Figure 2

Tumor necrosis factor (TNF) increases expression of vascular endothelial growth factor-C (VEGF-C) by osteoclast precursors (OCPs). Wild-type (WT) spleen cells were cultured with macrophage colony-stimulating factor (M-CSF) for 3 days to enrich for OCPs. OCPs were cultured in 10% serum with M-CSF and treated with phosphate-buffered saline (PBS) or TNF (10 ng/mL). (a) Expression of VEGF-C and β-actin mRNA was measured by real-time reverse transcription-polymerase chain reaction (RT-PCR) at various times (left panel), and protein levels were measured by Western blot analysis (right panels). (b) Expression of VEGF-A, -B, -C, -D, and placental growth factor (PLGF) mRNA was examined in samples treated for 24 hours. The fold increases in TNF-treated over PBS-treated cells at a given time were calculated. Values are the means of triplicate loadings plus standard deviation. The effect of different doses of TNF (c) or a combination with interleukin 1 (IL-1) (TNF and IL-1 10 ng/mL) (d) on VEGF-C expression at 24 hours was examined by RT-PCR. Experiments were repeated twice with similar results.