Fig. 1

TNF-α induces VCAM-1 expression in HCFs. a Time and concentration dependence of TNF-α-induced VCAM-1 expression, HCFs were treated with various concentrations of TNF-α (0.15, 1.5, 15, or 30 ng/ml) for the indicated time intervals. The cell lysates were prepared and blotted using VCAM-1 or GAPDH (as a control) antibody. b For VCAM-1 mRNA expression, cells were treated with TNF-α (15 ng/ml) for the indicated times. The VCAM-1 mRNA expression was analyzed by real time-qPCR. c For VCAM-1 promoter activity, cells were transiently transfected with a VCAM-1-Luc reporter gene and then treated with TNF-α (15 ng/ml) for the indicated time intervals. The promoter luciferase activity was analyzed and normalized to β-galactosidase activity as described in “Methods”. d For adhesion activity, cells were treated with TNF-α for 16 h and washed with PBS prior to the addition of THP-1 monocytes. Moreover, functional expression of VCAM-1 was determined by THP-1 monocyte adhesion assay in the absence or presence of VCAM-1 neutralizing antibody (VCAM-1 nAb, 2 μg/ml). Data are expressed as mean ± SEM of three individual experiments (n = 3). ^* P < 0.05, ^# P < 0.01 vs. vehicle (A-C) or TNF-α (d) alone