Loss of intestinal fatty acid binding protein increases the susceptibility of male mice to high fat diet-induced fatty liver

Mice lacking I-FABP (encoded by the Fabp2 gene) exhibit a gender dimorphic response to a high fat/cholesterol diet challenge characterized by hepatomegaly in male I-FABP-deficient mice. In this study, we determined if this gender-specific modification of liver mass in mice lacking I-FABP is attributable to the high fat content of the diet alone and whether hepatic Fabp1 gene (encodes L-FABP) expression contributes to this difference. Wild-type and Fabp2-/- mice of both genders were fed a diet enriched with either polyunsaturated or saturated fatty acids (PUFA or SFA, respectively) in the absence of cholesterol. Male Fabp2-/- mice, but not female Fabp2-/- mice, exhibited increased liver mass and hepatic triacylglycerol (TG) deposition as compared to corresponding wild-type mice. In wild-type mice that were fed the standard chow diet, there was no difference in the concentration of hepatic L-FABP protein between males and females although the loss of I-FABP did cause a slight reduction of hepatic L-FABP abundance in both genders. The hepatic L-FABP mRNA abundance in both male and female wild-type and Fabp2-/- mice was higher in the PUFA-fed group than in the SFA-fed group, and was correlated with L-FABP protein abundance. No correlation between hepatic L-FABP protein abundance and hepatic TG concentration was found. The results obtained demonstrate that loss of I-FABP renders male mice sensitive to high fat diet-induced fatty liver, and this effect is independent of hepatic L-FABP.     

Capsaicin suppresses liver fat accumulation in high-fat diet-induced NAFLD mice

ABSTRACT

Dietary capsaicin exhibits anti-steatosis activity in obese mice. High-fat diet (HFD)-induced mice is a highly studied approach to develop non-alcoholic fatty liver disease (NAFLD). In this study, we determined whether the topical application of capsaicin can improve lesions of NAFLD. The HFD-induced mice were treated with daily topical application of capsaicin for 8 weeks. Topical application of capsaicin reduced liver fat in HFD-fed mice. Capsaicin stimulated carnitine palmitoyl transferase (CPT)-1 and CD36 expression, which are associated with β-oxidation and fatty acids influx of liver while it decreased the expression of key enzymes involved in the synthesis of fatty acids, such as acetyl Co-A carboxylase (ACC) and fatty acid synthase (FAS). Immunohistochemical analysis revealed the elevated level of adiponectin in liver tissue of the capsaicin-treated mice. These results suggest that the topical application of capsaicin suppresses liver fat accumulation through the upregulation of β-oxidation and de novo lipogenesis in HFD-induced NAFLD mice.     

Reduced antioxidant capacity and diet-induced atherosclerosis in uncoupling protein-2-deficient mice

Vascular dysfunction in response to reactive oxygen species (ROS) plays an important role in the development and progression of atherosclerotic lesions. In most cells, mitochondria are the major source of cellular ROS during aerobic respiration. Under most conditions the rates of ROS formation and elimination are balanced through mechanisms that sense relative ROS levels. However, a chronic imbalance in redox homeostasis is believed to contribute to various chronic diseases, including atherosclerosis. Uncoupling protein-2 (UCP2) is a mitochondrial inner membrane protein shown to be a negative regulator of macrophage ROS production. In response to a cholesterol-containing atherogenic diet, C57BL/6J mice significantly increased expression of UCP2 in the aorta, while mice lacking UCP2, in the absence of any other genetic modification, displayed significant endothelial dysfunction following the atherogenic diet. Compared with wild-type mice, Ucp2(-/-) mice had decreased endothelial nitric oxide synthase, an increase in vascular cell adhesion molecule-1 expression, increased ROS production, and an impaired ability to increase total antioxidant capacity. These changes in Ucp2(-/-) mice were associated with increased aortic macrophage infiltration and more numerous and larger atherosclerotic lesions. These data establish that in the vasculature UCP2 functions as an adaptive antioxidant defense to protect against the development of atherosclerosis in response to a fat and cholesterol diet.     

