Expression validation of TAGs. (A) Transcriptional validation of TAG markers by quantitative real-time PCR. Expression ratios (± SEM) in TECs compared with NECs (▪) and TECs compared with PLECs (▦) normalized for cyclophilin A are shown. (B) Immunohistochemical staining of CD31/CD34 and different TAGs in colorectal tumor and normal colon tissue sections. Vimentin expression is detected in ECs of tumor and normal colon tissue, although heavily overexpressed on tumor endothelium. IGFBP7 and CD59 show predominant endothelial expression that is more abundant in tumor tissue. HMGB1 is also localized to stromal and epithelial cells, suggestive of secretion. Scale bar = 50 μm. (C) Protein-expression levels (mean fluorescence intensity, ± SEM) of HMGB1, IGFBP7, and vimentin on tumor ECs compared with normal ECs, assessed by flow cytometry (*P < .01; **P < .001).