Opti mem basal medium

In order to transfect hADSCs with modRNAs, MessengerMAX (Invitrogen, California, USA) transfection reagent was employed. For in vitro experiments, modRNAs and transfection reagents were first diluted separately in Opti-MEM basal medium (Invitrogen, California, USA) and incubated for 5 min at room temperature (RT). Afterwards, the two mixes were pooled together and incubated for 15 min to generate modRNA-lipid complexes. modRNA-lipid complexes were exposed to cells for 4 h, after which the medium was completely replaced with cell culture media or removed to collect cells. Specifically, for lipo-complexing, 2 μL of MessengerMAX transfection reagent was used per 1 μg modRNA to transfect per 1 × 10⁵ hADSCs in the in vitro and in vivo experiments.
For evaluating GFP modRNA (modGFP) expression kinetics in hADSCs, the transfection efficiency and mean fluorescence intensities of modGFP were recorded at 4, 8, 16, 24, and 48 h post-transfection, by C6 flow cytometry (Beckman Coulter, CA, USA).

mRNA transfection was carried out with RNAiMAX (Invitrogen) according to the manufacturer's instruction in a 6-well format. Briefly, mRNA and reagent were first diluted in Opti-MEM basal medium (Invitrogen). 1.2 ug (100 ng/μl) RNA was diluted in 48 ul Opti-MEM and 6 ul RNAiMAX was diluted in 54 ul Opti-MEM. Both dilutions were combined to a total of 120 ul, briefly vortexed and incubated for 15 min at room temperature. After complex formation, mix was drop-wise dispensed to culture medium. RNA transfection was performed in Pluriton media (Stemgent) supplemented with Pluriton supplement (Stemgent) and B18R interferon inhibitor (eBioscience) at 200 ng/ml.