Research Paper Volume 8, Issue 8 pp 1670—1689

Figure 5. BMI1 negatively regulates DUSP1/MKP-1 in hUCB-MSCs. (A) DUSP1 and p16^INK4a mRNA expression levels were investigated in replicatively senescent hUCB-MSCs in normoxia/hypoxia culture conditions after exposure to IFN-γ and TNF-α for 30 minutes. (B) The expression levels of MKP-1, p-p38 and BMI1 in normoxic- or hypoxic-cultured hUCB-MSCs were investigated via western blot analysis in replicatively senescent hUCB-MSCs after treatment with IFN-γ and TNF-α for 30minutes. (C) COX-2 and DUSP1 mRNA expression levels were assessed in GFP- and BMI1-overexpressing hUCB-MSCs after activation with IFN-γ and TNF-α for 30 minutes. (D) MKP-1 protein expression levels were investigated via western blot analysis in normoxic culture conditions with BMI1-overexpressing hUCB-MSCs. (E) MKP-1 protein expression levels were assessed via western blot analysis in hypoxic culture conditions with BMI1-down-regulated hUCB-MSCs. (F-G) MKP-1 expressions were analyzed by immunocytochemistry in GFP/BMI1 overexpressing and Luc/shBMI1 transfected hUCB-MSCs. The graphs show the fluorescence intensity of MKP-1 in each cells. Error bars represent mean±s.e.m. from three separate experiments. * P<0.05, ** P<0.01, *** P<0.005 using Student’s t-test.