Inhibition Of Gastric Cancer Cells Invasion And Metastasis By Zileuton In Viro

Objective1.To investigate the effects of Darbufelone on the growth, invasion, metastasis and inducing endothelial cell to form neovascularization of human gastric cancer cells SGC-7901 in vitro.2. To explore the possible mechanisms that Darbufelone affect the invasion and metastasis of SGC-7901 cells.MethodsTo investigate the growth and adhesion ability by MTT.2.To observe invasion and vertical metastasis ability in Transwell Chamber membrane invasion culture system3.To observe the transverse metastasis ability by wound-healing assays.4.To detect the capability of SGC-7901 cell condition medium inducing human umbilical vein endothelial cells ECV304 to form blood vessel.5.To analyse the mRNA expression of OPN and MMP-2 by RT-PCR .6.To analyse the protein expression of OPN and MMP-2 by cel1-immunohistochemistry and Western blotting.Results1. After treatment with Darbufelone at concentration of 1.5×10-5 mol/L for 72 h,the growth of SGC-7901 cells were decreased ( P<0.05).2. Cell adherence assays display,after treatment with Darbufelone at concentration of 1.5×10-5 mol/L for 72 h, the 30,60,90,120 min adhesion rate of SGC-7901 cells were (27.10±3.44)%,(46.36±4.52) %,(50.37±7.90)%,(58.01±9.22)%, which were obviously declined compared with the control groups [(34.99±2.12)%, (60.60±4.89)%, (70.65±5.20)%, (84.62±7.52)%] (P<0.05), respectively.3. In vitro invasive assays and cell migration reveal,after treatment with Darbufelone at concentration of 1.5×10-5 mol/L for 72 h, the number of invasion and vertical metastasis SGC-7901 cells were 50.00±3.67 and 93.00±6.56 ,which were significantly decreased compared with the control groups (82.00±7.14, 143.00±9.82)(P<0.01).4.Wound-healing assays show,after treatment with Darbufelone at concentration of 1.5×10-5 mol/L for 72 h, the 24,48 h transverse metastasis length of SGC-7901 cells were obviously shorter than the controls.5. The vessel number and the length of vessel of ECV304 [(82.70±4.58), (172.20±6.41)μm],induced by SGC-7901 cells after treatment with Darbufelone at concentration of 1.5×10-5 mol/L for 72 h, were significantly less than those in controls [142.00±5.87, (214.60±9.44)μm](P<0.01).6. RT-PCR display ,after treatment with Darbufelone at concentration of 1.5×10-5 mol/L for 72 h, the mRNA expression of OPN and MMP-2 (0.34 ±0.03, 0.33±0.04) were obviously lower than those in controls (0.54±0.01, 0.57±0.09)(P<0.05 ).7. Cel1- immunohistochemistry and Western blotting show,after treatment with Darbufelone at concentration of 1.5×10-5 mol/L for 72 h, the protein expression of OPN and MMP-2 were both declined.1)Cel1-immunohistochemistry show that the AOD of OPN and MMP-2 in Darbufelone treated groups(0.18±0.01,0.26±0.02) were obviously lower than those in controls (0.26±0.03,0.33±0.03) (P<0.05 ).2)Western blotting show that the OPN expression in Darbufelone treated group (0.27±0.02) was obviously lower than the controls (0.44±0.06) (P<0.01 );the MMP-2 expression in Darbufelone treated group(0.35±0.03) was also lower than the controls (0.48±0.06) (P<0.05 ).Conclusion1.Darbufelone can inhibit the growth, invasion, metastasis and inducing endothelial cell to form neovascularization of human gastric cancer cells SGC-7901 in vitro.2. Darbufelone inhibit the invasion and metastasis of SGC-7901 cells probably via down-regulation of OPN and MMP-2.