Figure 2
FN and TNC expression in different endothelial cell models. (a) Western analysis (cropped blots) of FN (top) and TNC (bottom) expression in indicated cells. Culture medium (25 µl), left; total cell extracts of equal protein concentration (40 µg), right. Equivalent amounts of FN-depleted serum-containing (10%) culture medium were deposited as control. α-tubulin was used as loading control for total cell extracts. Full-length blots are shown in Fig. S5. (b) Detection of cFN isoform transcripts (containing EDB+/EDA+, EDB+/EDA−, EDB−/EDA+, EDB−/EDA−, as schematically presented on the top) by RT-PCR in human endothelial cells and TIFs. 18S rRNA was used as internal control/housekeeping gene. Cropped blots are shown in (a) and (b). (c) QPCR analysis of FN, FN-EDB, FN-EDA and TNC mRNA expression in endothelial cells, normalized with GAPDH and presented as log ΔCt (±S.D., n = 5). (d) Double immunofluorescence staining (wide-field fluorescence) of total FN (red) and, in green, FN-EDA (left); FN-EDB (middle) or TNC (right) in confluent endothelial cells and TIFs. Cell density was evaluated after Hoechst 33342 staining (30–50 cells in each field). Bar = 100.