Increased expression of PPARgamma in high fat diet-induced liver steatosis in mice

The present study was performed to examine a hypothesis that peroxisome proliferator-activated receptor gamma (PPARgamma) is implicated in high fat diet-induced liver steatosis. Mice were fed with control or high fat diet containing approximately 10% or 80% cholesterol, respectively. Macroscopic and microscopic findings demonstrated that lipid accumulation in the liver was observed as early as 2 weeks after high fat diet and that high fat diet for 12 weeks developed a fatty liver phenotype, establishing a novel model of diet-induced liver steatosis. Gene profiling with microarray and real-time PCR studies demonstrated that among genes involved in lipid metabolism, adipogenesis-related genes, PPARgamma and its targeted gene, CD36 mRNA expression was specifically up-regulated in the liver by high fat diet for 2 weeks. Immunohistochemical study revealed that PPARgamma protein expression is increased in the nuclei of hepatocytes by high fat diet. It was also shown that protein expression of cAMP response element-binding protein (CREB), an upstream molecule of PPARgamma, in the liver was drastically suppressed by high fat diet. All these results suggest for the first time that the CREB-PPARgamma signaling pathway may be involved in the high fat diet-induced liver steatosis.     

The obesity and fatty liver are reduced by plant-derived Pediococcus pentosaceus LP28 in high fat diet-induced obese mice

We evaluated the effect of an oral administration of a plant-derived lactic acid bacterium, Pediococcus pentosaceus LP28 (LP28), on metabolic syndrome by using high fat diet-induced obese mice. The obese mice were divided into 2 groups and fed either a high fat or regular diet for 8 weeks. Each group was further divided into 3 groups, which took LP28, another plant-derived Lactobacillus plantarum SN13T (SN13T) or no lactic acid bacteria (LAB). The lean control mice were fed a regular diet without inducing obesity prior to the experiment. LP28 reduced body weight gain and liver lipid contents (triglyceride and cholesterol), in mice fed a high fat diet for 8 weeks (40%, 54%, and 70% less than those of the control group without LAB, and P = 0.018, P<0.001, and P = 0.021, respectively), whereas SN13T and the heat treated LP28 at 121°C for 15 min were ineffective. Abdominal visceral fat in the high fat diet mice fed with LP28 was also lower than that without LAB by 44%, although it was not significant but borderline (P = 0.076). The sizes of the adipocytes and the lipid droplets in the livers were obviously decreased. A real-time PCR analyses showed that lipid metabolism-related genes, such as CD36 (P = 0.013), SCD1 encoding stearoyl-CoA desaturase 1 (not significant but borderline, P = 0.066), and PPARγ encoding peroxisome proliferator-activated receptor gamma (P = 0.039), were down-regulated by taking LP28 continuously, when compared with those of the control group. In conclusion, LP28 may be a useful LAB strain for the prevention and reduction of the metabolic syndrome.     

Gelatinases impart susceptibility to high-fat diet-induced obesity in mice

Gelatinases play a role in adipose and muscle hypertrophy and could be involved in tissue remodeling in response to high-fat diet (HFD) intake. This study tested potential roles of gelatinases (matrix metalloproteinses-2 and ?9 [MMP-2 and -9]) in relationship to an antigrowth factor [myostatin (MSTN)] known to be dysregulated in relation to HFD-induced obesity (HFDIO) propensity. In vitro and ex vivo analyses demonstrated that MMP-9 increased mature MSTN levels, indicating a potential role of gelatinases in MSTN activation in vivo. HFD intake resulted in increased body weight and circulating blood glucose values in C57BL/6J and MMP-9 null mice, with no changes observed in SWR/J mice. HFD intake attenuated MMP-9 and MMP-2 mRNA levels in SWR/J mice while elevating MMP-2 levels in skeletal muscle in C57BL/6J mice. In MMP-9 null mice, the effects of HFD intake were muted. Consistent with changes in mRNA levels, HFD intake increased MMP-9 activity in muscle tissue of C57BL/6J mice, demonstrating a strong relationship between HFDIO susceptibility and local MMP regulation. Overall, resistance to HFDIO appears to correspond to low MMP-9 and MSTN levels, suggesting a role of MMP-9 in MSTN activation in local tissue responses to HFD intake.     

