Diet and nonalcoholic fatty liver disease

PURPOSE OF REVIEW: Nonalcoholic fatty liver disease is a common and serious form of chronic liver disease. It is characterized by lipid accumulation in the liver and is associated with all aspects - and may even be an initiating factor - of the metabolic syndrome. The purpose of this review is to summarize recent findings from human studies on dietary effects on hepatic lipid accumulation. RECENT FINDINGS: Epidemiological studies did not give consistent results. From intervention studies there is evidence to support a role for weight loss. Some studies have also suggested that decreasing total fat intake and increasing the intake of fish oils may be beneficial in the treatment of nonalcoholic steatohepatitis. SUMMARY: Only a few studies have focused on dietary effects on hepatic lipid accumulation. So far, there is only evidence to support a role for weight loss. Decreasing total fat intake and increasing the intake of fish oils may also be beneficial, but these conclusions are based on a limited number of studies, which sometimes lacked a proper control group. Also, other nutrients have not been studied in detail. Therefore, there is an urgent need for evidence-based dietary guidelines to prevent or even to treat nonalcoholic fatty liver disease.     

A proton nuclear magnetic resonance study of sulfmyoglobin cyanide

The proton nuclear magnetic resonance spectrum of sulfmyoglobin cyanide was studied at 400 MHz. The position of a methyl-group resonance at low field is consistent with a chlorin-like structure for the prosthetic group. The proton NMR spectrum of the cyanide derivative of the purified prosthetic group which decomposes upon extraction from the protein was found to be the same as that of the cyanide derivative of the prosthetic group extracted from myoglobin and a sample prepared from hemin-Cl.     

The hedgehog pathway in nonalcoholic fatty liver disease

Nonalcoholic fatty liver disease (NAFLD) encompasses a spectrum of obesity-associated liver diseases and it has become the major cause of cirrhosis in the Western world. The high prevalence of NAFLD-associated advanced liver disease reflects both the high prevalence of obesity-related fatty liver (hepatic steatosis) and the lack of specific treatments to prevent hepatic steatosis from progressing to more serious forms of liver damage, including nonalcoholic steatohepatitis (NASH), cirrhosis, and primary liver cancer. The pathogenesis of NAFLD is complex, and not fully understood. However, compelling evidence demonstrates that dysregulation of the hedgehog (Hh) pathway is involved in both the pathogenesis of hepatic steatosis and the progression from hepatic steatosis to more serious forms of liver damage. Inhibiting hedgehog signaling enhances hepatic steatosis, a condition which seldom results in liver-related morbidity or mortality. In contrast, excessive Hh pathway activation promotes development of NASH, cirrhosis, and primary liver cancer, the major causes of liver-related deaths. Thus, suppressing excessive Hh pathway activity is a potential approach to prevent progressive liver damage in NAFLD. Various pharmacologic agents that inhibit Hh signaling are available and approved for cancer therapeutics; more are being developed to optimize the benefits and minimize the risks of inhibiting this pathway. In this review we will describe the Hh pathway, summarize the evidence for its role in NAFLD evolution, and discuss the potential role for Hh pathway inhibitors as therapies to prevent NASH, cirrhosis and liver cancer.     

