Fig. 2.

TNFα safeguards MSC homeostasis in a receptor-independent manner through endocytosis. a–d Functional analyses of MSCs according to CFU, BrdU labeling, and osteogenic and adipogenic differentiation. MSCs were derived from WT, TNFα^−/− or TNFR^−/− mice (N = 5). Scale bars = 100 μm. e Calcein labeling for bone formation analysis in WT and TNFR^−/− mice (N = 5). Scale bar = 50 μm. f Oli red O staining for bone marrow adiposity in WT and TNFR^−/− mice (N = 5). Scale bar = 150 μm. g Endocytosis analysis of FITC-labeled TNFα uptake by MSCs for 24 h in vitro. Pitstop^® 2 was used to inhibit clathrin-mediated endocytosis at 12 μmol·L⁻¹. N = 3. Scale bars = 20 μm (top) and 7 μm (bottom). h Western blot analysis of protein expression levels (N = 3). i–k Functional analyses of MSCs according to BrdU labeling and osteogenic and adipogenic differentiation. N = 3. Scale bars = 100 μm. l Diagram showing that TNFα regulates MSC homeostasis in a receptor-independent manner through endocytosis. For quantification of Western blotting, a two-tailed Student’s t test was used for the comparison between the treatment and WT groups. *P < 0.05. N.S. not significant. Data represent the mean ± SD