A targeted apolipoprotein B-38.9-producing mutation causes fatty livers in mice due to the reduced ability of apolipoprotein B-38.9 to transport triglycerides

Nonphysiological truncations of apolipoprotein (apo) B-100 cause familial hypobetalipoproteinemia (FHBL) in humans and mice. An elucidation of the mechanisms underlying the FHBL phenotypes may provide valuable information on the metabolism of apo B-containing lipoproteins and the structure-function relationship of apo B. To generate a faithful mouse model of human FHBL, a subtle mutation was introduced into the mouse apo B gene by targeting embryonic stem cells using homologous recombination followed by removal of the selection marker gene by Cre-loxP-mediated site-specific recombination. The engineered mice bear a premature stop codon at residue 1767 and a 42-base pair loxP inserted into intron 24 of the apo B gene, thus closely resembling the apo B-38.9-producing mutation in humans. Apo B-38.9 was the sole apo B protein in homozygote (apob(38.9/38.9)) plasma. In heterozygotes (apob(+/)(38. 9)), apo B-100 and apo B-48 were reduced by 75 and 40%, respectively, and apo B-38.9 represented 20% of total circulating apo B. Hepatic apo B-38.9 mRNA levels were reduced by 40%. In cultured apob(+/)(38. 9) hepatocytes, apo B-100 was produced in trace quantities, and the synthesis rate of apo B-38.9 relative to apo B-48 was reduced by 40%. However, almost equimolar amounts of apo B-38.9 and apo B-48 were secreted into the media. Pulse-chase studies revealed that apo B-38. 9 was secreted at a faster rate and more efficiently than apoB-48. Nevertheless, both apob(+/)(38.9) and apob(38.9/38.9) mice had reduced hepatic triglyceride secretion rates and fatty livers. Thus, low mRNA levels or defective secretion of apo B-38.9 may not be responsible for the FHBL phenotypes caused by the apo B-38.9 mutation. Rather, a reduced capacity of apo B-38.9 for triglyceride transport may account for the fatty livers in these mice.     

Chromium attenuates high-fat diet-induced nonalcoholic fatty liver disease in KK/HlJ mice

Nonalcoholic fatty liver disease (NAFLD) is a chronic liver disease associated with insulin resistance, oxidative stress, and inflammation. Evidence indicates that chromium has a role in the regulation of glucose and lipid metabolism and may improve insulin sensitivity. In this study, we report that chromium supplementation has a beneficial effect against NAFLD. We found that KK/HlJ mice developed obesity and progressed to NAFLD after feeding with high-fat diet for 8 weeks. High-fat-fed KK/HlJ mice showed hepatocyte injury and hepatic triglyceride accumulation, which was accompanied by insulin resistance, oxidative stress, and inflammation. Chromium supplementation prevented progression of NAFLD and the beneficial effects were accompanied by reduction of hepatic triglyceride accumulation, elevation of hepatic lipid catabolic enzyme, improvement of glucose and lipid metabolism, suppression of inflammation as well as resolution of oxidative stress, probably through enhancement of insulin signaling. Our findings suggest that chromium could serve as a hepatoprotective agent against NAFLD.     

