Fig 2: FoxO1, FoxO3A, PPAR?, ADRP, ELOVL4 and FABP4 mRNA levels in the meibomian gland were measured by qPCR (A) (n = 6). Western blot assay of indicated FoxO1, FoxO3A, PPAR?, ADRP, ELOVL4, and FABP4 proteins in the meibomian gland homogenates (B) and densitometry analyses of the western blotting results (C) (n = 6). Data are presented as the mean ± SD. *P < 0.05, **P < 0.01, and ***P < 0.001 versus control group.

Fig 3: a Double immunofluorescence analysis of paraffin-embedded liver sections of 52-week-old mice was performed using specific anti-LC3B (green) and LAMP1 (red) antibodies. Nuclei were stained with DAPI (blue). Colocalization of these two proteins appears in yellow. Magnification ×1000 Scale bar 40 µm. b Representative double immunofluorescence analysis of paraffin-embedded liver sections from 52-week-old mice was performed using anti-p62 (green) and anti-LAMP1 (red). Arrowheads indicate colocalization of these proteins shown in yellow color and p62 accumulation in nuclear inclusion. Original image magnification ×1000, bars 40 µm. c Representative immunohistochemical analysis of paraffin-embedded liver sections from 52-week-old mice was performed using specific anti-LAL (lysosomal acid lipase) antibody. Original magnification ×200 and ×1000, bar 100 µm. d Representative immunohistochemical analysis of PLIN2 (blue) and LAMP1 (red). Arrows show costained areas (purple color). Original magnification ×200 and ×1000, bar 100 µm

Fig 4: Oleate induces polarization of human monocytes into CD206+ suppressive cells A1 × 106 CD14+ myeloid cells from healthy donors were polarized by 1 ng/ml GM-CSF in the presence of BSA control (C), 0.2 mM oleate (O), or 0.2 mM stearate (S) over a 6-day period. The expression of CD206, CD204, and CD38 was analyzed via flow cytometry.BThe level of lipid droplets was determined via BODIPY staining.CThe oxygen consumption rate (OCR) of differentiated myeloid cells was monitored as described in Materials and Methods.DThe purified CFSE-labeled autologous CD4+ T cells were co-cultured with polarized myeloid cells with indicated treatment for T-cell proliferation assay.ETumor tissue (tumor) as well as corresponding non-tumorous adjacent tissue (control) was collected from colorectal cancer patients and prepared for histopathology. The expression of CD68 (red), CD206 (green), and ADRP (blue) indicates the lipid droplets in tumor-infiltrating myeloid cells. Nuclei (white) were stained using DAPI. The absolute number of positive cells was quantified in 5 high-power fields (hpf; scale bar = 20 µm).Data information: Shown is the mean ± SD from two to four independent experiments and analyzed by either one-way ANOVA (A), Kruskal–Wallis test with Dunn's multiple comparisons test (D) or unpaired Student's two-tailed t-tests (E). *P < 0.05; **P = 0.01; ***P = 0.001.

Fig 5: Conditioned medium from Adipose-derived mesenchymal stromal cells (ASCcm) induce macrophage polarization and promote lipid droplet biogenesis. After differentiation with L929 medium, macrophages were seeded and cultured with fresh medium (CTRL) or fresh medium plus ASCcm (50%). After treatment, arginase expression was analyzed by western blot. (A) and arginase activity was measured in cell lysates. (B) The IL-10 content measured in supernatants by ELISA, after macrophage re-education with ASCcm for 24 h followed by LPS + IFN? stimulation for an extra 24 h. (C) ASCcm induces cellular elongation phenotype in macrophages, as shown by phalloidin staining. (D) A significant increase of Lipid droplet number is observed in response to ASCcm. Microscopy images obtained from control (non-treated) or ASCcm- treated macrophage stained with Plin2 (red) and Bodipy (green). The yellow dot represents the merge of Plin2 and Bodipy. (E) The images are representative of at least six different experiments. Labeled lipid droplets were quantified by the measurement of fluorescent area per cell using ImageJ software. (F) Analysis of PGE2 production by macrophage was performed by EIA in the supernatant. (G) Analysis of cPLA2 and COX-2 in total cell lysates of macrophages by Western blot. (H) ß-Actin levels were used for control of protein loading. Data are expressed as mean ± SEM of three independent experiment for supernatant dosages and immunofluorescence and six independent experiments for western blot analyses. *p < 0.05 versus non treated cells (CTRL).