The genes of interest were glyceraldehyde 3-phosphate dehydrogenase (GAPDH, control), osteocalcin (BGLAP
), transcription factor RunX2 (RunX), alkaline phosphatase (AP), and SMAD family member 4 (SMAD4) (Table 1).
All reactions were performed using SYBR Green PCR Master Mix (Applied Biosystems, Foster City, CA) on a C1000 Touch[TM] Thermal Cycler (Bio-Rad Laboratories, Hercules, CA) for the following genes: scleraxis (SCX), mohawk (MKX), tenomodulin (TNMD), thrombospondin-4 (THBS4), tenascin C (TNC), collagen type I (COL1A1), decorin (DCN), elastin (ELN), peroxisome proliferatoractivated receptor (PPAR[gamma]), adipocyte-binding protein 2 (aP2), runt-related transcription factor 2 (RUNX2), osteopontin (SPP1), osteocalcin (BGLAP
), sex-determining region Y-box9 (SOX9), collagen type II (COL2A1), aggrecan (ACAN), PouV, Nanog, and Sox2.
The expression of adipogenic differentiation-related genes (Adiponectin, Cebp[alpha], Pparyl, and Ppar[gamma]2) and osteogenetic differentiation-related genes (Runx2, Bglap
, Sppl, Ibsp, Sp7, and Alp) was significantly increased when cells were cultured in differentiation medium in 21% and 5% oxygen condition (Figures 2(e) and 2(g)).