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© 2022. This work is published under http://creativecommons.org/licenses/by/4.0/ (the “License”). Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License.

Abstract

Liposomes can efficiently deliver messenger RNA (mRNA) into cells. When mRNA cocktails encoding different proteins are needed, a considerable challenge is to efficiently deliver all mRNAs into the cytosol of each individual cell. In this work, two methods are explored to co‐deliver varying ratiometric doses of mRNA encoding red (R) or green (G) fluorescent proteins and it is found that packaging mRNAs into the same lipoplexes (mingle‐lipoplexes) is crucial to efficiently deliver multiple mRNA types into the cytosol of individual cells according to the pre‐defined ratio. A mixture of lipoplexes containing only one mRNA type (single‐lipoplexes), however, seem to follow the “first come – first serve” principle, resulting in a large variation of R/G uptake and expression levels for individual cells leading to ratiometric dosing only on the population level, but rarely on the single‐cell level. These experimental observations are quantitatively explained by a theoretical framework based on the stochasticity of mRNA uptake in cells and endosomal escape of mingle‐ and single‐lipoplexes, respectively. Furthermore, the findings are confirmed in 3D retinal organoids and zebrafish embryos, where mingle‐lipoplexes outperformed single‐lipoplexes to reliably bring both mRNA types into single cells. This benefits applications that require a strict control of protein expression in individual cells.

Details

Title
Together is Better: mRNA Co‐Encapsulation in Lipoplexes is Required to Obtain Ratiometric Co‐Delivery and Protein Expression on the Single Cell Level
Author
Zhang, Heyang 1   VIAFID ORCID Logo  ; Bussmann, Jeroen 2 ; Huhnke, Florian H 3 ; Devoldere, Joke 1 ; An‐Katrien Minnaert 1 ; Jiskoot, Wim 2 ; Serwane, Friedhelm 4 ; Spatz, Joachim 5 ; Röding, Magnus 6 ; De Smedt, Stefaan C 7 ; Braeckmans, Kevin 8 ; Remaut, Katrien 7   VIAFID ORCID Logo 

 Laboratory of General Biochemistry and Physical Pharmacy, Faculty of Pharmaceutical Sciences, Ghent University, Ghent, Belgium 
 Division of BioTherapeutics, Leiden Academic Center for Drug Research, Leiden University, Leiden, The Netherlands 
 Max Planck Institute for Medical Research, Department of Cellular Biophysics, Stuttgart, Germany 
 Max Planck Institute for Medical Research, Department of Cellular Biophysics, Stuttgart, Germany; Center for NanoScience, Ludwig‐Maximilian‐University Munich, Munich, Germany; Faculty of Physics, Ludwig‐Maximilian‐University, Munich, Germany; Munich Cluster for Systems Neurology (SyNergy), Munich, Germany 
 Max Planck Institute for Medical Research, Department of Cellular Biophysics, Stuttgart, Germany; Department of Biophysical Chemistry, University of Heidelberg, Heidelberg, Germany 
 RISE Research Institutes of Sweden, Bioeconomy and Health, Agriculture and Food, Göteborg, Sweden; Department of Mathematical Sciences, Chalmers University of Technology and University of Gothenburg, Göteborg, Sweden 
 Laboratory of General Biochemistry and Physical Pharmacy, Faculty of Pharmaceutical Sciences, Ghent University, Ghent, Belgium; Cancer Research Institute Ghent (CRIG), Ghent University, Ghent, Belgium 
 Laboratory of General Biochemistry and Physical Pharmacy, Faculty of Pharmaceutical Sciences, Ghent University, Ghent, Belgium; Center for Advanced Light Microscopy, Ghent University, Ghent, Belgium 
Section
Research Articles
Publication year
2022
Publication date
Feb 2022
Publisher
John Wiley & Sons, Inc.
e-ISSN
21983844
Source type
Scholarly Journal
Language of publication
English
ProQuest document ID
2624873676
Copyright
© 2022. This work is published under http://creativecommons.org/licenses/by/4.0/ (the “License”). Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License.