Angiogenesis is essential for tumour growth and
metastasis. It is controlled by angiogenic factors, one of the most important being
vascular endothelial growth factor (
VEGF)-A. Although its role has been demonstrated in many tumour types including
colorectal carcinoma (CRC), the importance of the newer family members in
adenoma, invasive tumour growth, and progression to a metastatic phenotype has been poorly characterized in CRC. The aim of this study was to determine the role and timing of the
VEGF angiogenic switch during CRC progression. We measured the gene expression of
VEGF ligands (
VEGF-A,
VEGF-B,
VEGF-C, and
VEGF-D) and their receptors (VEGFR-1, VEGFR-2, and VEGFR-3), in normal colorectal tissues (n = 20),
adenomas (n = 10), and in CRC (n = 71) representing different Duke's stages using
ribonuclease protection assay, semi-quantitative relative
reverse transcriptase polymerase chain reaction, together with the pattern of their expression by immunohistochemistry.
VEGF-A mRNA was the most abundant in colorectal tissue, followed by
VEGF-B,
VEGF-C, and
VEGF-D.
VEGF-A and
VEGF-B mRNAs were significantly more abundant in
adenomas (p = 0.0003 and p = 0.04 respectively) compared with normal tissues, while
VEGF-A and
VEGF-C were significantly increased in
carcinomas compared with normal tissues (p = 0.0006 and p = 0.0009 respectively). A significantly greater amount of
VEGF-C mRNA was present in
carcinomas compared with
adenomas (p = 0.03), whereas there was a significant reduction of
VEGF-B in
carcinomas compared with
adenomas (p = 0.0002).
VEGF-D mRNA was significantly more abundant in normal tissues than in
adenomas (p = 0.0001) and
carcinomas (p < 0.0001). In normal tissues distant from the primary tumour, there was a significantly greater amount of
VEGF-A and
VEGF-D mRNA in patients with Duke's B and Duke's C respectively, compared with Duke's A stage tumours (p = 0.04 and p = 0.01 respectively). Immunohistochemistry showed low basal levels of all
ligands in histologically normal tissues and their expression in the epithelium of tumours reflected the levels of
mRNA expression identified.
VEGF-A and
VEGF-C mRNA levels correlated significantly with tumour grade (p = 0.01 and p = 0.01 respectively) and tumour size (p = 0.001 and p = 0.01 respectively), but not with patient age, sex, presence of infiltrative margin, lymphocytic response, vascular invasion, Duke's stage, or lymph node involvement (p > 0.05).
VEGF-B mRNA correlated with an infiltrative margin (p = 0.04) but no other clinicopathological variable, and expression of
VEGF-D demonstrated no association with any parameter examined.
VEGFR-1 was significantly correlated with tumour grade (p = 0.02), Duke's stage (p < 0.001), and lymph node involvement (p = 0.004),
VEGFR-2 with lymph node involvement (p = 0.02), and
VEGFR-3 did not correlate with any of the clinicopathological variables tested. These results suggest that
VEGF-A and
VEGF-B play a role early in tumour development at the stage of
adenoma formation and that
VEGF-C plays a role in advanced disease when there is more likelihood of metastatic spread. The finding of increased levels of
VEGF-A and
VEGF-D expression in normal tissues collected from a site distant from the primary tumour indicates changes in the surrounding tumour environment that may enhance the subsequent spread of tumour cells.