Fig. 3: CK2 is required for maturation of osteoprogenitors in vitro and in vivo.
From: A RUNX2 stabilization pathway mediates physiologic and pathologic bone formation
a–d COBs isolated from P5 Csnk2bfl/fl pups were infected with lentiviruses expressing vector (WT) or Cre recombinase (Csnk2b KO) and cultured under osteogenic conditions. mRNA levels of Csnk2b (a) and osteogenic genes (d) were measured by RT-PCR at day 12. ALP activity (b, left) and staining (b, right) or alizarin red staining (c) were performed at day 7 or 18, respectively. Scale bar, 250 μm (b). a, d, n = 4; b, c, n = 6. e Csnk2b mRNA levels in P0 calvaria of Csnk2bfl/fl, Osx-Cre, and Csnk2bOsx pups. (n = 4). f Alizarin red/alcian blue staining of skeletal preparations of calvaria (top) and clavicles (bottom) obtained from P10 Csnk2bfl/fl, Osx-Cre, and Csnk2bOsx pups. Scale bar, 2 mm. g MicroCT analysis shows 3D-reconstruction of calvaria from 2-month-old Csnk2bfl/fl, Osx-Cre, and Csnk2bOsx male mice. The arrows indicate hypomineralization areas. Scale bar, 2 mm. h Radiographic images of clavicles from 2-month-old Osx-Cre and Csnk2bOsx male mice. The arrow indicates defective sternoclavicular ossification. Scale bar, 2 mm. i, j MicroCT analysis of femurs from 2-month-old Csnk2bfl/fl, Osx-Cre, and Csnk2bOsx male mice. Quantification (i) and 3D-reconstruction (j) are displayed. Trabecular bone volume/total volume Tb. BV/TV trabecular thickness, Tb.Th trabecular number per cubic millimeter, Tb.N, and C.Th cortical thickness. Scale bar, 1 mm (j). i Tb. BV/TV, Tb.Th, Tb.N; n = 12 (Csnk2bfl/fl), 7 (Osx-Cre) or 5 (Csnk2bOsx), C.Th; n = 13 (Csnk2bfl/fl), 5 (Osx-Cre) or 5 (Csnk2bOsx)). k H&E-stained longitudinal sections of femurs from 2-month-old Osx-Cre and Csnk2bOsx male mice. Scale bar, 50 μm. l, m Histomorphometric analysis of femurs from 2-month-old Osx-Cre and Csnk2bOsx female mice. Images of calcein/alizarin red-labeled sections of the femur (l) and quantification (m) are displayed. Scale bar, 50 μm (l). BFR/BS bone formation rate per bone surface, MAR mineral reposition rate, Ob.S/BS osteoblast surface per bone surface. (n = 9 (Osx-Cre) or 8 (Csnk2bOsx)). Similar skeletal phenotypes were observed in both male and female mice (f–m). Data are representative of three (a–h, j–l) independent experiments or are pooled from two experiments (i, m). A two-tailed unpaired Student’s t test for comparing two groups (a–e, i, m; error bars, SD of biological replicates).
