Fatty Acid Incorporation in the Muscle, Oxidative Markers, Lipid Peroxidation and PPAR-α and SREBP-2 Expression of Zebrafish Fed Linseed Oil and Clove Leaf Essential Oil.

The objective of this study is to assess, in zebrafish, the effects of combining linseed oil (LO) and clove leaf essential oil (CLEO) on the incorporation of fatty acids in the muscle, oxidative markers, lipid peroxidation and expression of the PPAR-α (Peroxisome Proliferator-Activated Receptor-α) and the SREBP-2 (Sterol Regulatory Element Binding Protein-2) genes. Six diets were prepared, containing combinations of LO (3, 6 and 9%) and CLEO (0.5 and 1%): 3% LO + 0.5% CLEO; 3% LO + 1% CLEO; 6% LO + 0.5% CLEO; 6% LO + 1% CLEO; 9% LO + 0.5% CLEO; 9% LO + 1% CLEO. Results showed increase in the incorporation of n-3 fatty acids in the muscle concomitantly with the addition of LO and CLEO. The activities of superoxide dismutase and catalase were reduced and the glutathione content had increased. Lipid peroxidation was lower in the treatment with 1% CLEO, regardless of LO content. The expression of the PPAR-α and the SREBP-2 genes was higher in animals fed 9% LO + 0.5% CLEO. Therefore, for a greater incorporation and protection against the oxidative damages of n-3 fatty acids, a combined use of 9% LO with 0.5% CLEO is recommended for zebrafish.

UBC12-mediated SREBP-1 neddylation worsens metastatic tumor prognosis.

Activation of sterol regulatory element-binding protein 1 (SREBP-1), a master lipogenic transcription factor, is associated with cancer metabolism and metabolic disorders. Neddylation, the process of adding NEDD8 to its substrate, contributes to diverse biological processes. Here, we identified SREBP-1 as a substrate for neddylation by UBC12 and explored its impact on tumor aggressiveness. In cell-based assays, SREBP-1 neddylation prolonged SREBP-1 stability with a decrease in ubiquitination. Consequently, NEDD8 overexpression facilitated proliferation, migration, and invasion of SK-Hep1 liver tumor cells. MLN4924 (an inhibitor of the NEDD8-activating enzyme-E1) treatment or UBC12 knockdown prevented SREBP-1 neddylation and tumor cell phenotype change. This effect was corroborated in an in vivo xenograft model. In human specimens, SREBP-1, UBC12, and NEDD8 were all upregulated in hepatocellular carcinoma (HCC) compared to nontumorous regions. Moreover, SREBP-1 levels positively correlated with UBC12. In GEO database analyses, SREBP-1 levels were greater in metastatic HCC samples accompanying UBC12 upregulation. In HCC analysis, tumoral SREBP-1 and UBC12 levels discriminated overall patient survival rates. Additionally, MLN4924 treatment destabilized SREBP-1 in MDA-MB-231 breast cancer cells and in the tumor cell xenograft. SREBP-1 and UBC12 were also highly expressed in human breast cancer tissues. Moreover, most breast cancers with lymph node metastasis displayed predominant SREBP-1 and UBC12 expressions, which compromised overall patient survival rates. In summary, SREBP-1 is neddylated by UBC12, which may contribute to HCC and breast cancer aggressiveness through SREBP-1 stabilization, and these events can be intervented by MLN4924 therapy. Our findings may also provide potential reliable prognostic markers for tumor metastasis.

Estrogen-related receptor γ controls sterol regulatory element-binding protein-1c expression and alcoholic fatty liver.

Although SREBP-1c regulates key enzymes required for hepatic de novo lipogenesis, the mechanisms underlying transcriptional regulation of SREBP-1c in pathogenesis of alcoholic fatty liver is still incompletely understood. In this study, we investigated the role of ERRγ in alcohol-mediated hepatic lipogenesis and examined the possibility to ameliorate alcoholic fatty liver through its inverse agonist. Hepatic ERRγ and SREBP-1c expression was increased by alcohol-mediated activation of CB1 receptor signaling. Deletion and mutation analyses of the Srebp-1c gene promoter showed that ERRγ directly regulates Srebp-1c gene transcription via binding to an ERR-response element. Overexpression of ERRγ significantly induced SREBP-1c expression and fat accumulation in liver of mice, which were blocked in Srebp-1c-knockout hepatocytes. Conversely, liver-specific ablation of ERRγ gene expression attenuated alcohol-mediated induction of SREBP-1c expression. Finally, an ERRγ inverse agonist, GSK5182, significantly ameliorates fatty liver disease in chronically alcohol-fed mice through inhibition of SREBP-1c-mediated fat accumulation. ERRγ mediates alcohol-induced hepatic lipogenesis by upregulating SREBP-1c expression, which can be blunted by the inverse agonist for ERRγ, which may be an attractive therapeutic strategy for the treatment of alcoholic fatty liver disease in human.

