Submitted on November 23, 2007
Revised on November 24, 2008
Accepted on December 22, 2008
Increased Susceptibility to Exacerbated Liver Injury in Hypercholesterolemic ApoE-deficient mice. Potential Involvement of Oxysterols
Natàlia Ferré1, Marcos Martínez-Clemente1, Marta López-Parra1, Ana González-Périz2, Raquel Horrillo3, Anna Planagumà3, Jordi Camps4, Jorge Joven4, Alba Tres5, Francesc Guardiola5, Ramón Bataller6, Vicente Arroyo6, and Joan Claria3*
1 Hospital Clinic, University of Barcelona
2 HHospital Clinic-University of Barcelona
3 Hospital Clinic-University of Barcelona
4 Universitat Rovira i Virgili
5 University of Barcelona
6 Hospital Clínic, University of Barcelona
* To whom correspondence should be addressed. E-mail: jclaria{at}clinic.ub.es.
The contribution of metabolic factors to the severity of liver disease is not completely understood. In this study, apolipoprotein-E deficient (ApoE-/-) mice were evaluated to define potential effects of hypercholesteromia on the severity of carbon tetrachloride (CCl4)-induced liver injury. Under baseline conditions, hypercholesterolemic ApoE-/- mice showed increased hepatic oxidative stress (SOD activity/4-HNE immunostaining) and higher hepatic TGF-
1, MCP-1 and TIMP-1 expression than wild-type control mice. After CCl4 challenge, ApoE-/- mice exhibited exacerbated steatosis (oil red-O staining), necroinflammation (hematoxylin/eosin staining), macrophage infiltration (F4/80 immunohistochemistry) and fibrosis (Sirius red staining and 
-SMA immunohistochemistry) and more severe liver injury (ALT and AST) than wild-type controls. Direct correlations were identified between serum cholesterol and hepatic steatosis, fibrosis and ALT levels. These changes did not reflect the usual progression of the disease in ApoE-/- mice, since exacerbated liver injury was not present in untreated age-paired ApoE-/- mice. Moreover, hepatic cytochrome P450 expression was unchanged in ApoE-/- mice. In order to explore potential mechanisms, cell types relevant to liver pathophysiology were exposed to selected cholesterol-oxidized products. Incubation of hepatocytes with a mixture of oxysterols representative of those detected by GC-MS in livers from ApoE-/- mice resulted in a concentration-dependent increase in total lipoperoxides and SOD activity. In hepatic stellate cells, oxysterols increased IL-8 secretion through a NF-
B-independent mechanism and up-regulated TIMP-1 expression. In macrophages, oxysterols increased TGF-1 secretion and MCP-1 expression in a concentration-dependent manner. Oxysterols did not compromise cell viability. Taken together, these findings demonstrate that hypercholesterolemic mice are sensitized to liver injury and that cholesterol-derived products (i.e. oxysterols) are able to induce proinflammatory and profibrogenic mechanisms in liver cells.
