Altered lipid metabolism in Hfe-knockout mice promotes severe NAFLD and early fibrosis

Abstract

Tan TC, Crawford DH, Jaskowski LA, Murphy TM, Heritage ML, Subramaniam VN, Clouston AD, Anderson GJ, Fletcher LM. Altered lipid metabolism in Hfe-knockout mice promotes severe NAFLD and early fibrosis. Am J Physiol Gastrointest Liver Physiol 301: G865-G876, 2011. First published August 4, 2011; doi:10.1152/ajpgi.00150.2011.-The HFE protein plays a crucial role in the control of cellular iron homeostasis. Steatosis is commonly observed in HFE-related iron-overload disorders, and current evidence suggests a causal link between iron and steatosis. Here, we investigated the potential contribution of HFE mutations to hepatic lipid metabolism and its role in the pathogenesis of nonalcoholic fatty liver disease. Wild-type (WT) and Hfe knockout mice (Hfe Ϫ/Ϫ ) were fed either standard chow, a monounsaturated low fat, or a high-fat, high-carbohydrate diet (HFD) and assessed for liver injury, body iron status, and markers of lipid metabolism. Despite hepatic iron concentrations and body weights similar to WT controls, Hfe Ϫ/Ϫ mice fed the HFD developed severe hypoxia-related steatohepatitis, Tnf-␣ activation, and mitochondrial respiratory complex and antioxidant dysfunction with early fibrogenesis. These features were associated with an upregulation in the expression of genes involved in intracellular lipid synthesis and trafficking, while transcripts for mitochondrial fatty acid ␤-oxidation and adiponectin signaling-related genes were significantly attenuated. In contrast, HFD-fed WT mice developed bland steatosis only, with no inflammation or fibrosis and no upregulation of lipogenesis-related genes. A HFD led to reduced hepatic iron in Hfe Ϫ/Ϫ mice compared with chow-fed mice, despite higher serum iron, decreased hepcidin expression, and increased duodenal ferroportin mRNA. In conclusion, our results demonstrate that Hfe Ϫ/Ϫ mice show defective hepatic-intestinal iron and lipid signaling, which predispose them toward diet-induced hepatic lipotoxicity, accompanied by an accelerated progression of injury to fibrosis.

Excess iron modulates endoplasmic reticulum stress-associated pathways in a mouse model of alcohol and high-fat diet-induced liver injury

“…Serum alanine aminotransferase (ALT), iron, cholesterol, and triglyceride measurements were performed as previously described. 19 Quantitative Real-Time PCR Analysis Total cellular RNA was isolated from hepatic tissue using the TRIzol reagent (Invitrogen, Melbourne, Australia) according to the manufacturer's instructions. Reverse transcription and quantitative RT-PCR (Applied Biosystems, Brisbane, Australia) were performed as previously described.…”

“…Reverse transcription and quantitative RT-PCR (Applied Biosystems, Brisbane, Australia) were performed as previously described. 19 The primer sequences used for the experiments are outlined in Supplementary Table S3. Gene expression was normalized to the internal standard basic transcription factor 3 (Btf3).…”

“…19 Immunoblotting was performed with specific primary antibodies (Supplementary Table S4). Blocking and antibody dilutions were performed in 10% skim milk in Tris-buffered saline with 0.1% Tween.…”

“…19,23 Complex 1 activity was measured as described but at 30 1C and normalized to citrate synthase activity. 23 Hepatic Iron Content Hepatic non-heme iron concentration (HIC) was measured as previously described.…”

Hfe-associated steatohepatitis: expression profiling and identifying the molecular basis of liver injury

“…Our research group has observed similar evidence in Hfe -/-mice fed a high calorie diet (HCD). This study showed a significant increase in serum alanine amino transferase (ALT), a marker of liver injury, and increased hepatic inflammatory and fibrogenic gene expression in Hfe -/-mice fed a HCD which develop steatohepatitis and early fibrosis (10). The mechanisms underlying the development of more severe injury are however not fully understood and require further investigation.…”