Effects of small interfering RNA tareting teratocarcinoma-derived growth factor-1 gene expression on cell adhesion and invasion and urokinase-type plasminogen activator of human malignant melanoma cell

Objective To study the effects of teratocarcinoma-derived growth factor-1 (TDGF-1) gene small interfering RNA (siRNA) on adhesion and invasion of human melanoma cell.Methods Realtime fluorescent quantitative polymerase chain reaction(FQ-PCR) was used to evaluate the TDGF-1 mRNA expression of human melanoma cell lines A-375,C-918 and M14.The highest expression of TDGF-1 was transfected with different dose of TDGF-1 siRNA.The expression of TDGF-1 mRNA and protein were were determined by Real-time quantitative PCR and Western blotting,respectively.Cell adhesion was exmined by methyl thiazol tetrazolium (MTT) assay,and cell invasion was evaluated by boyden chamber,respectiv ely.The matrix metalloproteinase-13 (MMP-13) were examined by enzyme linked immunosorbent assay (ELISA) assay.Results The results from FQ-PCR showed that the TDGF-1 mRNA of A-375,C-918 and M14 cell line is 0.589 ±0.081,0.712 ±0.065,and 1.517 ±0.085.After M14 cell line was transfected by TDGF-1 siRNA,the results of the MTT assay showed that the A Values of Con-A,Con-B,3.125 nmol/L,6.250 nmol/L,and 12.500 nmol/L siRNA were 0.89 ±0.15,0.85 ±0.12,0.62 ±0.09,0.51 ±0.08,0.33 ± 0.06,respectively (P ＜ 0.05) ; the results of Boyden assay showed that the membrane cell number of Con-A,Con-B,3.125 nmol/L siRNA,6.250 nmol/L siRNA,and 12.500 nmol/L siRNA were 37.8 ± 1.6,36.9±1.5,21.8±1.2,11.6±0.8,and 5.6±0.5,respectively (P＜0.05).The results from ELISA showed that urokinase-type plasminogen activator (uPA) content of Con-A,Con-B,3.125 nmol/L siRNA,6.250 nmol/L siRNA,and 12.500 nmol/L siRNA were (85.2 ± 1.8),(84.8 ± 1.5),(51.6 ± 1.2),(35.6 ± 0.8),(17.8 ± 0.6) ng/L.Conclusion TDGF-1 gene might play an important role in adhesion and invasion of human melanoma cancer cell.The siRNA targeted TDGF-1 could effectively inhibit adhesion and invasion of human melanoma cancer cell through downregulation uPA. Key words: Melanoma ;  Teratocarcinoma-derived growth factor-1 ;  Adhesion ;  Invasion ; Urokinase-type plasminogen activator