Oxidative stress stimulates proliferation and invasiveness of hepatic stellate cells via a MMP2-mediated mechanism.
Experimental evidence indicates that reactive oxygen species (ROS) are involved in the development of hepatic fibrosis; they induce hepatic stellate cells (HSC) proliferation and collagen synthesis. To address the role of matrix metalloproteinase (MMP)-2 in promoting HSC proliferation during hepatic injury, we investigated whether oxidative stress modulates the growth and invasiveness of HSC by influencing MMP-2 activation. Cell invasiveness and proliferation, which were studied using Boyden chambers and by counting cells under a microscope, were evaluated after treatment with a superoxide-producing system, xanthine plus xanthine oxidase (X/XO), in the presence or absence of antioxidants and MMP inhibitors. Expression and activation of MMP-2 were evaluated via gel zymography, immunoassay, and ribonuclease protection assay. The addition of X/XO induced proliferation and invasiveness of human HSC in a dose-dependent manner. The addition of antioxidants as well as MMP-2-specific inhibitors impaired these phenomena. X/XO treatment increased MMP-2 expression and secretion appreciably and significantly induced members of its activation complex, specifically membrane-type 1 MMP and tissue inhibitor metalloproteinase 2. To study the intracellular signaling pathways involved in X/XO-induced MMP-2 expression, we evaluated the effects of different kinase inhibitors. The inhibition of extracellular signal-regulated kinase 1/2 (ERK1/2) and phosphatidyl inositol 3-kinase (PI3K) abrogated X/XO-elicited MMP-2 upregulation and completely prevented X/XO-induced growth and invasiveness of HSC. In conclusion, our findings suggest that MMP-2 is required for the mitogenic and proinvasive effects of ROS on HSC and demonstrate that ERK1/2 and PI3K are the main signals involved in ROS-mediated MMP-2 expression.
Galli
Andrea
A
Gastroenterology Unit, Department of Clinical Pathophysiology, University of Florence, Florence, Italy. a.galli@dfc.unifi.it
Svegliati-Baroni
Gianluca
G
Ceni
Elisabetta
E
Milani
Stefano
S
Ridolfi
Francesco
F
Salzano
Renata
R
Tarocchi
Mirko
M
Grappone
Cecilia
C
Pellegrini
Giulia
G
Benedetti
Antonio
A
Surrenti
Calogero
C
Casini
Alessandro
A
D002455:Cell Division / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D022781:Hepatocytes / Q000166:cytology* / Q000201:enzymology*; D006801:Humans; D008103:Liver Cirrhosis / Q000378:metabolism / Q000473:pathology; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism*; D053504:Matrix Metalloproteinases, Membrane-Associated; D008666:Metalloendopeptidases / Q000378:metabolism; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D018384:Oxidative Stress / Q000187:drug effects / Q000502:physiology*; D038762:Ribosomal Protein S6 Kinases, 70-kDa / Q000378:metabolism; D019716:Tissue Inhibitor of Metalloproteinase-2 / Q000378:metabolism; D019820:Xanthine / Q000494:pharmacology; D014969:Xanthine Oxidase / Q000494:pharmacology
D019716:Tissue Inhibitor of Metalloproteinase-2; D019820:Xanthine; D014969:Xanthine Oxidase; D038762:Ribosomal Protein S6 Kinases, 70-kDa; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D053504:Matrix Metalloproteinases, Membrane-Associated; D008666:Metalloendopeptidases; D020778:Matrix Metalloproteinase 2