Insulin suppresses the AMPK signaling pathway to regulate lipid metabolism in primary cultured hepatocytes of dairy cows.

Dairy cows with type II ketosis display hepatic fat accumulation and hyperinsulinemia, but the underlying mechanism is not completely clear. This study aimed to clarify the regulation of lipid metabolism by insulin in cow hepatocytes. In vitro, cow hepatocytes were treated with 0, 1, 10, or 100 nm insulin in the presence or absence of AICAR (an AMP-activated protein kinase alpha (AMPKα) activator). The results showed that insulin decreased AMPKα phosphorylation. This inactivation of AMPKα increased the gene and protein expression levels of carbohydrate responsive element-binding protein (ChREBP) and sterol regulatory element-binding protein-1c (SREBP-1c), which downregulated the expression of lipogenic genes, thereby decreasing lipid biosynthesis. Furthermore, AMPKα inactivation decreased the gene and protein expression levels of peroxisome proliferator-activated receptor-α (PPARα), which upregulated the expression of lipid oxidation genes, thereby increasing lipid oxidation. In addition, insulin decreased the very low density lipoprotein (VLDL) assembly. Consequently, triglyceride content was significantly increased in insulin treated hepatocytes. Activation of AMPKα induced by AICAR could reverse the effect of insulin on PPARα, SREBP-1c, and ChREBP, thereby decreasing triglyceride content. These results indicate that insulin inhibits the AMPKα signaling pathway to increase lipid synthesis and decrease lipid oxidation and VLDL assembly in cow hepatocytes, thereby inducing TG accumulation. This mechanism could partly explain the causal relationship between hepatic fat accumulation and hyperinsulinemia in dairy cows with type II ketosis.

Non-alcoholic fatty liver disease in mice with heterozygous mutation in TMED2.

The transmembrane emp24 domain/p24 (TMED) family are essential components of the vesicular transport machinery. Members of the TMED family serve as cargo receptors implicated in selection and packaging of endoplasmic reticulum (ER) luminal proteins into coatomer (COP) II coated vesicles for anterograde transport to the Golgi. Deletion or mutations of Tmed genes in yeast and Drosophila results in ER-stress and activation of the unfolded protein response (UPR). The UPR leads to expression of genes and proteins important for expanding the folding capacity of the ER, degrading misfolded proteins, and reducing the load of new proteins entering the ER. The UPR is activated in non-alcoholic fatty liver disease (NAFLD) in human and mouse and may contribute to the development and the progression of NAFLD. Tmed2, the sole member of the vertebrate Tmed ß subfamily, exhibits tissue and temporal specific patterns of expression in embryos and developing placenta but is ubiquitously expressed in all adult organs. We previously identified a single point mutation, the 99J mutation, in the signal sequence of Tmed2 in an N-ethyl-N-nitrosourea (ENU) mutagenesis screen. Histological and molecular analysis of livers from heterozygous mice carrying the 99J mutation, Tmed299J/+, revealed a requirement for TMED2 in liver health. We show that Tmed299J/+ mice had decreased levels of TMED2 and TMED10, dilated endoplasmic reticulum membrane, and increased phosphorylation of eIF2α, indicating ER-stress and activation of the UPR. Increased expression of Srebp1a and 2 at the newborn stage and increased incidence of NAFLD were also found in Tmed299J/+ mice. Our data establishes Tmed299J/+ mice as a novel mouse model for NAFLD and supports a role for TMED2 in liver health.

Abnormal lipid/lipoprotein metabolism and high plasma testosterone levels in male but not female aromatase-knockout mice.

