Figure 10. FGF23 directly inhibits osteoblast differentiation.

(A–C) mRNA expression of markers of osteoblast differentiation in MC3T3-E1 osteoblasts cultured for 21 days and treated with recombinant FGF23 (0, 25, and 50 ng/mL) for the last 6 and 48 hours of culture (n ≥ 4). Levels of (D) total DMP1 (cDMP1) and (E) total FGF23 (cFGF23) measured by ELISA in conditioned culture media from bone marrow stromal cells (BMSCs) isolated from 12-week-old WT (n ≥ 4), Fgf23^Dmp1-cKO (n ≥ 5), Dmp1^KO (n ≥ 5), and Dmp1^KO Fgf23^Dmp1-cKO (n ≥ 5) mice, then cocultured for 21 days with BMSCs isolated from the same group (isogenic), or immortalized BMSCs displaying genetic overexpression of Fgf23 (Fgf23^TG) or Dmp1 (Dmp1^TG). (F and G) Alkaline phosphatase (ALP) staining and quantification and (H and I) alizarin red S (ARS) staining and quantification. Values are expressed as mean ± SEM; P < 0.05 vs. (A–C) time point–matched ^aCtr, ^bFGF23 25 ng/mL, (D–I) culture-matched ^aWT, ^bFgf23^cKO, ^cDmp1^KO, and *genotype-matched isogenic. Statistical tests were ANOVA test followed by post hoc t tests and multiple-testing correction using Holm-Bonferroni method (A–C) and by Bonferroni’s post hoc tests (D–I).