Hepatic triglyceride synthesis and nonalcoholic fatty liver disease

PURPOSE OF REVIEW: Nonalcoholic fatty liver disease is a spectrum of diseases ranging from simple steatosis to cirrhosis. The hallmark of nonalcoholic fatty liver disease is hepatocyte accumulation of triglycerides. We will review the role of triglyceride synthesis in nonalcoholic fatty liver disease progression and summarize recent findings about triglyceride synthesis inhibition and prevention of progressive disease. RECENT FINDINGS: Attempts to inhibit triglyceride synthesis in animal models have resulted in improvement in hepatic steatosis. Studies in animal models of nonalcoholic fatty liver disease demonstrate that inhibition of acyl-coenzyme A:diacylglycerol acyltransferase, the enzyme that catalyzes the final step in triglyceride synthesis, results in improvement in hepatic steatosis and insulin sensitivity. We recently confirmed that hepatic specific inhibition of acyl-coenzyme A:diacylglycerol acyltransferase with antisense oligonucleotides improves hepatic steatosis in obese, diabetic mice but, unexpectedly, exacerbated injury and fibrosis in that model of progressive nonalcoholic fatty liver disease. When hepatocyte triglyceride synthesis was inhibited, free fatty acids accumulated in the liver, leading to induction of fatty acid oxidizing systems that increased hepatic oxidative stress and liver damage. These findings suggest that the ability to synthesize triglycerides may, in fact, be protective in obesity. SUMMARY: Nonalcoholic fatty liver disease is strongly associated with obesity and peripheral insulin resistance. Peripheral insulin resistance increases lipolysis in adipose depots, promoting increased free fatty acid delivery to the liver. In states of energy excess, such as obesity, the latter normally triggers hepatic triglyceride synthesis. When hepatic triglyceride synthesis is unable to accommodate increased hepatocyte free fatty acid accumulation, however, lipotoxicity results. Thus, rather than being hepatotoxic, liver triglyceride accumulation is actually hepato-protective in obese, insulin-resistant individuals.     

Biotransformations monitored in situ by proton nuclear magnetic resonance spectroscopy

One-dimensional Fourier-transform proton nuclear magnetic resonance (1H-NMR) spectroscopy can be used to study biotransformations in situ, in vivo and in aqua (1H2O). Although an insensitive method, it rapidly provides solution-structural information of mixtures of diverse compounds that are used and formed during enzymic reactions and culture fermentations; the samples do not require any physical or chemical processing for analysis. The absolute stereochemistry of some reactions can also be determined, and assessments of metabolic fluxes made. This technique, with appropriate modifications, is of obvious value for on-line assessments of industrial fermentation processes.     

Mobile domains in ribosomes revealed by proton nuclear magnetic resonance

Ribosomes and subunits from eukaryotic and prokaryotic sources were studied by high-resolution proton magnetic-resonance spectroscopy. If all ribosomal components are firmly bound within the particle, then only broad spectra would be expected. However, relatively sharp resonances were found both in ribosomal subunits and in 70 or 80 S ribosomes. The regions of these mobile protein domains have been partially assigned in Escherichia coli ribosomes. Large and small ribosomal subunits were treated to remove selectively proteins L7/12 and S1, respectively. Sharp proton magnetic resonance spectra were not observed for the stripped large subunit showing that proteins L7/12 comprise the flexible protein region and that there is little other flexibility in the stripped subunit. Complete removal of S1 from the small subunit greatly reduced but did not abolish the sharp protein resonance peaks, indicating that protein S1 contains a substantial flexible component but that other flexible components remain in the stripped small subunit. Evidence for generality of these features of ribosome organization is provided by similar studies on ribosomes from eukaryotic sources.     

A proton nuclear magnetic resonance-based metabolomic approach in IgA nephropathy urinary profiles

Immunoglobulin A nephropathy (IgAN) is the most common form of primary glomerulonephritis worldwide. Both one-dimensional NOESY and transverse-relaxation filter CPMG NMR spectra were recorded to investigate the urine metabolome of 24 IgAN patients and to detect altered metabolic profiles in comparison with 68 healthy matched controls. The spectral data were analyzed using multivariate statistical techniques. The analysis revealed that the NMR spectra of IgAN patients were statistically different from those of the controls (P = 4 × 10^?7 for 1D-NOESY and P = 2 × 10^?7 for CPMG). The robustness of the determined statistical model was confirmed by its predictive performance (for the 1D-NOESY dataset: sensitivity = 67 %, specificity = 95 %; for the CPMG dataset sensitivity = 60 %, specificity = 94 %). For the first time we found metabolites, including betaine and citrate, that are differentially modulated in IgAN patients compared to controls and that may be directly involved in the pathogenesis of IgAN. These metabolites may influence, directly or indirectly, the TNF-α, a regulating factor of the Th1/Th2 cell balance that is relevant in the pathology. The involvement of metabolites such as betaine and citrate in TNF-α regulation supports the power of the identified metabolic profiles to discern IgAN from controls.     

