Figure 2

Cytotoxicity induced by RNA transfer is dependent on endocytic uptake. eGFP encoding mRNA was transferred into primary cortical neurons using endocytosis-dependent (lipofection, (a)) and -independent (fusion, (b)) transfection reagents. Transfer was performed in the absence and presence of endosome inhibitor β-MCD and cells were analyzed 24 h after treatment. Transfection efficiencies are indicated. Identically treated cells were analyzed for eGFP intensity by flow cytometry (c), relative amount of transferred eGFP mRNA (normalized to active endocytotic cells (c,d) and to the respective highest values of each measurement in (e,f)) (d), IL-6 expression (e,f) and number of dead cells (g,h). For better comparison, IL-6 values are directly compared to transferred eGFP mRNA levels (e,d). Percentage of dead cells is given in comparison to fluorescence intensity (g,h). L = lipofection, F = fusion. eGFP-mRNA was used in a concentration of 4 µg/mL for fusion and 2 µg/mL for lipofection. In all cases, a total of 1 µg was transferred per substrate. Scale bar: 200 µm, n = three independent experiments for each analysis. p-values: not significant (n.s.): p > 0.05, *: p ≤ 0.05, **: p ≤ 0.01, ***: p ≤ 0.001.