Sosnowska Malwina, Kutwin Marta, Zawadzka Katarzyna, Pruchniewski Michał, Strojny Barbara, Bujalska Zuzanna, Wierzbicki Mateusz, Jaworski Sławomir, Sawosz Ewa
Department of Nanobiotechnology, Institute of Biology, Warsaw University of Life Sciences, 02-776 Warsaw, Poland.
Cancers (Basel). 2023 Nov 23;15(23):5553. doi: 10.3390/cancers15235553.
The epithelial-mesenchymal transition (EMT) is a process in which epithelial cells acquire the ability to actively migrate via a change to the mesenchymal phenotype. This mechanism occurs in an environment rich in cytokines and reactive oxygen species but poor in nutrients. The aim of this study was to demonstrate that the use of a fullerene C nanofilm can inhibit liver cancer cell invasion by restoring their non-aggressive, epithelial phenotype. We employed epithelial and mesenchymal HepG2 and SNU-449 liver cancer cells and non-cancerous mesenchymal HFF2 cells in this work. We used enzyme-linked immunosorbent assays (ELISAs) to determine the content of glutathione and transforming growth factor (TGF) in cells. We measured the total antioxidant capacity with a commercially available kit. We assessed cell invasion based on changes in morphology, the scratch test and the Boyden chamber invasion. In addition, we measured the effect of C nanofilm on restoring the epithelial phenotype at the protein level with protein membranes, Western blotting and mass spectrometry. C nanofilm downregulated TGF and increased glutathione expression in SNU-449 cells. When grown on C nanofilm, invasive cells showed enhanced intercellular connectivity; reduced three-dimensional invasion; and reduced the expression of key invasion markers, namely MMP-1, MMP-9, TIMP-1, TIMP-2 and TIMP-4. Mass spectrometry showed that among the 96 altered proteins in HepG2 cells grown on C nanofilm, 41 proteins are involved in EMT and EMT-modulating processes such as autophagy, inflammation and oxidative stress. The C nanofilm inhibited autophagy, showed antioxidant and anti-inflammatory properties, increased glucose transport and regulated the β-catenin/keratin/Smad4/snail+slug and MMP signalling pathways. In conclusion, the C nanofilm induces a hybrid mesenchymal-epithelial phenotype and could be used in the prevention of postoperative recurrences.
上皮-间质转化（EMT）是一个上皮细胞通过转变为间质表型而获得主动迁移能力的过程。这种机制发生在富含细胞因子和活性氧但营养物质匮乏的环境中。本研究的目的是证明使用富勒烯C纳米膜可以通过恢复其非侵袭性的上皮表型来抑制肝癌细胞的侵袭。在这项工作中，我们使用了上皮和间质的HepG2和SNU-449肝癌细胞以及非癌性间质HFF2细胞。我们使用酶联免疫吸附测定（ELISA）来测定细胞中谷胱甘肽和转化生长因子（TGF）的含量。我们使用市售试剂盒测量总抗氧化能力。我们根据形态变化、划痕试验和博伊登小室侵袭试验评估细胞侵袭。此外，我们通过蛋白质膜、蛋白质免疫印迹和质谱法在蛋白质水平上测量了C纳米膜对恢复上皮表型的影响。C纳米膜下调了SNU-449细胞中的TGF并增加了谷胱甘肽的表达。当在C纳米膜上生长时，侵袭性细胞显示出细胞间连接增强；三维侵袭减少；关键侵袭标志物即基质金属蛋白酶-1（MMP-1）、基质金属蛋白酶-9（MMP-9）、金属蛋白酶组织抑制因子-1（TIMP-1）、金属蛋白酶组织抑制因子-2（TIMP-2）和金属蛋白酶组织抑制因子-4（TIMP-4）的表达降低。质谱分析表明，在C纳米膜上生长的HepG2细胞中96种改变的蛋白质中，有41种蛋白质参与EMT以及自噬、炎症和氧化应激等EMT调节过程。C纳米膜抑制自噬，具有抗氧化和抗炎特性，增加葡萄糖转运并调节β-连环蛋白/角蛋白/Smad4/蜗牛+蛞蝓和MMP信号通路。总之，C纳米膜诱导了一种混合的间质-上皮表型，可用于预防术后复发。
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