转录组学测序揭示WNK2促进体外培养卵巢癌细胞ROS产生

李凤杰; 贾永琴; 王延洲; 杜秀蓉; 曹兰琴; 周丹; 梁志清

doi:10.16016/j.2097-0927.202205141

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转录组学测序揭示WNK2促进体外培养卵巢癌细胞ROS产生

基础医学 | 更新时间：2026-01-09

- 转录组学测序揭示WNK2促进体外培养卵巢癌细胞ROS产生
- Transcriptomic sequencing reveals that WNK2 promotes reactive oxygen species production in ovarian cancer cells
- 陆军军医大学学报 2023年45卷第2期页码：130-138
- 作者机构：
  
  1.400038 重庆，陆军军医大学(第三军医大学)第一附属医院妇产科
  2.410008 长沙，中南大学湘雅医院妇科
- 作者简介：
  
  [梁志清, E-mail：zhi_qingliang@163.com LIANG Zhiqing, E-mail: zhi_qingliang@163.com
- 基金信息：
  
  国家重点研发计划项目(2019YFC1005202)
- DOI：10.16016/j.2097-0927.202205141
  中图分类号： R341;R394.3;R737.31
- 收稿：2022-05-26，
  
  修回：2022-07-20，
  
  纸质出版：2023-01-30
- 稿件说明：
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李凤杰, 贾永琴, 王延洲, 等. 转录组学测序揭示WNK2促进体外培养卵巢癌细胞ROS产生[J]. 陆军军医大学学报, 2023,45(2):130-138.

Fengjie LI, Yongqin JIA, Yanzhou WANG, et al. Transcriptomic sequencing reveals that WNK2 promotes reactive oxygen species production in ovarian cancer cells[J]. Journal of Army Medical University, 2023, 45(2): 130-138.
李凤杰, 贾永琴, 王延洲, 等. 转录组学测序揭示WNK2促进体外培养卵巢癌细胞ROS产生[J]. 陆军军医大学学报, 2023,45(2):130-138. DOI： 10.16016/j.2097-0927.202205141.

Fengjie LI, Yongqin JIA, Yanzhou WANG, et al. Transcriptomic sequencing reveals that WNK2 promotes reactive oxygen species production in ovarian cancer cells[J]. Journal of Army Medical University, 2023, 45(2): 130-138. DOI： 10.16016/j.2097-0927.202205141.

摘要

目的

转录组学测序分析WNK2在卵巢癌中调控的生物、代谢过程。

方法

在卵巢癌细胞SKOV3中

使用siRNA干扰WNK2

分为siNC和siWNK2 2组

每组设置3个重复。siRNA转染SKOV3细胞48 h后收集细胞提取总RNA。验证SKOV3细胞中的WNK2成功敲降后

将总RNA样品进行转录组学测序及生物信息分析

筛选padj < 0.01

fold change>1

FPKM>500范围内的基因

即siNC与siWNK2两组间的差异基因

从而获取在SKOV3细胞中被WNK2调控的基因。进一步通过DisGeNET和DO (disease ontology)富集分析得出差异基因与疾病间的关系; GO功能富集分析差异基因调控的细胞功能和代谢过程。通过RT-qPCR验证活性氧(reactive oxygen species

ROS)通路中10个差异基因表达变化是否与测序结果一致

并探究过表达、敲降WNK2后对卵巢癌细胞ROS产生的影响。

结果

DisGeNET以及DO富集分析表明WNK2敲降后

siNC和siWNK2两组的差异基因与多种恶性肿瘤密切相关(

0.05)。GO功能富集分析表明差异基因富集于ROS代谢过程、凋亡信号通路、泛素连接酶复合物形成、线粒体膜蛋白复合物形成、生长因子结合和转录激活因子活性等生物过程(

0.05)。在所有富集通路中

ROS代谢相关的生物过程数量最多且差异最为显著(

0.001)。RT-qPCR验证WNK2敲降后ROS通路中9个差异基因表达变化与测序结果一致

1个基因(PDGFB)与测序结果相反

1个基因在两组中无明显变化。在卵巢癌细胞SKOV3和CAOV3中敲降WNK2后

ROS产生下降; 过表达WNK2后

卵巢癌细胞的ROS产生增多。

结论

WNK2可能通过促进ROS代谢过程调控卵巢癌的恶性进程。

Abstract

Objective

To investigate the biological and metabolic roles of WNK2 in regulation of ovarian cancer based on transcriptome sequencing technology.

Methods

Ovarian cancer SKOV3 cells were divided into experimental group and control group

with 3 replicates in each group.The expression of WNK2 was knocked down with transfection of siRNA targeting WNK2 in SKOV3 cells.Then the total RNA was extracted and prepared for transcriptome sequencing and bioinformatics analysis after 48 h of transfection.The differential genes between siNC and siWNK2 were screened out in the range of padj < 0.01

fold change >1

and FPKM >500 to obtain the genes under the regulation of WNK2.DisGeNET and disease ontology (DO) enrichment analysis was used to analyze the relationship of the diseases and differentially expressed genes (DEGs).GO enrichment analysis was adopted to analyze the cellular functions and metabolic processes in which the DEGs were enriched.Finally

RT-qPCR was used to examine whether the 10 differential proteins in the reactive oxygen species (ROS) pathway were regulated by WNK2.The influence of WNK2 overexpression and knockdown on the ROS metabolic process of ovarian cancer cells was determined.

Results

The DisGeNET and DO enrichment analysis showed the obtained DEGs were closely related with malignant tumors (

0.05).GO functional enrichment analysis showed the obtained DEGs were enriched in biological processes such as ROS metabolism

apoptosis signaling pathway

formation of ubiquitin ligase complex and mitochondrial membrane protein complex

growth factor binding

and transcription activator activity and so on (

0.05).Among these enriched pathways

ROS related pathways were the most numerous and significant (

0.001).The results of RT-qPCR indicated that the changes in 9 DEGs were consistent with the sequencing results after WNK2 was knocked down

and one gene (PDGFB) was contrary to the sequencing results

and one gene had no significant change between the 2 groups.Knockdown of WNK2 in SKOV3 and CAOV3 cells resulted in decreased production of ROS

whereas overexpression of WNK2 increased the production.

Conclusion

WNK2 probably promotes ovarian cancer malignancy by inducing ROS metabolism.

关键词

Keywords

references

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