Overexpression of vascular endothelial growth factor164 and its co-receptor neuropilin-1 in estrogen-induced rat pituitary tumors and GH3 rat pituitary tumor cells.

We have shown previously that the VEGF system plays a crucial role in regulation of tumor angiogenesis during the development of estrogen-induced prolactin-secreting pituitary tumors in Fisher 344 rats. Studies also suggested that both endothelial and non-endothelial cells expressed VEGF. However, several questions concerning the VEGF signals in regulation of estrogen-induced angiogenesis in rat pituitary remained unanswered. VEGF exists in a number of isoforms in human and rodent tissue (i.e., VEGF206h/205r, VEGF189h/188r, VEGF165h/164r, VEGF145h/144r and VEGF121) that differ in their molecular masses and biological activities. The VEGF isoforms bind with two tyrosine-kinase receptors, KDR/flk-1 and flt-1. In addition, VEGF165 binds with a newly identified co-receptor, neuropilin-1, which is expressed in human endothelial cells and several types of non-endothelial cells including tumor cells. The present study was undertaken to elucidate which isoforms of VEGF are predominantly expressed in normal Fisher 344 rat pituitaries, estrogen-induced prolactin secreting rat pituitary tumors and in prolactin secreting rat pituitary tumor cell line (GH3 cell line). To identify the isoform, RT-PCR with primer pairs derived from exon 1 and exon 8 of the VEGF gene, cloning, sequencing and Western blot analysis were performed. The status of neuropilin-1 in the rat pituitaries (normal and transformed) and GH3 pituitary tumor cell line has also been investigated using RT-PCR and Western blot analysis. These studies demonstrate that normal rat pituitaries, estrogen-induced rat pituitary tumors and GH3 pituitary tumor cells expressed VEGF164 and co-receptor, neuropilin-1. The VEGF164 was the predominant form in all of these cells. The VEGF164 and neuropilin-1 mRNA and protein levels were significantly higher in the estrogen-induced pituitary tumors and GH3 tumor cell line, as compared to normal pituitary. The data suggest that both VEGF164 and neuropilin-1 may actively participate in modulation of tumor angiogenesis and the development of pituitary tumors in Fisher 344 rats.