High-resolution proton nuclear magnetic resonance studies of human gastrin

High-resolution 1H nuclear magnetic resonance (NMR) spectroscopy at 300 MHz has been used to study the behavior of human gastrin in aqueous solution. A large number of resonances have been assigned by analysis of one- and two-dimensional NMR spectra and the effects of pH and by comparison with the spectrum of des-less than Glu1-gastrin. In gastrin, the ratio of cis to trans conformations around the Gly-2 to Pro-3 peptide bond is 3:7. This is reflected in splitting of the resonances of several neighboring residues and of a residue distant in the sequence, Tyr-12. The pKa of Tyr-12 is 10.7. Sulfation of this residue perturbs the resonances of Tyr-12 and Gly-13 but has very little effect on the rest of the spectrum. A study of the temperature dependence shows that several perturbed resonances move toward their expected positions as the temperature is raised but with a linear dependence on temperature, consistent with a redistribution of populations among accessible local conformations rather than a cooperative conformational change. Addition of Na+ or Ca2+ causes only minor changes in the spectrum. The paramagnetic metal ion Co2+ produces a number of spectral changes, reflecting strong binding to at least one site involving the Glu residues and weaker binding to Asp-16.     

A proton nuclear magnetic resonance study on the solution structure of crotamine

Proton nuclear magnetic resonance (NMR) spectra of crotamine, a myotoxic protein from a Brazilian rattlesnake (Crotalus durissus terrificus), have been analyzed. All the aromatic proton resonances have been assigned to amino acid types, and those from Tyr-1, Phe-12, and Phe-25 to the individual residues. ThepH dependence of the chemical shifts of the aromatic proton resonances indicates that Tyr-1 and one of the two histidines (His-5 or His-10) are in close proximity. A conformational transition takes place at acidicpH, together with immobilization of Met-28 and His-5 or His-10. Two sets of proton resonances have been observed for He-17 and His-5 or His-10, which suggests the presence of two structural states for the crotamine molecule in solution.     

A proton nuclear magnetic resonance study of gamma-glutamyl-amino acid cyclotransferase in human erythrocytes

Spin-echo NMR spectroscopy was shown to be a reliable technique for the monitoring of the in situ cleavage of gamma-Glu-Ala by gamma-glutamyl-amino acid cyclotransferase in whole erythrocytes and hemolysates. Of particular importance was the difference in chemical shifts between peptide resonances and those of the constituent amino acids. Using lysates of varying dilution, it was shown that the specific activity of the enzyme was not concentration-dependent, thus suggesting a lack of cytosolic low-molecular-weight-effectors or enzyme dissociation. Furthermore, the initial velocities of the reaction as a function of substrate concentration obeyed Michaelis-Menten kinetics with a Km = 2.0 +/- 0.3 mmol/l and Vmax = 137 +/- 7 mmol/h/l of cell water in 1H2O medium. Similar analysis in 2H2O medium revealed a solvent kinetic isotope effect of 1.9 +/- 0.4 at low substrate concentrations. The implications of this observation for the mechanism of the reaction are discussed. Cleavage of the peptide by a suspension of intact erythrocytes was at a rate 300 times less than the corresponding lysate flux, thus indicating the rate limitation by transport in the coupled system.     

Strategies for biomarker discovery in fibrotic disease

The discovery and development of biomarkers for fibrotic diseases have potential utility in clinical decision-making as well as in pharmaceutical research and development. This review describes strategies for identifying diagnostic, prognostic and theranostic biomarkers. A range of technologies and platforms for biomarker discovery are highlighted, including several with specific relevance for fibrosis. Some challenges specific to fibrotic diseases are outlined including; benchmarking biomarkers against imperfect clinical measures of fibrosis, the complexity resulting from diverse aetiologies and target organs, and the availability of samples (including biopsy) from well-characterised patients with fibrotic disease. To overcome these challenges collaboration amongst clinical specialities as well as between academia and industry is essential. This article is part of a Special Issue entitled: Fibrosis: Translation of basic research to human disease.     

