FIGURE 1.
Brachyury (T) transfection provokes expression of cardiopoietic markers in human adipose‐derived mesenchymal stem cells (AMSCs). A, Immunofluorescent images demonstrate increased nuclear expressions of Nkx2.5 and Mef2c in AMSCs 72 hours post‐transfection with M3RNA encoding T (AMSC +T), compared to nontransfected AMSCs. Relative nuclear expressions of Nkx2.5 and Mef2c were compared based on fluorescence intensity normalized to nontransfected AMSCs. Numbers of biological replicates (n) were ≥17 for Nkx2.5 and ≥31 for Mef2c, per experiment condition. Scale bar = 20 μm. B, Increased levels of Nkx2.5 and Mef2c 72 hours post‐transfection were detected by real‐time quantitative polymerase chain reaction (n = 3). C, Western blot confirmed nuclear, not cytoplasmic, translocation of Nkx2.5 (n = 4) and Mef2c (n = 4) in AMSC +T at 72 hours post‐transfection. Protein expression was compared based on nuclear to cytoplasmic ratio of protein levels using (AMSC +T nuclear/AMSC +T cytoplasmic)/(AMSC nuclear/AMSC cytoplasmic) normalized to AMSC. Total protein stain was used as loading control. *P < .05; **P < .01; ***P < .001 with Student's t test