中国组织工程研究

中国组织工程研究 ›› 2026, Vol. 30 ›› Issue (19): 4902-4910.doi: 10.12307/2026.247

• 肿瘤干细胞 cancer stem cells • 上一篇    下一篇

乳酸化混合谱系激酶结构域样蛋白对乳腺肿瘤干细胞干性表达的影响

吴  芳1，陶  翔2，何文莹1，谢季欣1，廖海雷1，王立斌2，王丽娟1   

  1. 1宁夏医科大学基础医学院，宁夏回族自治区银川市   750004；2深圳市南山区人民医院，广东省深圳市   518000
  • 收稿日期:2025-09-02 接受日期:2025-11-29 出版日期:2026-07-08 发布日期:2026-02-14
  • 通讯作者: 王丽娟，博士，教授，宁夏医科大学基础医学院，宁夏回族自治区银川市 750004； 共同通讯作者：王立斌，博士，教授，深圳市南山区人民医院，广东省深圳市 518000
  • 作者简介:吴芳，女，2000年生，四川省宜宾市人，汉族，宁夏医科大学在读硕士，主要从事肿瘤细胞生物学相关研究。
  • 基金资助:
    宁夏自然科学基金项目(2024AAC03197)，项目负责人：王丽娟；国家自然科学基金项目(NSFC32160094)，项目负责人：王丽娟；国家自然科学基金项目(82260610)，项目负责人：王立斌；宁夏自然科学基金重点项目(2023AAC02059)，项目负责人：王立斌；深圳市南山区卫生健康系统科技重大项目(NSZD2024021，NSZD2025002)，项目负责人：王立斌

Effect of lactylated mixed lineage kinase domain-like protein on stemness expression of breast tumor stem cells

Wu Fang1, Tao Xiang2, He Wenying1, Xie Jixin1, Liao Hailei1, Wang Libin2, Wang Lijuan1   

  1. 1Basic Medical College, Ningxia Medical University, Yinchuan 750004, Ningxia Hui Autonomous Region, China; 2Nanshan District People's Hospital of Shenzhen City, Shenzhen 518000, Guangdong Province, China
  • Received:2025-09-02 Accepted:2025-11-29 Online:2026-07-08 Published:2026-02-14
  • Contact: Wang Lijuan, MD, Professor, Basic Medical College, Ningxia Medical University, Yinchuan 750004, Ningxia Hui Autonomous Region, China; Wang Libin, MD, Professor, Nanshan District People's Hospital of Shenzhen City, Shenzhen 518000, Guangdong Province, China
  • About author:Wu Fang, Master candidate, Basic Medical College, Ningxia Medical University, Yinchuan 750004, Ningxia Hui Autonomous Region, China
  • Supported by:
    Ningxia Natural Science Foundation, No. 2024AAC03197 (to WLJ); National Natural Science Foundation of China, No. NSFC32160094 (to WLJ); National Natural Science Foundation of China, No. 82260610 (to WLB); Ningxia Natural Science Foundation (Key Project), No. 2023AAC02059 (to WLB); Shenzhen Nanshan District Health System Science and Technology Major Project, No. NSZD2024021, NSZD2025002 (to WLB) 

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摘要：

文题释义：

混合谱系激酶结构域样蛋白(mixed lineage kinase domain-like protein，MLKL)：属于蛋白激酶超家族，是具有无催化活性激酶结构域的假激酶。受体相互作用丝氨酸苏氨酸激酶3磷酸化后激活混合谱系激酶结构域样蛋白，使它定位于质膜并执行以钙流入和质膜损伤为特征的坏死性凋亡。
蛋白质乳酸化修饰：是乳酸在辅酶A作用下形成乳酰辅酶A，然后在一系列酶的催化下将其乳酰基团结合到目的蛋白赖氨酸残基上的过程。

摘要
背景：混合谱系激酶结构域样蛋白是坏死性凋亡途径中关键执行蛋白之一，在多种疾病中发挥重要作用，但其乳酸化对乳腺肿瘤干细胞形成和分化的作用机制尚不清楚。
目的：探究混合谱系激酶结构域样蛋白第230位赖氨酸(K230)乳酸化对乳腺肿瘤干细胞干性表达的影响。
方法：质谱分析乳腺肿瘤MCF-7贴壁细胞和成球干细胞中蛋白乳酸化程度差异，筛选获得与肿瘤干细胞相关的混合谱系激酶结构域样蛋白及其乳酸化位点。构建混合谱系激酶结构域样蛋白K230R突变质粒载体，转染乳腺肿瘤MCF-7细胞，CCK-8和划痕实验检测细胞增殖和迁移能力；细胞悬浮成球实验验证混合谱系激酶结构域样蛋白K230R突变对乳腺肿瘤干细胞形成的影响；蛋白质免疫印迹检测肿瘤干细胞干性及上皮-间充质转化相关蛋白表达。
结果与结论：乳腺肿瘤干细胞中混合谱系激酶结构域样蛋白K230位乳酸化修饰程度较高；与野生型组相比，K230R突变组细胞划痕愈合率及干细胞成球率显著降低，上皮相关蛋白表达显著升高，间质相关蛋白和干性相关蛋白表达显著降低。研究表明：混合谱系激酶结构域样蛋白K230位点乳酸化促进乳腺肿瘤细胞上皮-间充质转化，增强乳腺肿瘤干细胞干性表达，从而促进乳腺肿瘤的发生和发展。

关键词: 乳腺肿瘤, 乳腺肿瘤干细胞, 混合谱系激酶结构域样蛋白, 乳酸化

Abstract: BACKGROUND: Mixed lineage kinase domain-like protein is one of the key executor proteins in the necroptosis pathway and plays an important role in various diseases. However, the mechanism by which its lactylation affects the formation and differentiation of breast tumor stem cells remains unclear.
OBJECTIVE: To investigate the effect of mixed lineage kinase domain-like protein K230 site lactylation on the stemness expression of breast tumor stem cells.
METHODS: The differences in protein lactylation between breast tumor MCF-7 adherent cells and spheroidal stem cells were analyzed by mass spectrometry. The mixed lineage kinase domain-like protein and its lactylation sites related to tumor stem cells were screened. A mixed lineage kinase domain-like protein K230R mutant plasmid vector was constructed and transfected into MCF-7 breast tumor cells. The proliferation and migration abilities of the cells were detected by CCK-8 and scratch assays. The effect of mixed lineage kinase domain-like protein K230R mutation on the formation of breast tumor stem cells was verified by cell suspension spheroid formation experiments. The expression of tumor stem cell-related and epithelial-mesenchymal transition-related proteins was detected by western blot assay.
RESULTS AND CONCLUSION: Mixed lineage kinase domain-like protein K230 is highly lactylated in breast cancer stem cells. Compared with wild-type cells, K230R mutant cells showed significantly reduced wound healing and stem cell sphere formation rates, significantly increased expression of epithelial-associated proteins, and significantly decreased expression of interstitial-associated proteins and stemness-associated proteins. It is indicated that lactic acidification at the K230 site of mixed lineage kinase domain-like protein promotes epithelial-mesenchymal transition, enhances the formation of breast tumor stem cells, and thereby contributes to the occurrence and development of breast tumors.

Key words: breast tumor, breast tumor stem cells, mixed lineage kinase domain-like protein, lactylation

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