Together is Better: mRNA Co-Encapsulation in Lipoplexes is Required to Obtain Ratiometric Co-Delivery and Protein Expression on the Single Cell LevelShow others and affiliations
2022 (English)In: Advanced Science, E-ISSN 2198-3844, Vol. 9, no 4, article id 2102072Article in journal (Refereed) Published
Abstract [en]
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. © 2021 The Authors.
Place, publisher, year, edition, pages
John Wiley and Sons Inc , 2022. Vol. 9, no 4, article id 2102072
Keywords [en]
cellular uptake, mingle/single-mRNA lipoplex, protein expression, single cell, theoretical modeling, Cells, Cytology, Encoding (symbols), Proteins, RNA, Signal encoding, Individual cells, Lipoplexes, Messenger RNA, Mingle/single-messenger RNA lipoplex, Protein expressions, Ratiometric, Single cells, Single-cell level, Liposomes
National Category
Cell and Molecular Biology
Identifiers
URN: urn:nbn:se:ri:diva-57507DOI: 10.1002/advs.202102072Scopus ID: 2-s2.0-85121388407OAI: oai:DiVA.org:ri-57507DiVA, id: diva2:1623698
Note
Funding details: Baden-Württemberg Stiftung, MIVT‐5; Funding details: European Research Council, ERC; Funding details: Deutsche Forschungsgemeinschaft, DFG, EXC‐2082/1 – 390761711; Funding details: Fonds Wetenschappelijk Onderzoek, FWO, 1S28420N; Funding details: China Scholarship Council, CSC; Funding details: Horizon 2020, 648124, 810685, 850691; Funding text 1: H.Z. acknowledges the financial support from China Scholarship Council. The authors gratefully appreciate the technical support by Mike Wels, Dr. Rein Verbeke and Dr. Toon Brans as well as the Center for Advanced Light Microscopy at Ghent University (Belgium). The authors thank the European Research Council (ERC) under the European Union's Horizon 2020 research and innovation program (No. 648124 and 850691) and DELNAM (No. 810685) for funding. F.H.H. acknowledges funding from the Baden‐Württemberg Stiftung (MIVT5). F.S., F.H., and J.P.S. acknowledge support from the Baden‐Württemberg Stiftung (MIVT‐5) and Deutsche Forschungsgemeinschaft (DFG, German Research Foundation) under Germany's Excellence Strategy via the Excellence Cluster 3D Matter Made to Order (EXC‐2082/1 – 390761711). A.‐K.M. acknowledges the financial support from the Research Foundation‐Flanders, Belgium (FWO Vlaanderen) with grant number 1S28420N (FWO‐SB).
2021-12-302021-12-302023-05-25Bibliographically approved