A proton nuclear magnetic resonance study of the binding of methylmercury in human erythrocytes

The binding of methylmercury, CH₃Hg(II), by small molecules in the intracellular region of human erythrocytes has been studied by ¹H-NMR spectroscopy. To suppress or completely eliminate interfering resonances from the much more abundant hemoglobin protons, spectra were measured by a technique based on the transfer of saturation throughout the envelope of hemoglobin resonances following a selective presaturation pulse or by the spin-echo Fourier transform method. With these techniques, ¹H-NMR spectra were measured for the more abundant intracellular small molecules, including glycine, alanine, creatine, lactic acid, ergothioneine and glutathione, in both intact and hemolyzed erythrocytes to which CH₃Hg(II) had been added. The results for intact erythrocytes indicate that part of the CH₃Hg(II) is complexed by intracellular glutathione. These results also indicate that exchange of CH₃Hg(II) among glutathione molecules is fast, with the average lifetime of a CH₃Hg(II)-glutathione complex estimated to be less than 0.01 s. From exchange-averaged chemical shifts of the resonance for the proton on the α-carbon of the cysteine residue of glutathione, it is shown that, in hemolyzed erythrocytes, the sulfhydryl group of glutathione binds CH₃Hg(II) more strongly than the sulfhydryl groups of hemoglobin.     

A proton nuclear magnetic resonance investigation of histidyl residues in human normal adult hemoglobin

High-resolution proton nuclear magnetic resonance (NMR) spectroscopy at 250 MHz has been used to titrate 22 individual surface histidyl residues (11 per alpha beta dimer) of human normal adult hemoglobin in both the deoxy and the carbon monoxy forms. The proton resonances of beta 2, beta 143, and beta 146 histidyl residues are assigned by a parallel 1H NMR titration of appropriate mutant and chemically modified hemoglobins. The pK values of the 22 histidyl residues investigated are found to range from 6.35 to 8.07 in the deoxy form and from 6.20 to 7.87 in the carbon monoxy form, in the presence of 0.1 M Bis-Tris or 0.1 M Tris buffer in D2O with chloride ion concentrations varying from 5 to 60 mM at 27 degrees C. Four histidyl residues in the deoxy form and one histidyl residue in the carbon monoxy form are found to have proton nuclear magnetic resonance titration curves that deviate greatly from that predicted by the simple proton dissociation equilibrium of a single ionizable group. The proton nuclear magnetic resonance data are used to ascertain the role of several surface histidyl residues in the Bohr effect of hemoglobin under the above-mentioned experimental conditions. Under these experimental conditions, we have found that (i) the beta 146 histidyl residues do not change their electrostatic environments significantly upon binding of ligand to deoxyhemoglobin and, thus, their contribution to the Bohr effect is negligible, (ii) the beta 2 histidyl residues have a negative contribution to the Bohr effect, and (iii) the total contribution of the 22 histidyl residues investigated here to the Bohr effect is, in magnitude, comparable to the Bohr effect observed experimentally. These results suggest that the molecular mechanism of the Bohr effect proposed by Perutz [Perutz, M.F. (1970) Nature (London) 228, 726-739] is not unique and that the detailed mechanism depends on experimental conditions, such as the solvent composition.     

Free radical biology for medicine: learning from nonalcoholic fatty liver disease

Reactive oxygen species, when released under controlled conditions and limited amounts, contribute to cellular proliferation, senescence, and survival by acting as signaling intermediates. In past decades there has been an epidemic diffusion of nonalcoholic fatty liver disease (NAFLD) that represents the result of the impairment of lipid metabolism, redox imbalance, and insulin resistance in the liver. To date, most studies and reviews have been focused on the molecular mechanisms by which fatty liver progresses to steatohepatitis, but the processes leading toward the development of hepatic steatosis in NAFLD are not fully understood yet. Several nuclear receptors, such as peroxisome proliferator-activated receptors (PPARs) α/γ/δ, PPARγ coactivators 1α and 1β, sterol-regulatory element-binding proteins, AMP-activated protein kinase, liver-X-receptors, and farnesoid-X-receptor, play key roles in the regulation of lipid homeostasis during the pathogenesis of NAFLD. These nuclear receptors may act as redox sensors and may modulate various metabolic pathways in response to specific molecules that act as ligands. It is conceivable that a redox-dependent modulation of lipid metabolism, nuclear receptor-mediated, could cause the development of hepatic steatosis and insulin resistance. Thus, this network may represent a potential therapeutic target for the treatment and prevention of hepatic steatosis and its progression to steatohepatitis. This review summarizes the redox-dependent factors that contribute to metabolism alterations in fatty liver with a focus on the redox control of nuclear receptors in normal liver as well as in NAFLD.     