Sex steroid hormones, such as estrogen and testosterone, are believed to play important roles in lipid metabolism. To elucidate the effects of estrogen depletion on lipid metabolism in male and female mice, we used aromatase-knockout (ArKO) mice, in which Cyp19 gene disruption prevented estrogen synthesis in vivo. These mice were divided into the following 4 groups: male and female ArKO mice and male and female wild-type (WT) mice. These mice were fed a normal-fat diet (13.6% fat) ad libitum. At 159 days after birth, the mice were tested for liver and plasma lipid content and hepatic hormone receptor- and lipid/lipoprotein metabolism-related gene expression. Interestingly, we found that hepatic steatosis was accompanied by markedly elevated plasma testosterone levels in male ArKO mice but not in female ArKO mice. Plasma lipoprotein profiles exhibited concurrent decreases in LDL- and small dense LDL-triglyceride (TG) levels in male ArKO mice. Moreover, male mice, but not female mice, exhibited marked elevations in androgen receptor (AR), sterol regulatory element-binding protein 1 (SREBP1), and CD36 expression. These results strongly suggest that Cyp19 gene disruption, which induces a sexually dimorphic response and high plasma testosterone levels in male mice, also induces hepatic steatosis.

Bioactive quinone derivatives from the marine brown alga Sargassum thunbergii induce anti-adipogenic and pro-osteoblastogenic activities.

BACKGROUND: Health problems related to the lack of bone formation are a major problem for ageing populations in the modern world. As a part of the ongoing trend to develop natural substances that attenuate bone loss in osteoporosis, the effects of the edible brown alga Sargassum thunbergii and its active contents on adipogenic differentiation in 3T3-L1 fibroblasts and osteoblast differentiation in MC3T3-E1 pre-osteoblasts were evaluated. RESULTS: Treatment with S. thunbergii significantly reduced lipid accumulation and expression of adipogenic differentiation markers such as peroxisome proliferator-activated receptor γ, CCAAT/enhancer-binding protein α and sterol regulatory element binding protein 1c. In addition, S. thunbergii successfully enhanced osteoblast differentiation as indicated by increased alkaline phosphatase activity along raised levels of osteoblastogenesis indicators, namely bone morphogenetic protein-2, osteocalcin and collagen type I. Two compounds, sargaquinoic and sargahydroquinoic acid, were isolated from active extract and shown to be active by means of osteogenesis inducement. CONCLUSION: S. thunbergii could be a source for functional food ingredients for improved treatment of osteoporosis and obesity.

Protective effect of thymol on high fat diet induced diabetic nephropathy in C57BL/6J mice.

Obesity is one of several factors implicated in chronic kidney disease (CKD). Thymol, a monoterpene phenolic compound found in the oils of thyme with multiple biological properties especially antidiabetic activity. The present study was undertaken to evaluate the thymol against diabetic nephropathy by high fat diet (HFD)-induced diabetic C57BL/6J mice. After 10 weeks of continuous dietary intervention, HFD (fat- 35.2%) to mice presented characteristic features of progressive nephropathy by significant increased in kidney weight, blood, and urinary parameters, glomerulosclerosis, oxidative stress, hyperlipidemia and subsequent renal injuries. After intragastric administration of thymol (40 mg/kg BW) daily for the subsequent 5 weeks significantly decreased the blood, urinary parameters and kidney weight. Thymol inhibited the activation of transforming growth factor-ß1 (TGF-ß1) and vascular endothelial growth factor (VEGF). Also, significantly increased the antioxidants and suppresses the lipid peroxidation markers in erythrocytes and kidney tissue compared to the diabetic mice. Thymol downregulated the expression level of sterol regulatory element binding protein-1c (SREBP-1c) and reduced the lipid accumulation in renal. Histopathological study of kidney tissues showed that extracellular mesangial matrix expansion, glomerulosclerosis in diabetic mice were suppressed by thymol. Further, our results indicate that administration of thymol afforded remarkable protection against HFD-induced diabetic nephropathy.

Optimized Rapeseed Oils Rich in Endogenous Micronutrients Protect High Fat Diet Fed Rats from Hepatic Lipid Accumulation and Oxidative Stress.

