961 Toll-Like Receptor 4 Activation May Promote Development of Esophageal Adenocarcinoma in Barrett's Esophagus Through Induction of COX2

Background: Limited studies indicate that VEGF expression is upregulated in colorectal cancer (CRC). Since normal colonic epithelial cells have minimal or no expression of VEGF, increased VEGF in CRC represents its aberrant expression. The mechanisms of this phenomenon are not explained. Importins mediate in some cells (e.g., HeLa) the nuclear transport of transcription factors and signaling proteins that potentially can activate the VEGF gene. We hypothesized that upregulation of importins α and/or β is the underlying mechanism for the aberrant VEGF expression in CRC and cell proliferation by facilitating the nuclear transport of transcription factors (P-STAT3 and P-CREB) and P-MAPK/Erk1/2, respectively. Methods: We used: (1) human colon tissue arrays (CRC and normal specimens, n=60), and (2) human CRC cell lines HCT116 & HT29 and normal colonic epithelial cells NCM356 & NCM460 cultured in nutrient media. Studies: 1) Expressions of VEGF, its receptors (VEGFR1 & VEGFR2), importins, P-CREB, P-STAT3 and P-MAPK/Erk1/2 proteins in CRC cells in tissue array, and in cultured CRC and normal epithelial cells by immunostaining and/or western blotting; 2) In cultured CRC and normal epithelial cells: a) downregulation of importins with specific siRNAs, b) expression of VEGF mRNA by real time RTPCR, c) cell proliferation by BrdU assay, d) apoptosis by TUNEL staining.Results: Expression of VEGF and its receptor was aberrant and was significantly increased by 2-4 fold (P 80% (p<0.001). Conclusions: 1) Expression of VEGF and its receptors is upregulated in CRC cells, 2) Aberrant VEGF expression in CRC cells is dependent on upregulation of importin and is mediated by increased nuclear transport of P-CREB and P-STAT3, which bind to and activate VEGF gene promoter, 3) Increased expression of VEGF and its receptors in CRC cells suggests a direct regulation of their growth by VEGF through an autocrine mechanism.