Platelet-derived endothelial cell growth factor in human colon cancer angiogenesis: Role of infiltrating cells

Background: Development of new blood vessels is essential for tumor growth and metastasis and depends on the production of angiogenic factors by tumor and/or infiltrating cells, We previously showed that vascular endothelial growth factor (VEGF) expression and vessel count correlate with metastasis in human colon cancer. Although most tumors with high vessel counts express high levels of VEGF, some do not. Recently, platelet-derived endothelial cell growth factor (PD-ECGF), another potent angiogenic factor, has been reported to be expressed in colon cancer. Purpose: In this study, we examined the role of PD-ECGF in colon cancer angiogenesis and whether PD-ECGF is derived from the tumor or infiltrating cells. Methods: Immunostaining for PD-ECGF was performed on 96 colon cancer specimens, some of which were previously stained for VEGF and factor VIII, a marker that is specific for endothelial cells. Double staining was done by using antibodies to PD-ECGF and to CD68 (macrophage specific) or CDS (lymphocyte specific) to confirm which infiltrating cells produce PD-ECGF. Northern blot analysis for PD-ECGF messenger RNA (mRNA) was performed on four colon cancer specimens and corresponding normal colon mucosae (same patients) and four human colon cancer cell lines (KM12SM, SW620, HT29, and NCI-H508) to determine whether colon cancer epithelium expresses PD-ECGF. Results: Immunohistochemical analysis demonstrated that PD-ECGF was expressed in infiltrating cells in most of the colon cancer specimens (80 [83%] of 96) but rarely in tumor epithelium (five [5%] of 96). Double staining demonstrated that infiltrating cells staining positive for both PD-ECGF and CD68 were more predominant than those staining positive for both PD-ECGF and CD3. The intensity of staining for PD-ECGF in infiltrating cells correlated with vessel counts (Spearman's rank correlation coefficient (R) = .29; P = .004), but did not correlate with the intensity of VEGF staining (R = .176, P = .086) or metastasis (Mann-Whitney U test, P = .253). PD-ECGF staining intensity was higher in specimens with a high vessel count (>50 at high magnification) and low VEGF-staining intensity (less than or equal to 2+) than in specimens with a high vessel count (again, >50) and high VEGF-staining intensity (3+). Northern blot analysis revealed that colon cancer specimens and normal mucosae expressed relatively high levels of PD-ECGF mRNA, whereas PD-ECGF mRNA transcripts were not detectable in colon cancer cell lines. Conclusions and Implications: PD-ECGF expression in human colon cancer specimens is associated with vessel count and may be responsible for tumor vascularity in those tumors with ion: VEGF expression. Infiltrating cells expressing PD-ECGF mas contribute to angiogenesis, thus providing an additional mechanism for tumor neovascularization.