UNLABELLED: Micronutrients in rapeseed exert a potential benefit to hepatoprotection, but most of them are lost during the conventional refining processing. Thus some processing technologies have been optimized to improve micronutrient retention in oil. The aim of this study is to assess whether optimized rapeseed oils (OROs) have positive effects on hepatic lipid accumulation and oxidative stress induced by a high-fat diet. METHODS: Rats received experiment diets containing 20% fat and refined rapeseed oil or OROs obtained with various processing technologies as lipid source. After 10 weeks of treatment, liver was assayed for lipid accumulation and oxidative stress. RESULTS: All OROs reduced hepatic triglyceride contents. Microwave pretreatment-cold pressing oil (MPCPO) which had the highest micronutrients contents also reduced hepatic cholesterol level. MPCPO significantly decreased hepatic sterol regulatory element-binding transcription factor 1 (SREBP1) but increased peroxisome proliferator activated receptor α (PPARα) expressions, and as a result, MPCPO significantly suppressed acetyl CoA carboxylase and induced carnitine palmitoyl transferase-1 and acyl CoA oxidase expression. Hepatic catalase (CAT) and glutathione peroxidase (GPx) activities as well as reduced glutathione (GSH) contents remarkably increased and lipid peroxidation levels decreased in parallel with the increase of micronutrients. CONCLUSION: OROs had the ability to reduce excessive hepatic fat accumulation and oxidative stress, which indicated that OROs might contribute to ameliorating nonalcoholic fatty liver induced by high-fat diet.

Amla prevents fructose-induced hepatic steatosis in ovariectomized rats: role of liver FXR and LXRα.

OBJECTIVES: Increased fructose consumption causes dyslipidemia and fatty liver in postmenopausal women, both independent risk factors for cardiovascular disease. This study explored the potential mechanisms by which amla (Emblica officinalis) reduced hypercholesterolemia and hypertriglyceridemia and prevented fatty liver in a fructose-fed, ovariectomized rat model of menopause. METHODS: Sham-operated and ovariectomized rats were put on a chow or high fructose diet. They were further divided into groups with or without amla. After 18 weeks of treatment, livers were harvested and subjected to Western blot and histological analyses. RESULTS: In all groups, amla increased the protein expression of liver farnesoid X receptor (FXR) and liver X receptor (LXR), key proteins involved in lipid metabolism. Fructose-fed rats developed fatty liver and amla prevented this. Here amla produced an exceptional rise in LXR and insulin-induced gene-2 (Insig-2) which prevented the maturation of sterol regulatory element-binding protein-1 and steroyl CoA desaturase-1, responsible for triglyceride synthesis. Amla also increased the protein expression of ATP binding cassette transporter A1 (ABCA1), involved in high density lipoprotein (HDL) synthesis as well as low density lipoprotein receptor (LDLR) responsible for uptake of LDL cholesterol. Besides this, amla increased the protein expression of peroxisome proliferator activated receptor α (PPARα) involved in ß oxidation of fatty acids. CONCLUSIONS: Amla increased the protein expression of liver FXR, LXRα, PPARα and their downstream proteins Insig-2, ABCA1 and LDLR. This property of amla to modulate some of the key proteins involved in lipid metabolism promises its usefulness as a preventive agent for dyslipidemia and hepatic steatosis.

SREBP-1 has a prognostic role and contributes to invasion and metastasis in human hepatocellular carcinoma.

Sterol regulatory element-binding protein 1 (SREBP-1) is a well-known nuclear transcription factor involved in lipid synthesis. Recent studies have focused on its functions in tumor cell proliferation and apoptosis, but its role in cell migration and invasion, especially in hepatocellular carcinoma (HCC), is still unclear. In this study, we found that the expression of SREBP-1 in HCC tissues was significantly higher than those in matched tumor-adjacent tissues (p < 0.05). SREBP-1 was expressed at significantly higher levels in patients with large tumor size, high histological grade and advanced tumor-node-metastasis (TNM) stage (p < 0.05). The positive expression of SREBP-1 correlated with a worse 3-year overall and disease-free survival of HCC patients (p < 0.05). Additionally, SREBP-1 was an independent factor for predicting both 3-year overall and disease-free survival of HCC patients (p < 0.05). In vitro studies revealed that downregulation of SREBP-1 inhibited cell proliferation and induced apoptosis in both HepG2 and MHCC97L cells (p < 0.05). Furthermore, wound healing and transwell assays showed that SREBP-1 knockdown prominently inhibited cell migration and invasion in both HepG2 and MHCC97L cells (p < 0.05). These results suggest that SREBP-1 may serve as a prognostic marker in HCC and may promote tumor progression by promoting cell growth and metastasis.