A theoretical study of lipid accumulation in the liver—implications for nonalcoholic fatty liver disease

A hallmark of the nonalcoholic fatty liver disease is the accumulation of lipids. We developed a mathematical model of the hepatic lipid dynamics to simulate the fate of fatty acids in hepatocytes. Our model involves fatty acid uptake, lipid oxidation, and lipid export. It takes into account that storage of triacylglycerol within hepatocytes leads to cell enlargement reducing the sinusoids radius and impairing hepatic microcirculation. Thus oxygen supply is reduced, which impairs lipid oxidation. The analysis of our model revealed a bistable behavior (two stable steady states) of the system, in agreement with histological observations showing distinct areas of lipid accumulation in lobules. The first (healthy) state is characterized by intact lipid oxidation and a low amount of stored lipids. The second state in our model may correspond to the steatotic cell; it is marked by a high amount of stored lipids and a reduced lipid oxidation caused by impaired oxygen supply. Our model stresses the role of insufficient oxygen supply for the development of steatosis. We discuss implications of our results in regard to the experimental design aimed at exploring lipid metabolism reactions under steatotic conditions. Moreover, the model helps to understand the reversibility of lipid accumulation and predicts the reversible switch to show hysteresis. The system can switch from the steatotic state back to the healthy state by reduction of fatty acid uptake below the threshold at which steatosis started. The reversibility corresponds to the observation that caloric restriction can reduce the lipid content in the liver.     

芳香烃受体在非酒精性脂肪性肝脏疾病中的作用

非酒精性脂肪性肝脏疾病(nonalcoholic fatty liver diseases,NAFLD)是目前备受关注的一种肝脏疾病。肥胖、2型糖尿病、高脂血症等是NAFLD的重要危险因素,但其发病机理仍不十分清楚。芳香烃受体(aryl hydrocarbon receptor,AHR)是由配体激活的转录因子,其在多种重要疾病活动中发挥了重要作用。近年来多项研究表明AHR激活促进了NAFLD的发病进展,对AHR参与NAFLD发病机制的探讨将有利于进一步阐明NAFLD的发病机理,为NAFLD的防治提出新的思路。本文就AHR与NAFLD关系的研究进展做一综述,以期为该领域的研究提供新的方向。     

Advances in top-down proteomics for disease biomarker discovery

Top-down mass spectrometry strategies allow identification and characterization of proteins and protein networks by direct fragmentation. These analytical processes involve a panel of fragmentation mechanisms, some of which preserve protein post-translational modifications. Thus top-down is of special interest in clinical biochemistry to probe modified proteins as potential disease biomarkers. This review describes separating methods, mass spectrometry instrumentation, bioinformatics, and theoretical aspects of fragmentation mechanisms used for top-down analysis. The biological interest of this strategy is extensively reported regarding the characterization of post-translational modifications in biochemical pathways and the discovery of biomarkers. One has to bear in mind that quantitative aspects that are beyond the focus of this review are also of critical important for biomarker discovery. The constant evolution of technologies makes top-down strategies crucial players in clinical and basic proteomics.     

Fatty acids and the endoplasmic reticulum in nonalcoholic fatty liver disease

Nonalcoholic fatty liver disease (NAFLD) represents a burgeoning public health concern in westernized nations. The obesity-related disorder is associated with an increased risk of cardiovascular disease, type 2 diabetes and liver failure. Although the underlying pathogenesis of NAFLD is unclear, increasing evidence suggests that excess saturated fatty acids presented to or stored within the liver may play a role in both the development and progression of the disorder. A putative mechanism linking saturated fatty acids to NAFLD may be endoplasmic reticulum (ER) stress. Specifically, excess saturated fatty acids may induce an ER stress response that, if left unabated, can activate stress signaling pathways, cause hepatocyte cell death, and eventually lead to liver dysfunction. In the current review we discuss the involvement of saturated fatty acids in the pathogenesis of NAFLD with particular emphasis on the role of ER stress.