Gene engineered mesenchymal stem cells: greater transgene expression and efficacy with minicircle vs. plasmid DNA vectors in a mouse model of acute lung injury

摘要

Comparison of plasmid or MC-based transfection in mouse MSCs. a Schematic illustration of method in generating minicircles (adapted from www.systembio.com). b Nuclear-targeted electroporation (Amaxa) of MC-ANGPT1 resulted in a 3.7-fold increase in human ANGPT1 protein in conditioned media using ELISA (R&D Systems) at 24 h after transfection, compared to the same amount of plasmid ANGPT1 (pANGPT1). n = 4 experiments. c MSCs transfected with nucleofection using reagent only (mock), empty plasmid (pVAX), human ANGPT1 plasmid (pANGPT1), empty minicircle DNA (MC), or human ANGPT1 minicircle DNA (MC-ANGPT1). MC-ANGPT1 peak values at D1 and D2 compared to pANGPT1 and mock transfected. n = 2 experiments. d MSC viability evaluated by trypan blue exclusion (bar graph) and cell morphology (photomicrographs) at 24 h after transfection. n = 5 experiments. Group comparisons were analyzed by one-way ANOVA with Holm-Sidak’s post hoc test. *p < 0.05, mock-transfection vs. MC-ANGPT1-transfected MSCs. #p < 0.05, pANGPT1-transfection versus MC-ANGPT1-transfected MSCs