VCAM-1 ablation in endothelial and fibroblast cell cultures. (A) VCAM-1 RNA expression is significantly decreased in endothelial cultures from bone or lung of VCAM-1^Δ/Δ mice compared with controls. By contrast, no significant differences are seen in Tie2 expression in all groups. Endothelial cultures were established from the bones and lungs of the wild type (+/+), VCAM-1^f/f, and VCAM-1^Δ/Δ mice. Total RNA was isolated from these cultures, as well as from uncultured bone marrow cells, and reverse transcribed using oligo-dT primer. PCR was performed on serially diluted RT templates (dilution factor of 4) with VCAM-1–, Tie2- or actin-specific primer sets. (B) Partial deletion of the VCAM-1 gene in the fibroblast cultures from the bone marrow of VCAM-1^Δ/Δ mice. PCR analysis of genomic DNA isolated from fibroblast cultures and uncultured BM cells of VCAM-1^f/f and VCAM-1^Δ/Δ mice was done to show the floxed and Δ alleles of the VCAM-1 gene. (C) VCAM-1 RNA expression is decreased in fibroblast cultures of VCAM-1^Δ/Δ mice compared with control-cultured fibroblasts. Total RNA was isolated from the bone marrow fibroblast cultures, as well as from uncultured BM cells of VCAM-1^f/f and VCAM-1^Δ/Δ mice, and reverse transcribed using oligo-dT primer. PCR was performed with VCAM-1–specific primers recognizing the long VCAM-1 isoform (7D VCAM-1) or GPI-linked VCAM-1 isoform (GPI/VCAM-1), as well as with primers for the fibroblast-specific receptor DDR2, and actin-specific primers as internal controls.