| 张天赐,郭 婷,孙秀璇,冯 转,李 勇,耿杰杰,吴 佼.铁死亡相关基因ACSL4、TFRC及Hippo通路关键分子YAP在间皮瘤组织中的表达及意义[J].现代生物医学进展英文版,2021,(9):1617-1622. |
| 铁死亡相关基因ACSL4、TFRC及Hippo通路关键分子YAP在间皮瘤组织中的表达及意义 |
| The Expression and Significance of ACSL4, TFRC and Crucial Mediator of Hippo Signaling YAP in Mesothelioma |
| Received:October 23, 2020 Revised:November 19, 2020 |
| DOI：10.13241/j.cnki.pmb.2021.09.004 |
| 中文关键词: 铁死亡 间皮瘤 组织芯片 ACSL4 TFRC YAP |
| 英文关键词: Ferroptosis Mesothelioma Tissue microarray ACSL4 TFRC YAP |
| 基金项目:国家自然科学基金项目（31401222，31800756） |
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| 中文摘要: |
| 摘要 目的：分析铁死亡相关基因长链酯酰辅酶A合成酶4（acyl-CoA synthetase long chain family member 4，ACSL4）、转铁蛋白受体（transferrin receptor, TFRC）及Hippo通路关键分子Yes相关蛋白（Yes-associated protein，YAP）在间皮瘤组织中的表达水平、相关性及意义，观察YAP活化对间皮瘤细胞系211H侵袭转移能力及铁死亡敏感性的影响。方法：应用组织芯片技术和免疫组化法观察ACSL4、TFRC及YAP在30例间皮瘤组织和10例正常组织中的表达，分析蛋白水平及分布的相关性。GEPIA数据库分析基因ACSL4、TFRC和YAP在间皮瘤组织中的mRNA水平相关性。应用逆转录病毒载体于211H细胞中过表达活化型YAP S127A (serine-127突变为alanine)，利用实时荧光定量PCR检测ACSL4及TFRC表达；利用划痕实验和transwell侵袭实验检测211H细胞迁移及侵袭能力；利用erastin刺激及SYTOX Green染色检测铁死亡敏感性。结果：间皮瘤组织中YAP细胞核表达、ACSL4及TFRC阳性表达率显著高于正常组织，差异均有统计学意义（P<0.001，P<0.001，P<0.0001）。经Spearman秩相关检验，ACSL4及TFRC蛋白在间皮瘤中的表达呈正相关(Spearman r=0.61，P=0.0003)；YAP在细胞核中的表达与ACSL4表达呈正相关(Spearman r=0.4872，P=0.0063)。GEPIA 数据库分析发现，YAP与ACSL4、ACSL4与TFRC在间皮瘤中的mRNA表达水平均呈正相关（Spearman r=0.33, P=0.0019；Spearman r=0.52，P=0.0000）。表达活化型YAP的211H细胞，ACSL4与TFRC mRNA水平与对照细胞相比均上调；迁移侵袭能力增强；铁死亡敏感性增强。结论：YAP、ACSL4及TFRC在间皮瘤组织中表达具有正相关性；YAP活化促进间皮瘤细胞侵袭转移能力，同时通过ACSL4及TFRC提高细胞对铁死亡诱导的敏感性。YAP、ACSL4及TFRC有望成为预测间皮瘤铁死亡敏感性的潜在生物学标志物。 |
| 英文摘要: |
| ABSTRACT Objective: This study was designed to explore the expression and significance of acyl-CoA synthetase long chain family member 4 (ACSL4), transferrin receptor (TFRC) and the crucial mediator of Hippo signaling Yes-associated protein (YAP) in mesothelioma tissues, and to study the regulation of YAP activation in migration, invasion and ferroptosis sensitivity of mesothelioma cells. Methods: Tissue microarray containing specimens from 30 mesothelioma patients and 10 normal mesothelium was used. Immunohistochemistry was applied to examine the expression of YAP, ACSL4 and TFRC, and the correlations between the expression of the proteins were analyzed. The GEPIA database was also applied to explore the expression of YAP, ACSL4 and TFRC. Constitutively active YAP mutant, S127A (serine-127 mutated to alanine) was transfected into mesothelioma 211H cells using retrovirus system, and the ACSL4 and TFRC mRNA levels were examined by real-time PCR. Cells were subjected to an invasion assay and a wound healing assay to examine the invasion and migration potential. Erastin was used to induce ferroptosis and cell death was analyzed by SYTOX Green staining followed by flow cytometry. Results: We found elevated nuclear YAP, ACSL4 and TFRC levels in mesothelioma tissues compared with normal mesothelium (nuclear YAP: P<0.001, ACSL4: P<0.001, TFRC: P<0.0001). ACSL4 was positively correlated with TFRC (Spearman r=0.61, P=0.0003) and nuclear YAP (Spearman r=0.4872, P=0.0063). In GEPIA database, the mRNA level of ACSL4 was positively correlated with YAP（Spearman r=0.33, P=0.0019） and TFRC （Spearman r=0.52, P=0.0000）. YAPS127A mutant increased mRNA levels of ACSL4 and TFRC, enhanced migration and invasion potentials of 211H cells, and sensitized ferroptosis of cells. Conclusion: YAP and ACSL4 showed positive correlation with TFRC expression, in mesothelioma tissues. YAP activation increased migration and invasion of cells, but at the same time sensitized ferroptosis of cells by upregulating expression of ACSL4 and TFRC. YAP, ACSL4 and TFRC might become potential bio-markers for indicating ferroptosis sensitivity of mesothelioma. |
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