Reduced susceptibility to ventricular tachyarrhythmias in rats selectively bred for high aerobic capacity

Reperfusion after a brief period of cardiac ischemia can lead to potentially lethal arrhythmias. Human epidemiological studies and experimental work with animals indicate that regular physical activity is associated with reductions in cardiovascular disease (CVD) risk factors and sudden cardiac death. Similarly, artificial selection of rats for high aerobic treadmill-running capacity (high-capacity runners; HCR) has been shown to reduce CVD risk factors relative to rats selected as low-capacity runners (LCR). Therefore, we tested the hypothesis that HCR, relative to LCR rats, would be less susceptible to ischemia-reperfusion-mediated ventricular tachyarrhythmias. To test this hypothesis, we measured the susceptibility to ventricular tachyarrhythmias produced by 3 min of occlusion and reperfusion of the left main coronary artery in conscious LCR and HCR rats. Results document a significantly lower incidence of ventricular tachyarrhythmias in HCR (3 of 11, 27.3%) relative to LCR (6 of 7, 85.6%) rats. The decreased susceptibility to tachyarrhythmias in HCR rats was associated with a reduced cardiac metabolic demand during ischemia (lower rate-pressure product and ST segment elevation) as well as a wider range for the autonomic control of heart rate. The HCR and LCR represent a unique substrate for evaluation of the mechanisms underlying ischemia-mediated cardiac arrhythmogenesis.     

Faecal microbiota transplantation-mediated jejunal microbiota changes halt high-fat diet-induced obesity in mice via retarding intestinal fat absorption

Faecal Microbiota Transplantation (FMT) is considered as a promising technology to fight against obesity. Wild boar has leanermuscle and less fat in comparison to the domestic pig, which were thought to be related with microbiota. To investigate the function and mechanism of the wild boar microbiota on obesity, we first analysed the wild boar microbiota composition via 16S rDNA sequencing, which showed that Firmicutes and Proteobacteria were the dominant bacteria. Then, we established a high-fat diet (HFD)-induced obesity model, and transfer low and high concentrations of wild boar faecal suspension in mice for 9 weeks. The results showed that FMT prevented HFD-induced obesity and lipid metabolism disorders, and altered the jejunal microbiota composition especially increasing the abundance of the Lactobacillus and Romboutsia, which were negatively correlated with obesity-related indicators. Moreover, we found that the anti-obesity effect of wild boar faecal suspension was associated with jejunal N6-methyladenosine (m⁶A) levels. Overall, these results suggest that FMT has a mitigating effect on HFD-induced obesity, which may be due to the impressive effects of FMT on the microbial composition and structure of the jejunum. These changes further alter intestinal lipid metabolism and m⁶A levels to achieve resistance to obesity.     

Proteoglycan 4 deficiency protects against glucose intolerance and fatty liver disease in diet-induced obese mice

Objective

Proteoglycan 4 (Prg4) has emerged from human association studies as a possible factor contributing to weight gain, dyslipidemia and insulin resistance. In the current study, we investigated the causal role of Prg4 in controlling lipid and glucose metabolism in mice.

The intestinal fatty acid binding protein is not essential for dietary fat absorption in mice.

The intestinal fatty acid binding protein (I-FABP) belongs to a family of 15 kDa clamshell-like proteins that are found in many different tissues. So far, nine types have been identified. Their primary structures are highly conserved between species but somewhat less so among the different types. The function of these proteins, many of which are highly expressed, is not well understood. Their ability to bind lipid ligands suggests a role in lipid metabolism, but direct evidence for this idea is still lacking. We tested the hypothesis that I-FABP serves an essential role in the assimilation of dietary fatty acids by disrupting its gene (Fabpi) in the mouse. We discovered that Fabpi-/- mice are viable, but they display alterations in body weight and are hyperinsulinemic. Male Fabpi-/- mice had elevated plasma triacylglycerols and weighed more regardless of the dietary fat content. In contrast, female Fabpi-/- mice gained less weight in response to a high-fat diet. The results clearly demonstrate that I-FABP is not essential for dietary fat absorption. We propose that I-FABP functions as a lipid-sensing component of energy homeostasis that alters body weight gain in a gender-specific fashion.     