RETRACTED: Reversion of hepatic steatosis by exercise training in obese mice: the role of sterol regulatory element-binding protein-1c.

AIM: The dysregulation of regulatory element-binding protein-1c (SREBP-1c) is associated with hepatic steatosis. However, effects of exercise on SREBP-1c protein level in liver have not been investigated. Thus, in this study we investigated if reversion of the hepatic steatosis-induced by exercise training is related with levels of SREBP-1c. MAIN METHODS: Mice were divided into two groups: control lean mice (CT), fed on standard rodent chow, and obese mice (HF), fed on a high-fat diet for 2months. After this period obese mice were divided in two groups: obese mice and obese mice submitted to exercise (HF+EXE). The HF+EXE group performed a running program of 50min per day, 5days per week, for 8weeks. Forty-eight hours after the last exercise session, biochemical, immunoblotting, histology and immunohistochemistry analyses were performed. KEY FINDINGS: Livers of HF mice showed increased SREBP-1c, FAS (Fatty Acid Synthase), SCD1 (Stearoyl-CoA Desaturase1) and CPT1 (Carnitine Palmitoyl Transferase1) protein levels (3.4, 5.0, 2.6 and 2.9 times, respectively), though ACC (Acetyl-CoA Carboxilase) phosphorylation dropped 4.2 times. In livers of HF+EXE, levels of SREBP-1c, FAS, SCDI and CPTI decreased 2.1, 1.9, 1.8, and 2.7 times, respectively), while ACC phosphorylation increased 3.0 times. Lower SREBP-1c protein levels after exercise were confirmed also by immunohistochemistry. Total liver lipids content was higher in HF (2.2 times) when compared to CT, and exercise training reduced it significantly (1.7 times). SIGNIFICANCE: Our study allows concluding that the reduction in SREBP-1c protein levels is associated with steatosis reversion induced by exercise training.

A2b adenosine receptor regulates hyperlipidemia and atherosclerosis.

BACKGROUND: The cAMP-elevating A(2b) adenosine receptor (A(2b)AR) controls inflammation via its expression in bone marrow cells. METHODS AND RESULTS: Atherosclerosis induced by a high-fat diet in apolipoprotein E-deficient mice was more pronounced in the absence of the A(2b)AR. Bone marrow transplantation experiments indicated that A(2b)AR bone marrow cell signals alone were not sufficient to elicit this effect. Intriguingly, liver expression of the A(2b)AR in wild-type mice was vastly augmented by a high-fat diet, raising the possibility that this upregulation is of functional significance. A(2b)AR genetic ablation led to elevated levels of liver and plasma cholesterol and triglycerides and to fatty liver pathology typical of steatosis, assessed by enzymatic assays and analysis of liver sections. Western blotting and quantitative polymerase chain reaction revealed elevated expression of the following molecules in the liver of A(2b)AR-null mice: the transcription factor sterol regulatory element binding protein-1 (SREBP-1) and its 2 downstream targets and regulators of lipogenesis, acetyl CoA carboxylase and fatty acid synthase. Pharmacological activation or inhibition of A(2b)AR in primary hepatocytes confirmed the regulation of SREBP-1 by this receptor. A(2b)AR-mediated changes in cAMP were found to regulate levels of the transcriptionally active form of SREBP-1. Finally, adenovirally mediated restoration of the A(2b)AR in the liver of A(2b)AR-null mice reduced the lipid profile and atherosclerosis. Similarly, in vivo administration of the A(2b)AR ligand BAY 60-6853 in control mice on a high-fat diet reduced the lipid profile and atherosclerosis. CONCLUSION: This study provides the first evidence that the A(2b)AR regulates liver SREBP-1, hyperlipidemia, and atherosclerosis, suggesting that this receptor may be an effective therapeutic target.

Hypoxia stimulates low-density lipoprotein receptor-related protein-1 expression through hypoxia-inducible factor-1α in human vascular smooth muscle cells.

