Intervention Effect Of Ursolic Acid And Derivatives On Adhesion Of Hepatoma Carcinoma Cells To Vascular Endothelium And Underlying Mechanisms

Objective:The aim of this study is to observe the effect of ursolic acid and its novel derivative US597(UA/US597)on adhesion ability of Human hepatoma cell in vitro,we also examined the effect of UA/US597 on the metastases of melanoma in vivo and the influence on the gene and protein expression and detected its anti-metastasis molecule mechanisms.Methods:The cell proliferation and toxicity were determined using MTT colorimetric assay.After the human liver cancer cell HepG2,MHCC-97H,MHCC-97L,normal human hepatocyte cell L02 and human umbilical vein endothelial cell were treated with the UA/US597 for 24h,high-efficiency compound concentration was selected.The cell adhesion,wound healing and transwell invasion assays in vitro were performed to evaluate the effects of UA/US597 on the adhesion,migration and invasion of HepG2 cells.B16-F10 cells experimental lung metastasis model was used to evaluate the anti-metastatic activity of UA/US597 in vivo.Expression of genes and proteins were determined by quantitative real-time reverse transcription-PCR,flow cytometry,western blotting,microarray hybridization assays and immunohistochemical staining,respectively.Results:Our results showed that UA/US597 significantly inhibited HepG2 cells proliferation.Compared with control group,at safe and effective concentrations(0.2-5μM),UA/US597 significantly inhibits HepG2 cells adhesion,migration and invasion in a dose-dependent manner in vitro.Both UA and US597,80 mg/kg for 28 days,could decline lung tumor node in the B16-F10/C57 BL/6 mouse melanoma lung metastasis model and the inhibitory rates were 14.45%and 60.24%,respectively.We did not detect side effects with UA/US597 in mice such as weight loss,viscera tissues toxicity and blood cell abnormalities.Meanwhile,the life span of these animals was significantly increased by UA/US597 treatment.Further studies showed that UA/US597 significantly inhibited the expression of ICAM-1,VCAM-1 and E-selectin in HUVECs at both the protein and RNA levels.In addition,this effect was accompanied by inhibition of focal adhesion signaling pathway including alterations in integrin α6β1,FAK,Src,paxillin and PTEN.Conclusion:Low dose range of UA/US597 is highly effective in suppressing cancer metastasis,including cell adhesion,invasion and migration,without apparent side effects both in vitro and in vivo.The mechanism probably attributes to its down-regulation of cell adhesion molecules(ICAM-1、VCAM-1 and E-selectin)expression in HUVECs and modulate expression of integrin α6β1 within focal adhesion signaling pathway in HepG2 cells.Thus UA/US597 intervenes the adhesion of hepatoma carcinoma cells to vascular endothelium and exerts significant anti-metastatic activity.