Effects of Sphingosine 1-Phosphate/Sphingosine-1-Phosphate Receptor on Pro-Angiogenesis in HT-29 Human Colon Cancer Cells

Abstract

To investigate the effects and mechanism of sphingosine-1-phosphate/sphingosine-1-phosphate receptor on pro-angiogenesis in human colon cancer cells is the main objective. The effects of sphingosine-1-phosphate on the pro-angiogenesis in human colon cancer cells HT-29 and SW480 were assessed by tube formation assay. Quantitative real-time polymerase chain reaction was used to detect the expression of interleukin-8, interleukin-6 and vascular endothelial growth factor in HT29 and SW480 cells after sphingosine-1-phosphate treatment. Small interfering ribonucleic acid was designed to silence the expression of sphingosine-1- phosphate receptor 1, sphingosine-1-phosphate receptor 2 and sphingosine-1-phosphate receptor 3 in HT-29 and SW480 cells. Western blot and quantitative real-time polymerase chain reaction were used to assess the down-regulation efficiency of sphingosine-1-phosphate receptor in HT-29 and SW480 cells. Later, the effects of sphingosine-1-phosphate receptor were detected by quantitative real-time polymerase chain reaction. Tube formation assay showed that the number of formed tubes in human umbilical vein endothelial cell-fused cells (EA.hy926) resuspended in the culture supernatant from HT-29 and SW480 cells treated with sphingosine-1-phosphate was significantly more than that of the control group (t=3.667 and 4.881, p=0.021 and 0.013), indicating that sphingosine-1-phosphate promoted the pro-angiogenic ability of colon cancer cells. Moreover, the messenger ribonucleic acid expression of interleukin-8, interleukin-6 and vascular endothelial growth factor were significantly increased after sphingosine-1-phosphate treatment compared to the control group (p<0.05) where sphingosine-1-phosphate receptor 1 gene silence could significantly decrease the messenger ribonucleic acid expression in HT-29 and SW480 cells (p<0.05), while sphingosine-1-phosphate receptor 2 and sphingosine-1-phosphate receptor 3 gene silence did not lead to significant messenger ribonucleic acid changes. Sphingosine-1-phosphate up-regulated the expression of interleukin-8, interleukin-6 and vascular endothelial growth factor in human colon cancer cells through sphingosine-1-phosphate receptor 1 and thereby enhancing the proangiogenic effect of colon cancer cells. The sphingosine-1-phosphate/sphingosine-1-phosphate receptor pathway is a promising novel therapeutic target for inhibiting colon cancer growth.