Decreased liver fatty acid binding capacity and altered liver lipid distribution in mice lacking the liver fatty acid-binding protein gene

Although liver fatty acid-binding protein (L-FABP) is an important binding site for various hydrophobic ligands in hepatocytes, its in vivo significance is not understood. We have therefore created L-FABP null mice and report here their initial analysis, focusing on the impact of this mutation on hepatic fatty acid binding capacity, lipid composition, and expression of other lipid-binding proteins. Gel-filtered cytosol from L-FABP null liver lacked the main fatty acid binding peak in the fraction that normally comprises both L-FABP and sterol carrier protein-2 (SCP-2). The binding capacity for cis-parinaric acid was decreased >80% in this region. Molar ratios of cholesterol/cholesterol ester, cholesteryl ester/triglyceride, and cholesterol/phospholipid were 2- to 3-fold greater, reflecting up to 3-fold absolute increases in specific lipid classes in the order cholesterol > cholesterol esters > phospholipids. In contrast, the liver pool sizes of nonesterified fatty acids and triglycerides were not altered. However, hepatic deposition of a bolus of intravenously injected [14C]oleate was markedly reduced, showing altered lipid pool turnover. An increase of approximately 75% of soluble SCP-2 but little or no change of other soluble (glutathione S-transferase, albumin) and membrane (fatty acid transport protein, CD36, aspartate aminotransferase, caveolin) fatty acid transporters was measured. These results (i) provide for the first time a quantitative assessment of the contribution of L-FABP to cytosolic fatty acid binding capacity, (ii) establish L-FABP as an important determinant of hepatic lipid composition and turnover, and (iii) suggest that SCP-2 contributes to the accumulation of cholesterol in L-FABP null liver.     

Deficiency in apolipoprotein E has a protective effect on diet-induced nonalcoholic fatty liver disease in mice

Apolipoprotein E (apoE) mediates the efficient catabolism of the chylomicron remnants very low-density lipoprotein and low-density lipoprotein from the circulation, and the de novo biogenesis of high-density lipoprotein. Lipid-bound apoE is the natural ligand for the low-density lipoprotein receptor (LDLr), LDLr-related protein 1 and other scavenger receptors. Recently, we have established that deficiency in apoE renders mice resistant to diet-induced obesity. In the light of these well-documented properties of apoE, we sought to investigate its role in the development of diet-induced nonalcoholic fatty liver disease (NAFLD). apoE-deficient, LDLr-deficient and control C57BL/6 mice were fed a western-type diet (17.3% protein, 48.5% carbohydrate, 21.2% fat, 0.2% cholesterol, 4.5 kcal·g(-)) for 24 weeks and their sensitivity to NAFLD was assessed by histological and biochemical methods. apoE-deficient mice were less sensitive than control C57BL/6 mice to diet-induced NAFLD. In an attempt to identify the molecular basis for this phenomenon, biochemical and kinetic analyses revealed that apoE-deficient mice displayed a significantly delayed post-prandial triglyceride clearance from their plasma. In contrast with apoE-deficient mice, LDLr-deficient mice fed a western-type diet for 24 weeks developed significant accumulation of hepatic triglycerides and NAFLD, suggesting that apoE-mediated hepatic triglyceride accumulation in mice is independent of LDLr. Our findings suggest a new role of apoE as a key peripheral contributor to hepatic lipid homeostasis and the development of diet-induced NAFLD.     

