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. 2020 Dec 29;9:e59659. doi: 10.7554/eLife.59659

Figure 5. Def6 regulates cellular response of osteoblast cells to IFNβ.

(A–D) Immunoblot analysis of type-I IFN receptor expression during osteoblast differentiation (A), the induction of p-STAT1 and 3 (B, C) and expression of STAT1 (D) in WT and Def6-/- osteoblast cells stimulated with mouse recombinant IFN-β (10 U/ml) at the indicated time points. p38 was used as a loading control. The relative density of each band to its corresponding loading control p38 band was calculated by Image J software, and then was normalized to the WT controls at time 0 (the 1st lanes). The relative density for p-STAT3 to total STAT3 was also calculated. (E) qPCR analysis of type-I IFN response gene expression after indicated time points of treatment with IFN-β (10 U/ml) in WT and Def6-/- osteoblast cells. (F) Osteoblast differentiation is induced by the osteogenic medium in the absence or presence of mouse recombinant IFN-β in the WT and Def6-/- osteoblast cells. Alizarin red staining at day 16 (left panel) and its quantification (right panel) were performed. Data are mean ± SEM. *p<0.05, **p<0.01. n.s., not statistically significant. (G) A model showing that the Def6-IFN axis regulates both osteoclast-mediated bone resorption (Binder et al., 2017) and osteoblast-mediated bone formation (current study) in bone homeostasis. Def6 deletion enhances both bone resorption (Binder et al., 2017) and formation (current study) via attenuated type-I IFN-mediated feedback inhibition of the differentiation of both cell types, leading to a high turn-over osteoporotic phenotype in Def6-/- mice.

Figure 5—source data 1. Source data for Figure 5.

Figure 5.

Figure 5—figure supplement 1. qPCR analysis of mRNA expression of Ifnar1 and Ifnar2 during osteoblast differentiation.

Figure 5—figure supplement 1.

Figure 5—figure supplement 1—source data 1. Source data for Figure 5—figure supplement 1.