Fig. 5: Determination of cell cycle- and endogenous autophagy-associated cellular accumulation of DiD-labeled lipid nanoparticles (DiD-LNP) using the newly established technical platform.

a Statistical analyses of Total Fluorescence Intensity (TFI) in G1-phase, S-phase, G2-phase and M-phase WT and ATG7 KO PIP-FUCCI cells, respectively, measured at two-dimensional (2D, upper panel) and three-dimensional (3D, lower panel) levels. b Statistical analyses of Mean Fluorescence Intensity (MFI) in WT and ATG7 KO PIP-FUCCI cells, respectively, measured at four distinct cell cycle phases, as well as at 2D (upper panel) and 3D (lower panel) levels. Cancer cells were incubated with DiD-LNP at a dose equivalent to 6 μg/mL mRNA for 2 h. The same volume of PBS was used as a control. All data were log₁₀-transformed and presented as mean ± SD (WT PIP-FUCCI: n = 30, 28, 26, 30; ATG7 KO PIP-FUCCI: n = 28, 30, 26, 27 cells for G1 phase, S phase, G2 phase, and M phase, respectively). Statistical analyses of DiD-LNP accumulation among four distinct cell cycle phases were conducted using one-way ANOVA followed by Bonferroni multiple comparisons test, while differences between WT and ATG7 KO PIP-FUCCI cell lines were evaluated with two-tailed independent sample t-tests (complete statistical results provided in Supplementary Table 12). The level of significance was set at p < 0.05. WT PIP-FUCCI, wild-type U2OS cells stably expressing PIP-FUCCI; ATG7 KO PIP-FUCCI, ATG7 knockout U2OS cells stably expressing PIP-FUCCI. Source data are provided as a Source Data file.