Fig. 2

CD146⁺ pericytes show tissue-intrinsic programming for osteogenic and adipogenic differentiation. a Alizarin red S staining of bone nodules (day 7 of differentiation). b Oil red O staining of intracellular lipid droplets (day 10 of differentiation). Scale bar: 100 μm. c Alcian blue staining of CD146⁺ pericytes as a high-density micromass (day 21 of differentiation). Scale bar: 50 μm. d, e Tubule formation assay. HUVECs were seeded in culture alone, or in co-culture with purified periosteal CD146⁺ pericytes for 2 h. Before co-culture, CD146⁺ periosteal pericytes were labeled with red fluorescent dye (PKH26). d Representative images at ×10 magnification. e Quantification of tubule length. f–h Osteogenic differentiation of total unpurified periosteal cells and CD146⁺ periosteal pericytes derived from the same patient sample. f Alizarin red S staining at day 7 of bone nodules (above) and photometric quantification (below). g Osteogenic gene expression between total periosteal cells and purified CD146⁺ periosteal pericytes, including runt-related transcription factor 2 (RUNX2) and osterix (SP7) at day 7 of differentiation. h Immunofluorescent staining for osteocalcin (OCN) in total periosteal cells and purified CD146⁺ periosteal pericytes. Scale bar: 50 μm. i–l Osteogenic differentiation of periosteal, adipose, and dermal CD146⁺ pericytes. i Alizarin red (AR) staining at day 7 of osteogenic differentiation among periosteal, adipose, and dermal CD146⁺ pericytes. j Photometric quantification of AR staining in i. k, l Osteogenic gene expression among periosteal, adipose, and dermal CD146⁺CD31⁻CD45⁻ pericytes, including (k) RUNX2, and (l) Type I collagen (COL1A1) at day 7 of differentiation. m, n Adipogenic differentiation of periosteal, adipose, and dermal CD146⁺ pericytes. m Oil red O (ORO) staining at day 10 of adipogenic differentiation. Black scale bar: 50 μm. n Photometric quantification of Oil red O staining in m. GM growth medium, ODM osteogenic differentiation medium, ADM adipogenic differentiation medium. *P < 0.05; **P < 0.01; ***P < 0.001