Skip to main content
. Author manuscript; available in PMC: 2010 Feb 9.
Published in final edited form as: Circ Res. 2009 Aug 27;105(7):639. doi: 10.1161/CIRCRESAHA.109.205120

Figure 3. CLIC4 lies downstream of BMPRII, not RAGE, and is necessary in S100A4/Mts1 and BMP-2 induced migration of hPASMCs.

Figure 3

Figure 3

(A) Quantitative RT-PCR of CLIC4 mRNA normalized to β2 Microglobulin (β2M) mRNA in hPASMCs without stimulation and following S100A4 and BMP-2 for 6h ± 30min pretreatment with anti-RAGE (1:1000) or 48h after transfection with BMPRII RNAi (80% knock-down) or CLIC4 RNAi (B). (C) Western immunoblot and densitometry show a reduction in protein to less than 10% after CLIC4 RNAi. (D) Relative migration ± stimulation with S100A4 or BMP-2 of hPASMC transfected with Con or CLIC4 RNAi. Bars represent mean±SEM for n=3 in A and n=4 in C. *p<0.001 vs. control unstimulated. (E) Immunolocalization of CLIC4 in hPASMCs under control conditions and following 2h stimulation with S100A4 and BMP-2. CLIC4, localized with a primary antibody and a secondary antibody conjugated to fluorescein (FITC) exhibits some nuclear staining under control conditions. In response to stimulation with BMP-2 and S100A4, CLIC4 staining becomes increasingly perinuclear and diffusely cytoplasmic, and localizes to the cell membrane, particularly in filopodiae (arrow). Nuclei counterstained with DAPI. (Bars represent 10μm).