Fig. 9: OPN induces lysosomal-dependent lipid metabolism in BMDMs.

a Representative western blots (left) and quantification (right, n = 3 biologically independent samples) of OPN receptor αv and β3 expression in BMDMs and metabolically activated macrophages (MMes) after culture for 4 days in vitro. b Schematic diagram illustrating the co-culture system as indicated below (c). c, d Representative images (c) and quantification (n = 5 biologically independent samples) (d) of crystal violet staining of BMDMs (ctrl) and MMes treated with or without rOPN for 24 h in a Transwell system. Scale bar: 50 μm. e Immunofluorescence images of buoyant BMDMs (F4/80) containing lipid vesicles (Perilipin 1) isolated from bone marrow supernatant at week 12. Scale bar: 15 μm. f FACS analysis of FC + (F4/80+ and CD11b+ , top) and Perilipin1+ (bottom) BMDMs from NFD- (left) and HFD-fed (right) mice. P9, FC + and Perilipin1+ BMDM population from Q2. g Percentage of FC + and Perilipin1+ BMDMs (n = 5 biologically independent samples). h Lactate concentration in bone marrow supernatant (n = 5 biologically independent samples). i Schematic diagram illustrating the co-culture system as described below (j). j Immunofluorescence staining of the co-cultured BMDMs for 4 days in vitro; the eWAT from mice fed the HFD for 8 weeks was placed in the upper chamber; Neu Ab (2.0 μg/ml) was added into the lower chamber. Scale bar: 50 μm. The right-most image is an enlarged view of the field in the white dashed box. Scale bar: 12.5 μm. k Total intracellular ATP concentration (left, co-culture for 0.5 h) and lactate concentration (right, co-culture for 10 h) in BMDMs with the treatments indicated in j (n = 4 biologically independent samples). l Relative expression of different V-ATPase subunits in ctrl (BMDMs), MMes, and MMes+ rOPN after culture for 24 h (n = 3 biologically independent samples). rOPN, 0.5 μg/ml. Images are representative of 3 independent experiments. All data are presented as mean ± SD. Two-tailed Welch’s t-test (a), one-way ANOVA with Tukey’s post-hoc test (d, k), two-way ANOVA with Sidak’s post-hoc test (g, h), and two-way ANOVA with Tukey’s post-hoc test (l) were used. Source data are provided as a Source Data file.