OBJECTIVE: Hypoxia is considered a key factor in the progression of atherosclerotic lesions. Low-density lipoprotein receptor-related protein (LRP1) plays a pivotal role in the vasculature. The aim of this study was to investigate the effect of hypoxia on LRP1 expression and function in vascular smooth muscle cells (VSMC) and the role of hypoxia-inducible factor-α (HIF-1α). METHODS AND RESULTS: Real-time polymerase chain reaction and Western blot analysis demonstrated that hypoxia (1% O(2)) time-dependently induced LRP1 mRNA (maximum levels at 1 to 2 hours) and protein expression (maximum levels at 12 to 24 hours). The delayed hypoxic upregulation of LRP1 protein versus mRNA may be explained by the long half-life of LRP1 protein. Luciferase assays demonstrated that hypoxia and HIF-1α overaccumulation induced LRP1 promoter activity and that 2 consensus hypoxia response element sites located at -1072/-1069 and -695/-692 participate in the induction. Chromatin immunoprecipitation showed the in vivo binding of HIF-1α to LRP1 promoter in hypoxic VSMC. Hypoxia effects on LRP1 protein expression were functionally translated into an increased cholesteryl ester (CE) accumulation from aggregated low-density lipoprotein (agLDL) uptake. The blockade of HIF-1α expression inhibited the upregulatory effect of hypoxia on LRP1 expression and agLDL-derived intracellular CE overaccumulation, suggesting that both LRP1 overexpression and CE overaccumulation in hypoxic vascular cells are dependent on HIF-1α. Immunohistochemical analysis showed the colocalization of LRP1 and HIF-1α in vascular cells of human advanced atherosclerotic plaques. CONCLUSION: Hypoxia upregulates LRP1 expression and agLDL-derived intracellular CE accumulation in human VSMC through HIF-1α induction.

Beneficial effects of IH-901 on glucose and lipid metabolisms via activating adenosine monophosphate-activated protein kinase and phosphatidylinositol-3 kinase pathways.

IH-901 is an intestinal metabolite of ginsenosides found in Panax ginseng. In the present study, effects of IH-901 on glucose and lipid metabolisms were examined using C2C12 myotubes and C57BL/ksJ db/db mice. A significant increase in phosphorylated adenosine monophosphate-activated protein kinase was observed when differentiated C2C12 myotubes were treated with IH-901. Glucose transporter 4 protein expressions were also up-regulated when muscle cells were treated with of IH-901 up to 60 minutes, resulting in stimulation of glucose uptake by 25% as compared with untreated cells. In addition, phosphatidylinositol-3 kinase and Akt protein expressions were increased when C2C12 myotubes were exposed to IH-901 for up to 3 hours; and these effects including glucose uptake were attenuated by pretreatment with LY294002, a selective phosphatidylinositol-3 kinase inhibitor. In animal study, IH-901 at 25 mg/kg lowered the plasma glucose, triglyceride, cholesterol, and nonesterified fatty acid levels by 20.7%, 41.6%, 20.2%, and 24.6%, respectively, compared with control mice. In the meantime, plasma insulin levels were significantly increased by 2.2 and 3.4 times in 10 and 25 mg/kg-treated mice, respectively, compared with control mice, in parallel with the histologic observation showing a preserved architecture of the pancreatic islet. Protein and gene expression patterns for adenosine monophosphate-activated protein kinase, sterol regulatory element binding protein-1a, and glucose transporter 4 in the liver and skeletal muscles were similar to those in cell studies. In summary, IH-901 might be a promising therapeutic agent improving altered glucose and lipid metabolisms revealed in type 2 diabetes mellitus patients.

Effect of the feeding system on the fatty acid composition, expression of the Delta9-desaturase, Peroxisome Proliferator-Activated Receptor Alpha, Gamma, and Sterol Regulatory Element Binding Protein 1 genes in the semitendinous muscle of light lambs of the Rasa Aragonesa breed.

