Increased susceptibility to exacerbated liver injury in hypercholesterolemic ApoE-deficient mice: potential involvement of oxysterols

Ferre N, Martinez-Clemente M, Lopez-Parra M, Gonzalez-Periz A, Horrillo R, Planaguma A, Camps J, Joven J, Tres A, Guardiola F, Bataller R, Arroyo V, Claria J. Increased susceptibility to exacerbated liver injury in hypercholesterolemic ApoE-deficient mice: potential involvement of oxysterols. Am J Physiol Gastrointest Liver Physiol 296: G553-G562, 2009. First published January 8, 2009; doi:10.1152/ajpgi.00547.2007.-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 hypercholesterolemia on the severity of carbon tetrachloride (CCl4)-induced liver injury. Under baseline conditions, hypercholesterolemic ApoE(-/-) mice showed increased hepatic oxidative stress (SOD activity/4-hydroxy-2-nonenal immunostaining) and higher hepatic TGF-beta 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 alpha-smooth muscle actin immunohistochemistry) and more severe liver injury [alanine aminotransferase (ALT) and aspartate aminotransferase] 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 P-450 expression was unchanged in ApoE(-/-) mice. 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-kappa B-independent mechanism and upregulated TIMP-1 expression. In macrophages, oxysterols increased TGF-beta 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.