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. 2009 Nov 4;42(1):21–29. doi: 10.3858/emm.2010.42.1.002

Figure 2.

Figure 2

Effect of SHP on TGF-β-stimulated PAI-1 mRNA expression and Smad3 activity. (A) Northern blot analysis of the effect of overexpression of SHP on PAI-1 m RNA expression. VSMCs were infected at the indicated moi with Ad-SHP or GFP for 4 h. Cells were incubated in medium containing 0.5% FBS 24 h, and then treated with TGF-β (1 ng/ml) for 3 h before harvest. Data represents the means ± SEM. *P < 0.05, **P < 0.01 as compared to TGF-β alone. (B) Luciferase gene reporter assay of the effect of SHP on PAI-1 promoter activity. (Upper) VSMCs were transfected with -840 PAI-1 promoter Luc (300 ng) together with the indicated amounts of an SHP expression vector (pcDNA3) for 5 h. Cells were incubated in medium containing 0.5% FBS for 24 h, and then treated with TGF-β (1 ng/ml) for 1 h before harvest. Data represents the means ± SEM. *P < 0.01 as compared to the control. **P < 0.05, #P < 0.01 as compared to TGF-β alone. (Lower) VSMCs were co-transfected with -840 PAI-1 promoter Luc (300 ng) and expression vectors for Smad3 (100 ng), Smad4 (100 ng) and ALK5 (100 ng), together with the indicated amounts of SHP expression vector for 5 h. Cells were then incubated in medium containing 0.5% FBS for 24 h. Data represents the means ± SEM. *P < 0.001 as compared to control. **P < 0.01, #P < 0.001 as compared to Smad3/4/ALK5 alone.