrome oxidase Ⅳ subunit，COX Ⅳ）和低氧诱导因子1α亚基（hypoxia inducible factor 1α subunit，HIF1α）的蛋白表达量。结果：低氧与CAF细胞EMR呈剂量效应和时间效应关系，1%[O2]作用24 h为最佳低氧处理条件。与NF常氧组相比，低氧处理（1%[O2]作用24 h）使CAF葡萄糖吸收和乳酸产生能力分别增强2倍和3.1倍，明显抑制其氧化磷酸化活性，使CAF细胞中MCT4和HIF1α蛋白表达量分别上升3.1倍和3.9倍及COX Ⅳ下调90%。结论：低氧（1%[O2]作用24 h）使CAF具有典型的EMR表型。本研究成功构建低氧诱导的乳腺癌CAF细胞EMR模型。

Abstract

Objective：To establish the cell model of energy metabolism reprogramming（EMR） of breast cancer-associated fibroblasts（CAF） induced by hypoxia and to lay a firm foundation for subsequently exploring its molecular mechanism and biological functions. Methods：Based on the O2 concentration，paired immortalized normal fibroblasts（NF） and CAF derived from the same breast cancer patient were divided into hypoxic group and normoxic group. NF/CAF were treated with different O2 concentrations for 0，1，3，6，12 and 24 h. The glycolysis activity of NF and CAF was tested by the glucose consumption assay and the lactate generation assay；the oxidative phosphorylation activity of NF and CAF was detected by the mitochondrial activity assay；the protein expression levels of MCT4，COX Ⅳ and HIF1α were determined by Western blot；the data were analyzed with the SPSS 17.0 software. Results：There was dose-effect and time-effect between hypoxic levels and the EMR degree of CAF. 1%[O2] for 24 h was optimum for inducing the EMR of CAF. Hypoxic treatment increased the glycolysis activity，suppressed the oxidative phosphorylation levels，in－creased the protein expression of MCT4 and HIF1α 2 times or 3.9 times respectively，and decreased the protein expression of COX Ⅳ by 10% in CAF compared with NF. Conclusion：Hypoxic treatment（1%[O2] for 24 h） maked CAF have a typical EMR pheno－type. It is successfully established that the cell model of the EMR of breast cancer-derived CAF induced by hypoxia（1%[O2] for 24 h）.

关键词

乳腺癌癌相关成纤维细胞低氧能量代谢重塑细胞模型

Keywords

breast cancercancer-associated fibroblastshypoxiaenergy metabolism reprogrammingcell model

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