Osteo-, adipo-, and chondrogenic potential of fractionated endosteal cells. (A) ALP staining of freshly isolated ALCAM⁺Sca-1⁻, ALCAM⁻Sca-1⁻, and ALCAM⁻Sca-1⁺ populations. Scale bar: 50 μm. (B) Differentiation of osteoblasts from ALCAM⁺Sca-1⁻, ALCAM⁻Sca-1⁻, and ALCAM⁻Sca-1⁺ populations after osteogenic induction. Osteoblastic differentiation was determined by ALP staining (top) and calcium deposition (alizarin red staining, bottom) after osteogenic induction. Scale bar: 50 μm (upper), 0.5 mm (lower). (C) Differentiation of adipocytes (oil red O-positive cells) from each population after adipogenic induction. Scale bar: 200 μm. (D) Alcian blue staining after chondrogenic induction. Scale bar: 200 μm. (E) qPCR analysis of the osteoblastic markers Runx2, Opn, and Ocn in fractionated endosteal populations. Data represent means ± SD (*P < .01, **P < .05). Representative data from 3 independent experiments are shown. (F) qPCR analysis of the MPC marker Eng in fractionated endosteal populations. Data represent means ± SD (**P < .05). Representative data from 3 independent experiments are shown. (G) Representative results of flow cytometric analysis of the expression of CD90 (top) and Endoglin (bottom) in ALCAM⁺Sca-1⁻, ALCAM⁻Sca-1⁻, and ALCAM⁻Sca-1⁺ populations. Red line, specific Abs; gray dotted line, isotype control. Data represent means ± SD.