BACKGROUND: Conjugated linoleic acids (CLAs) are receiving increasing attention because of their beneficial effects on human health, with milk and meat products derived from ruminants as important sources of CLA in the human diet. SCD gene is responsible for some of the variation in CLA concentration in adipose tissues, and PPARgamma, PPARalpha and SREBP1 genes are regulator of SCD gene. The aim of this work was to evaluate the effect of the feeding system on fatty acid composition, CLA content and relative gene expression of Delta9-desaturase (SCD), Peroxisome Proliferator-Activated Receptor Gamma (PPARgamma), Peroxisome Proliferator-Activated Receptor Alpha, (PPARalpha) and Sterol Regulatory Element Binding Protein (SREBP1) in Rasa Aragonesa light lambs in semitendinous muscle. Forty-four single-born male lambs were used to evaluate the effect of the feeding system, varying on an intensity gradient according to the use of concentrates: 1. grazing alfalfa, 2. grazing alfalfa with a supplement for lambs, 3. indoor lambs with grazing ewes and 4. drylot. RESULTS: Both grazing systems resulted in a higher concentration of vaccenic acid (VA), CLA, CLA/VA acid ratio, and a lower oleic content, oleic acid (C18:1)/stearic acid (C18:0) ratio, PUFA n-6/n-3 ratio and SCD expression compared to other diets. In addition feeding system affected the fatty acid composition and SCD expression, possibly due to CLA concentration or the PUFA n-6/n-3 ratio. Both expression of the SCD gene and the feeding system were important factors affecting CLA concentration in the animal's semitendinous muscle. PPARgamma, PPARalpha and SREBP1 expression seemed to be unaffected by the feeding system. Although no significant results were found, PPARgamma, PPARalpha and SREBP1 showed similar expression pattern as SCD. Moreover, the correlation results between SCD expression and PPARgamma (p < 0.01), as well as SREBP1 (p < 0.01) expression, may suggest that these genes were affecting SCD expression in a different way. CONCLUSIONS: The data indicated that the feeding system is the main factor affecting the fatty acid composition and SCD gene expression, which is also affected by CLA and possibly by n-6/n-3 PUFAs.

Dietary polyunsaturated fatty acids slow the progression of diabetic nephropathy in streptozotocin-induced diabetic rats.

Diabetic nephropathy is associated with lipid deposits in the kidney. We hypothesized that a diet containing polyunsaturated fatty acids (PUFAs) could ameliorate pathogenesis of diabetic kidney diseases associated with lipid depositions in the kidneys. We examined if the pathogenesis and progression of diabetic nephropathy are affected by the type of dietary fat using streptozotocin (45 mg/kg body weight, intravenous)-induced diabetic rats (5-week-old male Sprague-Dawley rats). Streptozotocin-induced diabetic rats were fed a lard diet containing saturated fatty acids or a rapeseed oil diet containing PUFAs (DML and DMR, respectively) for 11 days. Similarly, streptozotocin-nontreated rats were fed a lard diet or a rapeseed oil diet (NL and NR, respectively) for 11 days. Hyperglycemia was induced in DML and DMR, compared with NL and NR groups. The levels of plasma ketone, total cholesterol, and triglyceride (TG) were significantly increased in the DML group. Moreover, albuminuria and renal TG content were enhanced in the DML group. The renal TG content correlated positively with urinary albumin excretion (P < .001). Oil-Red O staining of kidney sections indicated a marked accumulation of neutral lipids in both glomerular and tubular cells in the DML group. In addition, a renal sterol regulatory element-binding protein-1 mature protein increment was induced in the DML group. Conversely, sterol regulatory element-binding protein-1 expression in the kidney was maintained at normal levels in the DMR group. These results suggest that dietary PUFAs may slow the progression of diabetic nephropathy associated with lipid depositions in the kidney.

Hepatitis C virus protein and iron overload induce hepatic steatosis through the unfolded protein response in mice.

BACKGROUND/AIM: Hepatic iron overload and steatosis play critical roles in the progression of hepatitis C virus (HCV)-associated chronic liver disease. However, how these two pathophysiological features affect each other remains unknown. The aim of this study was to investigate how hepatic iron overload contributes to the development of hepatic steatosis in the presence of HCV proteins. METHODS: Male C57BL/6 transgenic mice expressing the HCV polyprotein and nontransgenic littermates were fed an excess-iron diet or a control diet. Mice in each group were assessed for the molecules responsible for fat accumulation in the liver. RESULTS: Hepatic iron levels were positively correlated with triglyceride concentrations in the liver for all mice. As compared with the livers of nontransgenic mice fed the control diet, the livers of transgenic mice fed the excess-iron diet showed a lower expression of carnitine palmitoyl transferase I, a higher expression of sterol-regulatory element-binding protein 1 and fatty acid synthetase and an activated unfolded protein response indicated by a higher expression of unspliced and spliced X-box DNA-binding protein 1 (XBP-1), phosphorylated eukaryotic initiation factor-2alpha (p-eIF2alpha), CCAAT/enhancer-binding protein homology protein (CHOP) and abundant autophagosomes concomitant with increased production of reactive oxygen species. Six-month treatment with the anti-oxidant N-acetyl cysteine dramatically reduced hepatic steatosis in transgenic mice fed the excess-iron diet through decreased expression of unspliced and spliced XBP-1, p-eIF2alpha, and CHOP. CONCLUSIONS: The iron-induced unfolded protein response appears to be one of the mechanisms responsible for fat accumulation in the liver in transgenic mice expressing the HCV polyprotein.

