TY - JOUR
T1 - Cell uptake and intracellular trafficking of bioreducible poly(amidoamine) nanoparticles for efficient mRNA translation in chondrocytes
AU - Pontes, Adriano P
AU - van der Wal, Steffen
AU - Ranamalla, Saketh R
AU - Roelofs, Karin
AU - Tomuta, Ioan
AU - Creemers, Laura B
AU - Rip, Jaap
N1 - Publisher Copyright:
Copyright © 2023 Pontes, van der Wal, Ranamalla, Roelofs, Tomuta, Creemers and Rip.
PY - 2023
Y1 - 2023
N2 - Disulfide-containing poly(amidoamine) (PAA) is a cationic and bioreducible polymer, with potential use as a nanocarrier for mRNA delivery in the treatment of several diseases including osteoarthritis (OA). Successful transfection of joint cells with PAA-based nanoparticles (NPs) was shown previously, but cell uptake, endosomal escape and nanoparticle biodegradation were not studied in detail. In this study, C28/I2 human chondrocytes were transfected with NPs co-formulated with a PEG-polymer coating and loaded with EGFP mRNA for confocal imaging of intracellular trafficking and evaluation of transfection efficiency. Compared with uncoated NPs, PEG-coated NPs showed smaller particle size, neutral surface charge, higher colloidal stability and superior transfection efficiency. Furthermore, endosomal entrapment of these PEG-coated NPs decreased over time and mRNA release could be visualized both
in vitro and in live cells. Importantly, cell treatment with modulators of the intracellular reducing environment showed that glutathione (GSH) concentrations affect translation of the mRNA payload. Finally, we applied a D-optimal experimental design to test different polymer-to-RNA loading ratios and dosages, thus obtaining an optimal formulation with up to ≈80% of GFP-positive cells and without toxic effects. Together, the biocompatibility and high transfection efficiency of this system may be a promising tool for intra-articular delivery of therapeutical mRNA in OA treatment.
AB - Disulfide-containing poly(amidoamine) (PAA) is a cationic and bioreducible polymer, with potential use as a nanocarrier for mRNA delivery in the treatment of several diseases including osteoarthritis (OA). Successful transfection of joint cells with PAA-based nanoparticles (NPs) was shown previously, but cell uptake, endosomal escape and nanoparticle biodegradation were not studied in detail. In this study, C28/I2 human chondrocytes were transfected with NPs co-formulated with a PEG-polymer coating and loaded with EGFP mRNA for confocal imaging of intracellular trafficking and evaluation of transfection efficiency. Compared with uncoated NPs, PEG-coated NPs showed smaller particle size, neutral surface charge, higher colloidal stability and superior transfection efficiency. Furthermore, endosomal entrapment of these PEG-coated NPs decreased over time and mRNA release could be visualized both
in vitro and in live cells. Importantly, cell treatment with modulators of the intracellular reducing environment showed that glutathione (GSH) concentrations affect translation of the mRNA payload. Finally, we applied a D-optimal experimental design to test different polymer-to-RNA loading ratios and dosages, thus obtaining an optimal formulation with up to ≈80% of GFP-positive cells and without toxic effects. Together, the biocompatibility and high transfection efficiency of this system may be a promising tool for intra-articular delivery of therapeutical mRNA in OA treatment.
KW - chondrocyte
KW - mRNA delivery
KW - nanoparticle
KW - non-viral gene delivery
KW - osteoarthritis
KW - poly(amidoamine)
UR - http://www.scopus.com/inward/record.url?scp=85177684062&partnerID=8YFLogxK
U2 - 10.3389/fbioe.2023.1290871
DO - 10.3389/fbioe.2023.1290871
M3 - Article
C2 - 38026902
SN - 2296-4185
VL - 11
JO - Frontiers in bioengineering and biotechnology
JF - Frontiers in bioengineering and biotechnology
M1 - 1290871
ER -