Figure 3
Ang II-Induced nonmuscle myosin II expression in keloid fibroblasts. (A) Keloid fibroblasts were serum starved overnight, followed by 6 h of stimulation. Total RNA was harvested at indicated conditions and subjected to quantitative real-time PCR analysis. The expression of each gene was normalized to the expression of housekeeping gene S9 and reported as percent expression relative to the control of DMEM alone. A representative Western blot is shown above the mean ± SEM of three experiments in separate primary cell lines. (B) Ang II induces dose-dependent expression of NMM IIA (white columns) and IIB (gray columns). Cells were incubated in Ang II (10−5 to 10−8 mol/L) for 24 h prior to lysate collection. (C) Ang II-induced expression of NMM IIA and IIB is time dependent. Cells were stimulated with 10−6 mol/L Ang II for 24–120 h, and lysates were collected. NMM IIA was increased two-fold (white columns) and IIB more than four-fold (gray columns). (D) Ang II-induced expression of NMM IIA and IIB in keloid fibroblasts is mediated by AT1. EMD66684 (AT1 antagonist) or PD123319 (AT2 antagonist) was added to the medium at a concentration of 10−5 mol/L and cells were stimulated with 10−6 mol/L Ang II for 24 h. EMD66684, but not PD123319, inhibited Ang II-induced NMM IIA (white columns) and IIB expression (gray columns). *P ≤ 0.05.