Figure 6. PTH and BMP induce commitment of GFP-labeled Sca-1⁺CD45⁻CD11b⁻ MSCs to osteoblast lineage.

(A) Colonies formed from GFP-labeled Sca-1⁺CD45⁻CD11b⁻ MSCs as indicated in CFU-F and CFU-Ob assays. MSCs were plated on 6-well plates at a density of 5 cells/cm² and cultured in culture medium or osteogenic medium. 100 nM PTH, 50 ng/ml BMP, and/or 50 ng/ml noggin were presented from day 1 to day 7 and thereafter with each change of culture medium or osteogenic medium for the entire culture period. The top panels show 6-well plates containing CFU-Fs stained with crystal violate. The bottom panels show 6-well plates containing CFU-Obs stained with Alizarin Red.

(B,C) The colony-forming efficiency was determined by counting the numbers of colonies. The colonies containing 50 or more cells were counted. Each bar correlates in position to treatment group of CFU-F and CFU-OB assays. n = 3. ^*p < 0.05 versus BMP2 alone; ^#p < 0.05 versus BMP2 and noggin.

(D) Illustration of vehicle or intermittent PTH treatment for 7 days in mice with bone marrow cavity transplantation. Total number of 5×10⁵ GFP-labeled Sca-1⁺CD45⁻CD11b⁻ MSCs were transplanted into the femur cavity of Rag2^−/− mice.

(E,F) Intermittent administration of PTH for 7 days stimulated osterix expression in GFP-labeled Sca-1⁺CD45⁻CD11b⁻ MSCs in representative one-parameter histogram of FACS analysis (E) and quantification (F) after transplantation. Quantification of osterix positive GFP-labeled Sca-1⁺CD45⁻CD11b⁻ MSCs is presented as percentage of GFP⁺ cells. n = 6. *p < 0.05 versus Veh.