Omacor, n-3 polyunsaturated fatty acid, attenuated albuminuria and renal dysfunction with decrease of SREBP-1 expression and triglyceride amount in the kidney of type II diabetic animals.

BACKGROUND: We assumed that n-3 polyunsaturated fatty acid (n-3 PUFA) would attenuate the tissue dyslipidemic condition through suppression of sterol regulatory element-binding protein (SREBP-1) in the kidney and would prevent renal progression in diabetic animals. METHODS: We gavaged Omacor, composed of docosahexaenoic acid and eicosapentaenoic acid, to db/db mice for 2 weeks (0.2 g/100 g/day). We measured the markers of renal function, triglyceride amount and expressions of SREBP-1, liver X-activated receptor alpha (LXRalpha), collagen IV and TGFbeta-1 in kidney lysate, and performed immunohistochemical staining for SREBP-1, desmin and WT-1 in the renal sections. We measured collagen IV in primary mesangial cells cultured with high glucose media (25 mM), both with and without a transient transfection of small interfering RNA (siRNA) SREBP-1. RESULTS: Omacor decreased the concentration of serum free fatty acid, and the amount of renal triglyceride, which was associated with decreased expression of SREBP-1 in the kidney, albuminuria and renal dysfunction in db/db mice. Omacor attenuated the expansion of mesangial matrix and the expression of desmin, preserved the WT-1 positive cells, and inhibited the phosphorylation of nuclear factor kappaB in renal tissue. In mesangial cells cultured in high glucose media, the suppression of SREBP-1 expression decreased the collagen IV in the cells. CONCLUSIONS: Our study results demonstrated that n-3 PUFA prevented renal progression with attenuation of SREBP-1 and reduction of triglyceride in the diabetic kidney. This suggests that the regulation of dyslipidemic signals in the kidney could be a possible mechanism by which PUFA preserves renal function in the diabetic condition.

Increased carnitine palmitoyl transferase 1 expression and decreased sterol regulatory element-binding protein 1c expression are associated with reduced intramuscular triglyceride accumulation after insulin therapy in high-fat-diet and streptozotocin-induced diabetic rats.

We previously reported that early insulin treatment reduced intramuscular triglyceride content in type 2 diabetes mellitus Sprague-Dawley rats; the underlying mechanisms are, however, not completely understood. Here we investigated the regulation of insulin on molecular expressions involved in lipid metabolism pathways in skeletal muscle of high-fat-diet and streptozotocin-induced diabetic Sprague-Dawley rats. Neutral protamine Hagedorn insulin and gliclazide were initiated at the third day after streptozotocin injection and lasted for 3 weeks. Compared with normal rats, untreated diabetic rats had a 30% and 61% increase in lipoprotein lipase protein expression and activity, which were decreased by insulin and gliclazide (P < .05). Fatty acid translocase protein was down-regulated by 45% in untreated diabetic rats, which was up-regulated by 31% and 26% with insulin and gliclazide, respectively (P < .05). Insulin failed to affect fatty acid transport protein 1 and fatty acid binding protein expressions. Carnitine palmitoyl transferase 1 had a 47% decrease in untreated diabetic rats, which was normalized by insulin (P < .05). Moreover, compared with normal rats, untreated diabetic rats had higher expressions of sterol regulatory element-binding protein 1c, tumor necrosis factor alpha, and Tyr(705) phosphorylation of signal transducer and activator of transcription 3 levels, which all were down-regulated after insulin treatment. These results suggested that early insulin reduced intramuscular triglyceride levels in diabetic rats potentially through amelioration of lipid dysfunction and inhibition of lipid synthesis.