Induction of Mesenchymal-Epithelial Transitions in Sarcoma Cells

Abstract

Phenotypic plasticity refers to a phenomenon in which cells transiently gain traits of another lineage. During carcinoma progression, phenotypic plasticity drives invasion, dissemination and metastasis. Indeed, while most of the studies of phenotypic plasticity have been in the context of epithelial-derived carcinomas, it turns out sarcomas, which are mesenchymal in origin, also exhibit phenotypic plasticity, with a subset of sarcomas undergoing a phenomenon that resembles a mesenchymal-epithelial transition (MET). Here, we developed a method comprising the miR-200 family and grainyhead-like 2 (GRHL2) to mimic this MET-like phenomenon observed in sarcoma patient samples.We sequentially express GRHL2 and the miR-200 family using cell transduction and transfection, respectively, to better understand the molecular underpinnings of these phenotypic transitions in sarcoma cells. Sarcoma cells expressing miR-200s and GRHL2 demonstrated enhanced epithelial characteristics in cell morphology and alteration of epithelial and mesenchymal biomarkers. Future studies using these methods can be used to better understand the phenotypic consequences of MET-like processes on sarcoma cells, such as migration, invasion, metastatic propensity, and therapy resistance.

In-Cell Double Electron–Electron Resonance at Nanomolar Protein Concentrations

“…Interestingly, all four tested mammalian cell lines revealed Mn-dominated echo-detected field swept (EDFS) spectra with comparable intensity per cell (Figure S7, Table S3). However, we found that HEK cells are advantageous for the electroporation experiments because of easy growth and less adherent architecture as compared to other tested lines, which allows for easier handling and shorter recovery time. , …”

“…We show that four different mammalian cell lines exhibit comparable intracellular manganese concentrations; therefore, the achieved sensitivity is also expected for other mammalian cells. However, we found that HEK cells are particularly suited for electroporation experiment because their weak adhesion to the culture plastic dish , facilitates handling of the sample and analysis after shorter recovery times. Notably, suspension HEK cell line derivatives are also available, e.g., HEK293-F, , which could offer further advantages for in-cell EPR.…”

“…However, we found that HEK cells are advantageous for the electroporation experiments because of easy growth and less adherent architecture as compared to other tested lines, which allows for easier handling and shorter recovery time. 41,42 We used an electroporation protocol which was previously used for EPR 9,14,29 but decreased the starting protein concentrations by one order of magnitude (protocol given in SI Section 4). We delivered Alexa647-labeled T4 lysozyme at C start = 5.6 μM or Gd-T4 lysozyme at C start = 11.0 μM into HEK cells.…”

ALDH5A1 acts as a tumour promoter and has a prognostic impact in papillary thyroid carcinoma

“…Epithelial‐mesenchymal transition (EMT) is a process in which epithelial cells acquire mesenchymal features 20 . In cancer, EMT is associated with tumour initiation, invasion, metastasis 21‐23 . Many studies had shown that EMT was associated with invasion and metastasis of thyroid cancer and lateral cervical lymph node metastasis 24‐26 .…”

Functional Role of Non-Coding RNAs during Epithelial-To-Mesenchymal Transition

“…Therefore, regulation of EMT is a complex process that is tightly regulated in different biological events. Epithelial-to-mesenchymal transition is a reversible process, and therefore in certain circumstances mesenchymal-to-epithelial transition (MET) can also take place, particularly during oncogenic processes [ 3 , 4 , 5 ]. Regulatory mechanisms driving MET fall beyond the scope of the present review (see for excellent reviews on MET [ 6 , 7 , 8 ]).…”