Fig. 6.
MAP4K4 expression is correlated with TNFα-induced VSMC migration, proinflammatory gene expression and atherosclerotic lesion size in Noxa1SMC-/-/Apoe-/- mice. A, Western blot analysis of MAP4K4 protein levels in cells treated with 100 ng/ml TNFα for 6 h with and without 10 μM DPI. Densitometry analysis of MAP4K4 normalized to β-tubulin levels (mean±SEM; n = 4). B, Western blot analysis of MAP4K4 protein levels in VSMC untransfected and transfected with nontargeting or Map4k4 siRNA. C, VSMC migration assessed by scratch wound assay in cells untransfected and transfected with nontargeting or Map4k4 siRNA in the presence and absence of 100 ng/ml TNFα for 24 h. Maximal wound healing distance is presented (mean ± SEM; n = 3). D, Relative mRNA expression levels as assessed by real time-PCR (mean±SEM, n = 4). E, Representative immunofluorescence images and quantification of aortic root sections stained for MAP4K4 (red), SM-α-actin (green) and with DAPI (blue). Data presented as integrated density of fluorescence per number of cells (mean ± SEM, n = 6). Scale is 100 µm.