Fig. 1: TGFβ priming switches the action of TNF to effectively drive osteoclastogenesis. | Nature Communications

Fig. 1: TGFβ priming switches the action of TNF to effectively drive osteoclastogenesis.

From: TGFβ reprograms TNF stimulation of macrophages towards a non-canonical pathway driving inflammatory osteoclastogenesis

Fig. 1

a Human osteoclast differentiation determined by TRAP staining (left) and the relative area of TRAP-positive multinuclear osteoclasts (MNCs, ≥3 nuclei/cell) per well (right) in the cell cultures using human CD14(+)-monocytes treated with or without TGFβ for 3 days, followed by TNF stimulation for 6 days in the presence or absence of TGFβ. Mono-Mφ: monocyte to macrophage stage. TRAP-positive cells: red. (n = 5/group). b Relative area of TRAP-positive-MNCs per well in human TNF-induced osteoclast differentiation with or without TGFβ priming (n = 20/group). c Quantitative real-time PCR (qPCR) analysis of mRNA expression of the indicated genes during osteoclastogenesis using human CD14(+)-monocytes treated with or without TGFβ priming for 3 days, followed by TNF stimulation for the indicated days (n = 5/group). d Von Kossa staining (left) and the resorption area (%) (right) of human osteoclast cultures induced by TNF with or without TGFβ priming (n = 5/group). Mineralized area: black; resorption area: white. e Human TNF-induced osteoclastogenesis using human CD14(+)-monocytes treated with or without TGFβ priming for 3 days, followed by TNF stimulation for 6 days in the presence or absence of recombinant OPG (100 ng/ml). Left: TRAP staining; Right: quantification of the relative area of TRAP-positive-MNCs/well (n = 5/group). f Phagocytosis of zymosan particles (left: zymosan staining; right: quantification of zymosan-containing cells) in the cell cultures using human CD14(+)-monocytes with or without TGFβ priming, followed by TNF stimulation for one day. Fluorescent zymosan particles: green; nuclei: blue; F-actin: red. (n = 6/group). g qPCR analysis of mRNA expression of IL1B and IL6 using human CD14(+)-monocytes treated with or without TGFβ priming for 3 days followed by TNF stimulation for the indicated times (n = 5/group). h Schematic of TGFβ priming effects on TNF-mediated macrophage response. a, c, e, g **p < 0.01; ***p < 0.001; ****p < 0.0001; n.s. not statistically significant by two-way ANOVA with Bonferroni’s multiple comparisons test. b, d, f **p < 0.01; ****p < 0.0001 by two-sided Student’s t test. Error bars: a, c, e, g, Data are mean ± SD. b, d, f Data are mean ± SEM. Scale bars: a, e, f 200 µm; d 100 µm. Source data are provided as a Source Data file.

Back to article page