Fig 1: Recombinant IGFBP5 protein suppressed osteoblast differentiation and facilitated osteoclastogenesis in vitro. (A) Osteoblast differentiation experimental protocol using MC3T3-E1 cells. DM: differentiation medium (B) representative image of ALP staining by 0 or 200 ng/mL IGFBP5 administration (left) and quantitative data of ALP staining area. n = 9 biological replicates from 3 technical replicates for each condition. (C) Gene expression level in 7 d cultured MC3T3-E1 cells with or without IGFBP5 by RT-qPCR. (D) Experimental protocol for the differentiation of RAW264 cells into osteoclasts by RANKL. (E) Representative image of TRAP staining of multinucleated osteoclasts (MNCs) and quantitative data of number of TRAP+ MNCs/well. n = 8 biological replicates from 2 technical replicates for each strain. Scale bars indicate 100 μm. Data are presented as means ± SD. *, p < .05 by nonparametric Mann–Whitney U test.
Fig 2: Igfbp5 expression was higher in PDGFRα+ cells derived from muscle in Esr1ΔPα mice. (A) Female mice injected with TMX at 12 wk of age were sacrificed at 23 wk and PDGFRα+ cells were isolated from lower limb muscles for RNA-seq. n = 3 biological replicates for each strain. (B) Sorting gate of PDGFRα+ cells by FACS (left) and number of PDGFRα+ cells in Esr1ΔPα and control mice (right). (C) Principal component analysis (PCA) plot. (D) Volcano plot. (E) Heat map of top 15 differentially expressed genes. (F) Enrichment analysis of upregulated genes. The bars represent -log10 q-values of these genes. (G) Genes enriched for regulation of osteoblast differentiation. (H) Serum IGFBP5 concentration. Data are presented as means ± SD. *, p < .05 by nonparametric Mann–Whitney U test.
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