Fig. 2

The specific lipid composition of the nanosystems has an influence on M2 macrophage reprogramming. a Experimental design for macrophage polarization and treatment with the lipid nanosystems: murine immortalized bone marrow-derived macrophages (IBMDM) were induced to an M1 phenotype using LPS (10 ng/mL) + IFN-γ (10 ng/mL) or to an M2 phenotype with IL4 (10 ng/mL) for 24 h. Following polarization, lipid nanosystems (1 mg/mL) were incubated with macrophages for a 4-h duration: VitE:SM, M:SM, VitE:PC, VitE:SM:PI, VitE:SM:PS, VitE:SM:PG, and VitE:SM:PA at different concentrations of PA. Subsequently, IBMDM cells were washed and cultured in RPMI medium for qRT-PCR and toxicity assays for 24 h, or Western blot analyses for 48 h. b Fold change in relative luciferase activity (i.e., toxicity) ± SD determined in IBMDM cultures treated with the indicated different nanoemulsion compositions compared to untreated M2 polarized macrophages, set as 1.0 (n = 3). c Analysis of the levels of the principal M2 marker Arg1 by qRT-PCR. Bars represent the mean fold change ± SD (n = 3), with untreated M2 set as 1.0. d Top: Representative Western immunoblots of ARG1 protein expression levels. Bottom: Densitometric analysis of the immunoblots is represented in the bar diagram. Bars represent the mean fold change ± SD (n = 4), with untreated M2 set as 1.0. e Summary table of three parameters chosen for evaluating the best nanoemulsion composition. For toxicity, low < 4 AU, medium = 4–7 AU, high > 7 AU. ∗  = p < 0.05; ∗  ∗  = p < 0.01; ∗  ∗  ∗  = p < 0.001; ∗  ∗  ∗  ∗  = p < 0.0001; ns = not significant. One-way ANOVA test for multiple comparisons with Dunnett’s post hoc test, compared to the untreated M2 sample