自噬抑制剂Spautin-1增加人胰腺癌细胞系PaTu 8988t凋亡率

doi:10.16352/j.issn.1001-6325.2026.07.0932

摘要/Abstract

摘要： 目的探究泛素特异性肽酶10(USP10)抑制剂Spautin-1对人胰腺癌细胞系PaTu 8988t增殖、迁移、自噬及凋亡的影响及其机制。方法使用GEPIA 2在线工具,对源自加州大学圣克鲁兹分校功能基因组学浏览器(UCSC Xena)的癌症基因组图谱(TCGA)与基因型-组织表达项目(GTEx)的胰腺导管腺癌(PAAD)标准化整合数据进行分析,评估USP10的表达、预后价值及与自噬基因(MAP1LC3B和SQSTM1)的相关性;使用R语言分析GEO数据集进行验证和可视化。以PaTu 8988t细胞为对象,CCK8法检测Spautin-1(0~200 μmol/L)细胞毒性并选定20 μmol/L进行后续实验。划痕实验和Transwell小室法评估迁移和侵袭能力;流式细胞测量术(annexin V/PI双染法)检测凋亡,DCFH-DA探针检测活性氧(ROS);Western blot分析USP10、LC3-Ⅱ、p62蛋白表达。结果 1)GEPIA 2分析显示USP10在PAAD中表达上调,提示不良预后,且与自噬基因MAP1LC3B、SQSTM1表达正相关;GSE16515数据验证其表达趋势,聚类分析提示USP10可能参与自噬调控;2)Spautin-1浓度依赖性抑制细胞活力;3)20 μmol/L Spautin-1显著抑制划痕愈合率和Transwell穿膜细胞数(P＜0.05);4)Spautin-1处理下调LC3-Ⅱ、上调p62蛋白,并降低USP10蛋白(P＜0.05);5)Spautin-1显著升高细胞内ROS水平并增加细胞凋亡率(P＜0.05)。结论 Spautin-1有效抑制PaTu 8988t细胞的增殖和迁移侵袭能力,其作用伴随USP10蛋白下调、自噬流受阻、ROS积累及凋亡增加。

关键词: 胰腺癌, 泛素特异性蛋白酶10, 自噬抑制剂, 凋亡, 活性氧

Abstract: Objective To investigate the effects of Spautin-1, an inhibitor of ubiquitin-specific peptidase 10 (USP10), on the proliferation, migration, autophagy, and apoptosis of human pancreatic cancer cell line PaTu 8988t, and to explore the underlying mechanisms. Methods Analysis of standardized TCGA/GTEx pancreatic ductal adenocarcinoma (PAAD) data from UCSC Xena was performed with GEPIA 2 to assess USP10 expression, its prognostic value and its potential correlation with autophagy genes (MAP1LC3B and SQSTM1). Validation and visualization were conducted with R language on the GEO dataset. PaTu 8988t cells were incubated with Spautin-1 (0~200 μmol/L). Cytotoxicity was determined by CCK-8 assay, and 20 μmol/L was selected for subsequent experiments. Cell migration and invasion were evaluated by wound healing and Transwell assays, respectively. Apoptosis was detected by flow cytometry (annexin V-FITC/PI double staining). Intracellular reactive oxygen species (ROS) level was measured with the DCFH-DA probe. Protein expression of USP10, LC3-Ⅱ and p62 was measured by Western blot. All experiments were independently repeated at least three times. Results 1)GEPIA 2 USP10 was up-regulated in PAAD accompanied by a poor prognosis/The expression was positively correlated with the autophagy genes MAP1LC3B and SQSTM1. Validation with GSE16515 dataset confirmed this expression trend, and clustering analysis suggested USP10's potential involvement in autophagy regulation. 2)Spautin-1 dose-dependently inhibited cell viability. 3)Spautin-1(20 μmol/L) significantly suppressed scratch healing rates and Transwell invasion (P＜0.05). 4)Spautin-1 down regulated LC3-Ⅱ and USP10 protein levels while up-regulating p62, indicating impaired autophagy flux(P＜0.05). 5)Spautin-1 significantly increased intracellular ROS level and apoptosis rates(P＜0.05). Conclusions Spautin-1 effectively inhibits proliferation and migration/invasion of PaTu 8988t cells. Its effects are accompanied by a reduced USP10 expression, blocked autophagy flux, ROS accumulation and increased apoptosis which suggest a potential mechanism involving USP10 inhibition, autophagy disruption and ROS-dependent apoptosis.

Key words: pancreatic cancer, ubiquitin-specific peptidase 10, autophagy inhibitor, apoptosis, reactive oxygen species

中图分类号:

R735.9

董泽耀, 林淑瑜, 戴立. 自噬抑制剂Spautin-1增加人胰腺癌细胞系PaTu 8988t凋亡率[J]. 基础医学与临床, 2026, 46(7): 932-938.

DONG Zeyao, LIN Shuyu, DAI Li. Autophagy inhibitor Spautin-1 increases apoptosis rate in human pancreatic cancer cell line PaTu 8988t[J]. Basic & Clinical Medicine, 2026, 46(7): 932-938.

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