MuSCs were differentiated in differentiation medium (DMEM (Invitrogen) with 2% horse serum (Sigma)).
Two days after plating, cells were differentiated using "Differentiation Medium" (catalog # DM-2, ZenBio) and considered as Day '0'.
At confluence (day 0), preadipocytes were differentiated in differentiation media (DM) for 72 h (day 03) and were maintained in nutrition media (NM) thereafter for 11 days (day 14).
Fifty thousand cells per well were seeded on Seahorse plate and differentiated in differentiation media containing 20% FBS, 20 n m insulin and 1 n m triiodothyronine for 48 h, than were supplemented with 0.5 m m dexamethasone, 0.5 m m isobutylmethylxanthine, and 0.125 m m indomethacin for 48 h, and returned to differentiation media.
We, therefore, argue that the CSCs are not on the apex of the hierarchy but rather exist in parallel to the differentiated cells, with differentiation and de-differentiation being common features within a solid tumor.
To characterize the cells, lnASCs or obASCs transfected with the leptin shRNA or control shRNA were differentiated in osteogenic differentiation medium or adipogenic differentiation medium, plated to determine colony forming unit capacity, and analyzed for the expression of cell surface markers.
Tumors were classified also by their degree of histological differentiation: (well differentiated [I]), moderately differentiated [II], or poorly differentiated [III]), the presence or absence of perineural invasion, the presence or absence of venous emboli, and the number of lymph nodes involved by the tumor.
Whole-cell perforated patch recordings were performed from sorted HUES-9 NSC that were differentiated in neuron differentiation medium for 3 weeks.
ESC chondrogenesis can be characterized by five main stages of cellular, extracellular and molecular events: (1) Condensation of differentiated ESCs, (2) Differentiation into chondrocytes and fibril scaffold formation, (3) ECM deposition and cartilage formation, (4) Hypertrophy and degradation of cartilage, and (5) Bone replacement.
The accelerated cell division of undifferentiated E/M cells might be supported by the high production of extracellular matrix proteins, observed in early passages (Figure 3C), whereas low cell densities of E/M-MP cells, or growth among more differentiated cells, enforces differentiation and the rapid loss of CD133 (Figures 7A,B).
