Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: Repressors NFI and NFY Participate in Organ-Specific Regulation of von Willebrand Factor Promoter Activity in Transgenic Mice

doi: 10.1161/atvbaha.110.206680

Figure Lengend Snippet: Figure 4. Immunofluorescent analyses of NFI iso- form expression in organs of mice. Sections, 5 m, from OCT frozen heart, kidney, lung, and brain of a control C57Bl/6 mouse that were arranged on tissue arrays were treated with anti- bodies specific to NFIA, NFIB, NFIC, and NFIX iso- forms of NFI (red) and simultaneously with anti- PECAM (green) antibodies to detect endothelial cells, as described in the “Methods” section. The results are representative of 2 independent experi- ments for each NFI antibody plus PECAM antibod- ies (magnification 600). The insets show magni- fied endothelial cell nuclei (4,6-diamidino-2- phenylindole).

Article Snippet: Primary antibodies used were polyclonal rabbit antibodies specifically detecting NFIA, NFIB, NFIC, NFIX (Aviva System Biology, San Diego, CA), anti-β galactosidase antibody (Fitzgerald, Concord, MA) and goat polyclonal anti- mouse PECAM antibody (Santa Cruz Biotechnology Inc., Santa Cruz, CA).

Techniques: Expressing, Control

Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: Repressors NFI and NFY Participate in Organ-Specific Regulation of von Willebrand Factor Promoter Activity in Transgenic Mice

doi: 10.1161/atvbaha.110.206680

Figure Lengend Snippet: Figure 5. ChIP detects the association of NFI and YY1 with their cognate binding sites and chromatin looping. ChIP and quantitative PCR analyses were performed on LMEC and HEK 293 cells, as described in the “Methods” section. A, Schematic of VWF chromatin looping that could bring NFI in proximity to the intron 51 YY1-binding site. B and C, Cross-linked chromatin from LMEC (B) and HEK 293 (C) cells were immunoprecipitated with antibodies specific to NFIA, NFIB, NFIC, NFIX, and YY1. Rabbit IgG antibody was used as a negative control. Immuno- precipitates were subjected to quantitative RT-PCR using prim- ers that specifically amplified an 89-bp upstream region of the VWF promoter containing the NFI binding site (NFI primers) or a 182-bp fragment of the VWF intron 51 sequence containing the YY1-binding site (I51 primers). Results for each antibody are presented as fold increase vs control IgG antibody, and all ChIP represents analyses of 2 to 4 independent ChIP experiments for each antibody.

Article Snippet: Primary antibodies used were polyclonal rabbit antibodies specifically detecting NFIA, NFIB, NFIC, NFIX (Aviva System Biology, San Diego, CA), anti-β galactosidase antibody (Fitzgerald, Concord, MA) and goat polyclonal anti- mouse PECAM antibody (Santa Cruz Biotechnology Inc., Santa Cruz, CA).

Techniques: Binding Assay, Real-time Polymerase Chain Reaction, Immunoprecipitation, Negative Control, Quantitative RT-PCR, Sequencing, Control