Lack of UCP1 stimulates fatty liver but mediates UCP1-independent action of beige fat to improve hyperlipidemia in Apoe knockout mice

Brown adipose tissue (BAT) plays a critical role in lipid metabolism and may protect from hyperlipidemia; however, its beneficial effect appears to depend on the ambient temperature of the environment. In this study, we investigated the effects of uncoupling protein 1 (UCP1) deficiency on lipid metabolism, including the pathophysiology of hyperlipidemia, in apolipoprotein E knockout (APOE-KO) mice at a normal (23 °C) and thermoneutral (30 °C) temperature. Unexpectedly, UCP1 deficiency caused improvements in hyperlipidemia, atherosclerosis, and glucose metabolism, regardless of an increase in hepatic lipid deposition, in Ucp1/Apoe double-knockout (DKO) mice fed a high-fat diet at 23 °C, with BAT hyperplasia and robust browning of inguinal white adipose tissue (IWAT) observed. Proteomics and gene expression analyses revealed significant increases in many proteins involved in energy metabolism and strong upregulation of brown/beige adipocyte-related genes and fatty acid metabolism-related genes in browned IWAT, suggesting an induction of beige fat formation and stimulation of lipid metabolism in DKO mice at 23 °C. Conversely, mRNA levels of fatty acid oxidation-related genes decreased in the liver of DKO mice. The favorable phenotypic changes were lost at 30 °C, with BAT whitening and disappearance of IWAT browning, while fatty liver further deteriorated in DKO mice compared with that in APOE-KO mice. Finally, longevity analysis revealed a significant lifespan extension of DKO mice compared with that of APOE-KO mice at 23 °C. Irrespective of the fundamental role of UCP1 thermogenesis, our results highlight the importance of beige fat for the improvement of hyperlipidemia and longevity under the atherogenic status at normal room temperature.     

Quantitative trait locus analysis of susceptibility to diet-induced atherosclerosis in recombinant inbred mice

Quantitative trait locus (QTL) analysis is a statistical method that can be applied to identify loci making a significant impact on a phenotype. For the phenotype of susceptibility to diet-induced atherosclerosis in the mouse, we have studied four quantitative traits: area of aortic fatty streaks and serum concentrations of high-density lipoprotein-bound cholesterol (HDL-cholesterol), apolipoprotein A-I, and apolipoprotein A-II (apo A-II). QTL analysis revealed a significant locus on chromosome 1 distal impacting serum apo A-II concentration on a high-fat diet and serum HDL-cholesterol concentration on a chow diet. This locus is presumablyApoa-2, the structural gene for apo A-II. QTL analysis of aortic fatty streaks failed to reveal a significant locus.     

Niemann-Pick C1-Like 1 deletion in mice prevents high-fat diet-induced fatty liver by reducing lipogenesis

Niemann-Pick C1-Like 1 (NPC1L1) mediates intestinal absorption of dietary and biliary cholesterol. Ezetimibe, by inhibiting NPC1L1 function, is widely used to treat hypercholesterolemia in humans. Interestingly, ezetimibe treatment appears to attenuate hepatic steatosis in rodents and humans without a defined mechanism. Overconsumption of a high-fat diet (HFD) represents a major cause of metabolic disorders including fatty liver. To determine whether and how NPC1L1 deficiency prevents HFD-induced hepatic steatosis, in this study, we fed NPC1L1 knockout (L1-KO) mice and their wild-type (WT) controls an HFD, and found that 24 weeks of HFD feeding causes no fatty liver in L1-KO mice. Hepatic fatty acid synthesis and levels of mRNAs for lipogenic genes are substantially reduced but hepatic lipoprotein-triglyceride production, fatty acid oxidation, and triglyceride hydrolysis remain unaltered in L1-KO versus WT mice. Strikingly, L1-KO mice are completely protected against HFD-induced hyperinsulinemia under both fed and fasted states and during glucose challenge. Despite similar glucose tolerance, L1-KO relative WT mice are more insulin sensitive and in the overnight-fasted state display significantly lower plasma glucose concentrations. In conclusion, NPC1L1 deficiency in mice prevents HFD-induced fatty liver by reducing hepatic lipogenesis, at least in part, through attenuating HFD-induced insulin resistance, a state known to drive hepatic lipogenesis through elevated circulating insulin levels.