Translation, but not transfection limits clinically relevant, exogenous mRNA based induction of alpha-4 integrin expression on human mesenchymal stem cells

Nowakowski, Adam; Andrzejewska, A; Boltze, Johannes; Nitzsche, Franziska; Cui, Li Li; Jolkkonen, Jukka; Walczak, Piotr; Łukomska, Barbara; Janowski, Mirosław

doi:10.1038/s41598-017-01304-3

Cited by 23 publications

(24 citation statements)

References 52 publications

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“…To date, EMT/MET is routinely studied by the use of exogenous reporter genes that are heterologously expressed, exhibiting interference of cisand trans-regulatory elements, alternative promoters, or epigenetic events, which result in obtaining unreliable data [32,33]. These limitations are overcome by using genome engineering of endogenous genes.…”

Section: Discussionmentioning

confidence: 99%

Genetically Engineered Lung Cancer Cells for Analyzing Epithelial–Mesenchymal Transition

Kiełbus

Czapiński

Kałafut

et al. 2019

Cells

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Cell plasticity, defined as the ability to undergo phenotypical transformation in a reversible manner, is a physiological process that also exerts important roles in disease progression. Two forms of cellular plasticity are epithelial–mesenchymal transition (EMT) and its inverse process, mesenchymal–epithelial transition (MET). These processes have been correlated to the poor outcome of different types of neoplasias as well as drug resistance development. Since EMT/MET are transitional processes, we generated and validated a reporter cell line. Specifically, a far-red fluorescent protein was knocked-in in-frame with the mesenchymal gene marker VIMENTIN (VIM) in H2170 lung cancer cells. The vimentin reporter cells (VRCs) are a reliable model for studying EMT and MET showing cellular plasticity upon a series of stimulations. These cells are a robust platform to dissect the molecular mechanisms of these processes, and for drug discovery in vitro and in vivo in the future.

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Section: Discussionmentioning

confidence: 99%

Genetically Engineered Lung Cancer Cells for Analyzing Epithelial–Mesenchymal Transition

Kiełbus

Czapiński

Kałafut

et al. 2019

Cells

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show abstract

“…To-date EMT/MET is routinely studied by the use of exogenous reporter genes that are heterologously expressed exhibiting interference of cis- and trans-regulatory elements, alternative promoters or epigenetic events that resulted in obtaining unreliable data [32,33]. These limitations are overcome by using genome engineering of endogenous genes.…”

Section: Discussionmentioning

confidence: 99%

Genetical engineered lung cancer cell for analyzing Epithelial-Mesenchymal transition

Kiełbus

Czapiński

Kałafut

et al. 2019

Preprint

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Cell plasticity, defined as the ability to undergo phenotypical transformation in a reversible 12 manner, is a physiological processes that also exert important roles in disease progression Two forms 13 of cellular plasticity are epithelial-mesenchymal transition (EMT) and its inverse process, 14 mesenchymal-epithelial transition (MET). These processes have been correlated to the poor outcome 15 of different types of neoplasias as well as drug resistance development. Since EMT/MET are 16 transitional processes, we have generated and validated a reporter cell line. Specifically, a far-red 17 fluorescent protein was knocked-in in-frame with the mesenchymal gene marker VIMENTIN (VIM) 18 in H2170 lung cancer cells. The vimentin reporter cells (VRCs) are a reliable model for studying EMT 19 and MET showing cellular plasticity upon a series of stimulations. These cells are a robust platform 20 to dissect the molecular mechanisms of these processes, and for drug discovery in vitro and in the 21 future in vivo.22 Keywords: epithelial-mesenchymal transition (EMT); mesenchymal-epithelial transition (MET); 23 cancer cell line; vimentin; reporter; 24 25 29 transition (EMT) and mesenchymal-epithelial transition (MET), respectively. EMT has been 30 correlated to the metastasis and invasion potential of many types of malignancies [2]. The fact that 31 the poor outcome of many types of neoplasias correlates with EMT/MET, makes these molecular 32 phenomena important focus for research and drug targeting [3-6]. Despite the clinical association,33 the role of EMT/MET in metastasis is inconclusive, for example mesenchymal-like prostate cancer 34 cells survive in circulation but, unlike epithelial or cells undergoing EMT, they are unable to form 35 macrometastases [7]. In fact, breast cancer cell metastasis to lung tissue in mice was not affected by36 decreasing EMT -by targeting EMT-triggering transcription factors such as SNAI1 (Snail) and SNAI2 37 (Slug) by overexpressing miR-200 [8]. An additional role of EMT in tumorigenesis might be the 38 development of apoptotic tolerance and increased resistance to chemotherapy as has been found in 39animal models and patients [8,9]. Recently, a hybrid stage between epithelial and mesenchymal 40 2 of 19 phenotypes (hybE/M) has been recognized, and such hybE/M cells migrate outside the primary 41 tumors displaying some mesenchymal features such as spindle-like morphology, increased nuclear 42 levels of ZEB1 transcription factor, and epithelial ones such as cell-cell adhesion potential [10,11]. 43In order to study the transitory stages of EMT, MET and hybE/M, in particular at in vivo settings, 44 there is need to generate reporter cells to visualize phenotypical changes. There have been different 45 approaches to this, for example the use of heterologous fluorescent proteins driven under stage-46 specific promoters such as mesenchymal (ZEB1) and epithelial (CDH1) [12], mesenchymal snai1 and 47 epithelial sox10 in zebrafish [13], or mesenchymal Vimentin (Vim) in mice [14]. Nevertheless, 48 hete...

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“…IVT mRNA-based modulation of homing MSCs to target ischemic areas has been assessed by in vitro transfection of different surface molecules, including the chemokine receptor CXCR4 [253], and the integrin α4 (ITGA4) [254]. MSCs may be also used as a vehicle to target biological molecules, such as interleukins (IL), to specific tissues.…”

Section: Regenerative Medicine and Cell Engineeringmentioning

confidence: 99%

Nanomedicines to Deliver mRNA: State of the Art and Future Perspectives

et al. 2020

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The use of messenger RNA (mRNA) in gene therapy is increasing in recent years, due to its unique features compared to plasmid DNA: Transient expression, no need to enter into the nucleus and no risk of insertional mutagenesis. Nevertheless, the clinical application of mRNA as a therapeutic tool is limited by its instability and ability to activate immune responses; hence, mRNA chemical modifications together with the design of suitable vehicles result essential. This manuscript includes a revision of the strategies employed to enhance in vitro transcribed (IVT) mRNA functionality and efficacy, including the optimization of its stability and translational efficiency, as well as the regulation of its immunostimulatory properties. An overview of the nanosystems designed to protect the mRNA and to overcome the intra and extracellular barriers for successful delivery is also included. Finally, the present and future applications of mRNA nanomedicines for immunization against infectious diseases and cancer, protein replacement, gene editing, and regenerative medicine are highlighted.

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Translation, but not transfection limits clinically relevant, exogenous mRNA based induction of alpha-4 integrin expression on human mesenchymal stem cells

Cited by 23 publications

References 52 publications

Genetically Engineered Lung Cancer Cells for Analyzing Epithelial–Mesenchymal Transition

Genetically Engineered Lung Cancer Cells for Analyzing Epithelial–Mesenchymal Transition

Genetical engineered lung cancer cell for analyzing Epithelial-Mesenchymal transition

Nanomedicines to Deliver mRNA: State of the Art and Future Perspectives

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