南方医科大学学报

南方医科大学学报 ›› 2024, Vol. 44 ›› Issue (10): 1947-1954.doi: 10.12122/j.issn.1673-4254.2024.10.13

• • 上一篇    

顺铂诱导TNF-α自分泌引发头颈部鳞状癌细胞RIP1/RIP3/MLKL的坏死性凋亡

汪虹晓1(), 陶德韬2,3(), 马俊杰1, 张东林1,3, 沈左媛1, 邓超1,3(), 周静萍1,3()   

  1. 1.皖南医学院口腔医学院，安徽 芜湖 241002
    2.皖南医学院第一附属医院（皖南医学院弋矶山医院）口腔颌面外科，安徽 芜湖 241100
    3.安徽省口腔材料与应用转化工程研究中心，安徽 合肥 230031
  • 收稿日期:2024-07-25 出版日期:2024-10-20 发布日期:2024-10-31
  • 通讯作者: 邓超,周静萍 E-mail:1492673419@qq.com;taodetao@wnmc.edu.cn;20120015@wnmc.edu.cn;19950008@wnmc.edu.cn
  • 作者简介:汪虹晓，本科，E-mail:1492673419@qq.com
    陶德韬，硕士，副教授，副主任医师，硕士生导师，E-mail: taodetao@wnmc.edu.cn
    第一联系人：汪虹晓、陶德韬共同为第一作者
  • 基金资助:
    安徽省高等学校科学研究项目(2024AH040246);皖南医学院弋矶山医院引进人才基金项目(YR202108);芜湖市科技计划项目(2022JC30);安徽省大学生创新创业训练项目计划(S202210368003)

Cisplatin promotes TNF‑α autocrine to trigger RIP1/RIP3/MLKL-dependent necroptosis of human head and neck squamous cell carcinoma cells

Hongxiao WANG1(), Detao TAO2,3(), Junjie MA1, Donglin ZHANG1,3, Zuoyuan SHEN1, Chao DENG1,3(), Jingping ZHOU1,3()   

  1. 1.School of Stomatology, Wannan Medical College, Wuhu 241002, China
    2.Department of Oral and Maxillofacial Surgery, First Affiliated Hospital (Yijishan Hospital) of Wannan Medical College, Wuhu 241100, China
    3.Anhui Engineering Research Center for Oral Materials and Application, Hefei 230031, China
  • Received:2024-07-25 Online:2024-10-20 Published:2024-10-31
  • Contact: Chao DENG, Jingping ZHOU E-mail:1492673419@qq.com;taodetao@wnmc.edu.cn;20120015@wnmc.edu.cn;19950008@wnmc.edu.cn

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摘要：

目的 探讨顺铂是否能够诱导头颈部鳞状癌细胞产生TNF-α，进而激活RIP1/RIP3/MLKL依赖性的坏死性凋亡通路，抑制鳞癌细胞的增殖，并探讨其分子机制。 方法 选取头颈部鳞状癌细胞系HN4和SCC4作为实验对象，分为对照组、顺铂组、caspases抑制组、坏死性凋亡抑制组。利用CCK-8法检测顺铂刺激后24 h的细胞存活率。随后采用Western blotting检测caspase-8以及坏死性凋亡通路蛋白（RIP1/RIP3/MLKL）和NF-κB（p65）、TNF-α表达情况；结合细胞划痕实验和Western blotting检测上皮间充质转化相关蛋白（N-cadherin、Vimentin、E-cadherin）的表达情况，评估坏死性凋亡对头颈部鳞状癌细胞迁移能力的影响。 结果 顺铂对HN4、SCC4细胞毒性IC50分别约为10 μg/mL、15 μg/mL。在顺铂刺激下，与坏死性凋亡抑制剂组相比，caspase-8表达降低（P<0.05），N-cadherin、Vimentin表达降低（P<0.05）、E-cadherin表达升高（P<0.05），坏死性凋亡通路蛋白（RIP1/RIP3/MLKL）表达升高（P<0.05），TNF-α和NF-κB（p65）蛋白表达均升高（P<0.05）。在顺铂组中，细胞愈合率显著降低于对照组和坏死性凋亡抑制剂组。 结论 顺铂作用可激活头颈部鳞状癌细胞NF-κB信号通路，并促进TNF-α自分泌，从而引发鳞癌细胞经由RIP1/RIP3/MLKL通路依赖性的坏死性凋亡反应，并抑制肿瘤细胞性增殖。

关键词: 头颈部鳞状癌细胞, 顺铂, z-VAD-fmk, 坏死性凋亡, Nec-1

Abstract:

Objective To investigate whether cisplatin induces tumor necrosis factor‑α (TNF‑α) secretion in human head and neck squamous cell carcinoma (HNSCC) cells to trigger RIP1/RIP3/MLKL-dependent necroptosis of the cells. Methods HNSCC cell lines HN4 and SCC4 treated with cisplatin (CDDP) or the combined treatment with CDDP and z-VAD-fmk (a caspase inhibitor) or Nec-1 (a necroptosis inhibitor) for 24 h were examined for changes in cell viability using CCK8 assay and expressions of caspase-8 and necroptosis pathway proteins (RIP1/RIP3/MLKL) using Western blotting. The changes in migration of the cells were assessed with cell scratch assay, and the expressions of epithelial-mesenchymal transition (EMT) marker proteins N-cadherin, vimentin, and E-cadherin as well as the expressions of NF‑κB (p65) and TNF‑α were detected with Western blotting. Results The IC50 of cisplatin was 10 μg/mL in HN4 cells and 15 μg/mL in SCC4 cells. Cisplatin treatment significantly decreased the expressions of caspase-8, N-cadherin and vimentin and increased the expressions of E-cadherin, the necroptosis pathway proteins (RIP1/RIP3/MLKL), TNF‑α, and NF‑κB (p65), and these changes were obviously inhibited by treatment with Nec-1. Cisplatin stimulation also significantly lowered migration of the cells, and this inhibitory effect was strongly attenuated by Nec-1 treatment. Conclusion Cisplatin activates nuclear factor‑κB signaling in HNSCCs to promote TNF‑α autocrine and induce RIP1/RIP3/MLKL-dependent necroptosis, thus leading to inhibition of cell proliferation.

Key words: human head and neck squamous cell carcinoma, cisplatin, z-VAD-fmk, necroptosis, Nec-1