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. 2023 Mar 15;15(3):952. doi: 10.3390/pharmaceutics15030952

Figure 1.

Figure 1

Nanoparticles formed between NF55/mRNA achieve high transfection efficacy in the spleen following systemic administration. (a) In vitro transfection of CHO-K1 cells in serum containing conditions. Analyzed 24 h post-transfection. Two-way ANOVA: F(2, 127) = 15.7, p = 0.00, Tukey post-hoc comparison: *** = p < 0.001. (b) CPP/mRNA nanoparticles and c) CPP/pDNA formed at a charge ratio (CR) 2:1 led to different biodistributions of the expressed reporter. pDNA and mRNA dose 2.5 mg/kg. Compared to CPP/pDNA nanoparticles, at the same nucleic acid dose, the CPP/mRNA nanoparticles led to significantly higher expression in the spleen. In (b) two-way ANOVA F(3, 16) = 8201.4, p = 0.00, Tukey post-hoc comparison: ** = p < 0.01, *** = p < 0.001. (c) mRNA dose reduction in NF55/mRNA nanoparticles results in a significant reduction of expression in the lungs, whereas in the spleen relatively high expression can be achieved even with the lower dose. Comparison of the distribution of reporter between the spleen, liver, and the lungs. NF55/mRNA and NF55/pDNA nanoparticles formed at a charge ratio (CR) 2:1 or 4:1 and with 2.5 mg/kg or 0.5 mg/kg. The distribution of the reporter shown in percentages. (d) Reporter levels were detected from spleen cells collected from NF55/mRNA-treated mice. Suspension—RBC stands for cell suspension where red blood cells have been removed. B + T cells—B cells and T cells collected together. MF + DC—macrophages and dendritic cells collected together. For in vivo experiments shown in (b–d) the tissues were harvested 24 h post-injection. (e) The biodistribution and dime-dependency of expression from delivered mRNA and pDNA shows that NF55/mRNA is expressed mainly in the spleen. NF55/mRNA nanoparticles or NF55/pDNA nanoparticles formed at a charge ratio (CR) 2:1. For mRNA-treated groups a 0.5 mg/kg dose was used and for pDNA-treated groups a 2.5 mg/kg NA dose was used. Left panel shows average radiance in the defined spleen region of treated (NF55/NA) and control (mRNA and pDNA) mice over a 1 to 48 h period. Right panels are representative images from whole-body imaging with equalized radiance scales.