Basement membrane proteins play an active role in the invasive process of human hepatocellular carcinoma cells with high metastasis potential.
OBJECTIVE
Cell-matrix adhesive interaction has an important role in the invasive process of tumor cells, and integrins are the major receptors mediating cell-matrix adhesion. The current study is to investigate the modulation of basement membrane (BM) proteins, especially collagen IV (C IV), laminin (LN), and fibronectin (FN) in the invasive processes of human hepatocellular carcinoma (HCC) cells in vitro, and to reveal the roles of beta1 integrins and RGD-containing oligopeptide in the cell-matrix interaction.


METHODS
Static adhesion assay was performed to study the rates of adhesion of MHCC97-H cells, treated or untreated with anti-beta1 (2 microg ml(-1)) and GRGDS, to C IV (50 microg ml(-1)), LN (50 microg ml(-1)) or FN (50 microg ml(-1)). Gelatin zymography was used to detect the secretion of MMPs in the conditioned medium of MHCC97-H cells incubated 24 h by C IV, LN or FN, and interactions with anti-beta1 and GRGDS. Transwell chamber assay was used to investigate the influence of C IV, LN or FN, interacting with anti-beta1 and GRGDS, on the cellular mobility of MHCC97-H cells.


RESULTS
Compared with blank control group, MHCC97-H cells showed significantly higher rates of adhesion to C IV, LN, and FN. Pretreatment with anti-beta1 could suppress adhesion to C IV, LN or FN, but GRGDS inhibited adhesion to FN (P<0.05) only. LN and FN could stimulate the secretion of MMPs by MHCC97-H cells cultured in vitro, especially MMP-9 and its activated type. Treatment with anti-beta1 could partly counteract the effects of LN and FN. GRGDS could prominently induce the secretion of MMPs, but the effect could be inhibited by pretreatment of anti-beta1. The results of Transwell chamber assay showed that LN, FN, and GRGDS could increase the number of tumor cells penetrating the microporous membrane, but the data of C IV did not reach significance. The effects were partly counteracted by anti-beta1.


CONCLUSIONS
BM proteins play an active role in the invasive process of human hepatocellular carcinoma cells. Integrin beta1 is an important molecule which mediates the cell-matrix adhesive interaction of tumor cells. RGD-containing peptides competitively combine with the binding site of integrin beta1, and the effects of FN are RGD sequence-dependent.
Liver Cancer Institute & Zhongshan Hospital, Fudan University, 136 Yixueyuan Road, 200032 Shanghai, P.R. China. zytang@scrap.stc.sh.cn
Bo
B
Tian
Yan
Y
Li
Xue-Ning
XN
Ji
Jie
J
Chen
Qiong
Q
Xue
Sheng-Long
SL
Ye
Yin-Kun
YK
Liu
Zhao-You
ZY
Tang
D001485:Basement Membrane / Q000502:physiology*; D006528:Carcinoma, Hepatocellular / Q000503:physiopathology*; D002448:Cell Adhesion; D024141:Collagen Type IV / Q000502:physiology*; D016326:Extracellular Matrix Proteins / Q000502:physiology; D005353:Fibronectins / Q000502:physiology*; D006801:Humans; D039641:Integrin beta Chains / Q000502:physiology; D007797:Laminin / Q000502:physiology*; D008113:Liver Neoplasms / Q000503:physiopathology*; D009361:Neoplasm Invasiveness / Q000503:physiopathology*; D009362:Neoplasm Metastasis / Q000503:physiopathology; D016212:Transforming Growth Factor beta / Q000502:physiology; D014407:Tumor Cells, Cultured; D000099222:betaIG-H3 Protein
D024141:Collagen Type IV; D016326:Extracellular Matrix Proteins; D005353:Fibronectins; D039641:Integrin beta Chains; D007797:Laminin; D016212:Transforming Growth Factor beta; D000099222:betaIG-H3 Protein