Paullin Trillitye, Powell Chase, Menzie Christopher, Hill Robert, Cheng Feng, Martyniuk Christopher J, Westerheide Sandy D
Department of Cell Biology, Microbiology, and Molecular Biology, College of Arts and Sciences, University of South Florida, Tampa, Florida, United States of America.
Department of Pharmaceutical Sciences, College of Pharmacy, University of South Florida, Tampa, Florida, United States of America.
PLoS One. 2017 Aug 9;12(8):e0182930. doi: 10.1371/journal.pone.0182930. eCollection 2017.
Ovarian cancer is the most lethal gynecological cancer, with over 200,000 women diagnosed each year and over half of those cases leading to death. These poor statistics are related to a lack of early symptoms and inadequate screening techniques. This results in the cancer going undetected until later stages when the tumor has metastasized through a process that requires the epithelial to mesenchymal transition (EMT). In lieu of traditional monolayer cell culture, EMT and cancer progression in general is best characterized through the use of 3D spheroid models. In this study, we examine gene expression changes through microarray analysis in spheroid versus monolayer ovarian cancer cells treated with TGFβ to induce EMT. Transcripts that included Coiled-Coil Domain Containing 80 (CCDC80), Solute Carrier Family 6 (Neutral Amino Acid Transporter), Member 15 (SLC6A15), Semaphorin 3E (SEMA3E) and PIF1 5'-To-3' DNA Helicase (PIF1) were downregulated more than 10-fold in the 3D cells while Inhibitor Of DNA Binding 2, HLH Protein (ID2), Regulator Of Cell Cycle (RGCC), Protease, Serine 35 (PRSS35), and Aldo-Keto Reductase Family 1, Member C1 (AKR1C1) were increased more than 50-fold. Interestingly, EMT factors, stress responses and epigenetic processes were significantly affected by 3D growth. The heat shock response and the oxidative stress response were also identified as transcriptome responses that showed significant changes upon 3D growth. Subnetwork enrichment analysis revealed that DNA integrity (e.g. DNA damage, genetic instability, nucleotide excision repair, and the DNA damage checkpoint pathway) were altered in the 3D spheroid model. In addition, two epigenetic processes, DNA methylation and histone acetylation, were increased with 3D growth. These findings support the hypothesis that three dimensional ovarian cell culturing is physiologically different from its monolayer counterpart.
卵巢癌是最致命的妇科癌症，每年有超过20万名女性被诊断出患有此病，其中超过一半的病例会导致死亡。这些糟糕的数据与缺乏早期症状以及筛查技术不足有关。这导致癌症在肿瘤通过上皮-间质转化（EMT）过程发生转移的后期阶段才被发现。代替传统的单层细胞培养，EMT和癌症进展总体上通过使用3D球体模型能得到最佳表征。在本研究中，我们通过微阵列分析检测了用TGFβ处理以诱导EMT的球体与单层卵巢癌细胞中的基因表达变化。包括卷曲螺旋结构域包含蛋白80（CCDC80）、溶质载体家族6（中性氨基酸转运体）成员15（SLC6A15）、信号素3E（SEMA3E）和PIF1 5'-至-3' DNA解旋酶（PIF1）的转录本在3D细胞中下调超过10倍，而DNA结合抑制因子2、HLH蛋白（ID2）、细胞周期调节因子（RGCC）、丝氨酸蛋白酶35（PRSS35）和醛酮还原酶家族1成员C1（AKR1C1）则增加超过50倍。有趣的是，EMT因子、应激反应和表观遗传过程受到3D生长的显著影响。热休克反应和氧化应激反应也被确定为在3D生长时表现出显著变化的转录组反应。子网富集分析显示，DNA完整性（如DNA损伤、基因不稳定、核苷酸切除修复和DNA损伤检查点途径）在3D球体模型中发生了改变。此外，随着3D生长，DNA甲基化和组蛋白乙酰化这两个表观遗传过程增加。这些发现支持了三维卵巢细胞培养在生理上与其单层对应物不同的假设。
