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Introduction: Obesity is associated with comorbidities such as cardiovascular disease and type 2 diabetes. HDAC3 regulates adipose tissue physiology (WAT), and its genetic inactivation causes metabolic reprogramming of white adipocytes toward browning. The aim of this work is to evaluate the effect of HDAC3 silencing at different stages of differentiation and investigate the influence of adipocyte metabolism on the immunophenotype of WAT. Materials and Methods: Following HDAC3 silencing in mesenchymal stem cells and mature adipocytes, adipocyte function, RNA, DNA and protein levels, and proliferation at the end of differentiation were analyzed. Visceral WAT immunophenotype (vWAT) of Hdac3 KO mice in WAT (Hdac3fatKO) and controls (FL) was performed by FACS. Results: Silencing HDAC3 in precursors amplifies the expression of genes and proteins that regulate differentiation, oxidative metabolism, browning and mitochondrial activity. Following silencing, we found increased 1)phosphorylation of AKT (1.64 fold change, P

Riboflavin is an essential water-soluble vitamin that needs to be provided through the diet because of the conversion into flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN), important cofactors in hundreds of flavoenzymes. The adsorption and distribution of riboflavin is mediated by transmembrane transporters of the SLC52 family, namely RFVT1-3, whose mutations are mainly associated with two diseases, MADD and the Brown-Vialetto-Van Laere syndrome. Interest in RFVTs as pharmacological targets has increased in the last few years due to their overexpression in several cancer cells, which can be exploited both by blocking the uptake of riboflavin into the cancerous cells, and by performing cancer targeted delivery of drugs with a high affinity for RFVTs. In this work we propose three-dimensional structural models for all three human riboflavin transporters obtained by state-of-the-art artificial intelligence-based methods, which were then further refined with molecular dynamics simulations. Furthermore, two of the most notable mutations concerning RFVT2 and RFVT3 (W31S and N21S respectively) were investigated studying the interactions between the wild-type and mutated transporters with riboflavin. This article is protected by copyright. All rights reserved.

The adipose organ comprises two main fat depots termed white and brown adipose tissues. Adipogenesis is a process leading to newly differentiated adipocytes starting from precursor cells, which requires the contribution of many cellular activities at the genome, transcriptome, proteome, and metabolome levels. The adipogenic program is accomplished through two sequential phases; the first includes events favoring the commitment of adipose tissue stem cells/precursors to preadipocytes, while the second involves mechanisms that allow the achievement of full adipocyte differentiation. While there is a very large literature about the mechanisms involved in terminal adipogenesis, little is known about the first stage of this process. Growing interest in this field is due to the recent identification of adipose tissue precursors, which include a heterogenous cell population within different types of adipose tissue as well as within the same fat depot. In addition, the alteration of the heterogeneity of adipose tissue stem cells and of the mechanisms involved in their commitment have been linked to adipose tissue development defects and hence to the onset/progression of metabolic diseases, such as obesity. For this reason, the characterization of early adipogenic events is crucial to understand the etiology and the evolution of adipogenesis-related pathologies, and to explore the adipose tissue precursors’ potential as future tools for precision medicine.

Obesity is a condition characterized by uncontrolled expansion of adipose tissue mass resulting in pathological weight gain. Histone deacetylases (HDACs) have emerged as crucial players in epigenetic regulation of adipocyte metabolism. Previously, we demonstrated that selective inhibition of class I HDACs improves white adipocyte functionality and promotes the browning phenotype of murine mesenchymal stem cells (MSCs) C3H/10T1/2 differentiated to adipocytes. These effects were also observed in db/db and diet induced obesity mouse models and in mice with adipose-selective inactivation of HDAC3, a member of class I HDACs. The molecular basis of class I HDACs action in adipose tissue is not deeply characterized and it is not known whether the effects of their inhibition are exerted on adipocyte precursors or mature adipocytes. Therefore, the aim of the present work was to explore the molecular mechanism of class I HDAC action in adipocytes by evaluating the effects of HDAC3-specific silencing at different stages of differentiation. HDAC3 was silenced in C3H/10T1/2 MSCs at different stages of differentiation to adipocytes. shRNA targeting HDAC3 was used to generate the knock-down model. Proper HDAC3 silencing was assessed by measuring both mRNA and protein levels of mouse HDAC3 via qPCR and western blot, respectively. Mitochondrial DNA content and gene expression were quantified via qPCR. HDAC3 silencing at the beginning of differentiation enhanced adipocyte functionality by amplifying the expression of genes regulating differentiation, oxidative metabolism, browning and mitochondrial activity, starting from 72 h after induction of differentiation and silencing. Insulin signaling was enhanced as demonstrated by increased AKT phosphorylation following HDAC3 silencing. Mitochondrial content/density did not change, while the increased expression of the transcriptional co-activator Ppargc1b suggests the observed phenotype was related to enhanced mitochondrial activity, which was confirmed by increased maximal respiration and proton leak linked to reduced coupling efficiency. Moreover, the expression of pro-inflammatory markers increased with HDAC3 early silencing. To the contrary, no differences in terms of gene expression were found when HDAC3 silencing occurred in terminally differentiated adipocyte. Our data demonstrated that early epigenetic events mediated by class I HDAC inhibition/silencing are crucial to commit adipocyte precursors towards the above-mentioned metabolic phenotype. Moreover, our data suggest that these effects are exerted on adipocyte precursors.

Obesity is characterized by uncontrolled expansion of adipose tissue mass, resulting in adipocyte hypertrophy (increased adipocyte size) and hyperplasia (increased number of adipocytes). The number of adipose cells is directly related to adipocyte differentiation process from stromal vascular cells to mature adipocytes. It is known that epigenetic factors influence adipose differentiation program. However, how specific epigenome modifiers affect white adipocyte differentiation and metabolic phenotype is still matter of research. Here, we provide evidence that class I histone deacetylases (HDACs) are involved both in the differentiation of adipocytes and in determining the metabolic features of these cells. We demonstrate that inhibition of class I HDACs from the very first stage of differentiation amplifies the differentiation process and imprints cells toward a highly oxidative phenotype. These effects are related to the capacity of the inhibitor to modulate H3K27 acetylation on enhancer regions regulating Pparg and Ucp1 genes. These epigenomic modifications result in improved white adipocyte functionality and metabolism and induce browning. Collectively, our results show that modulation of class I HDAC activity regulates the metabolic phenotype of white adipocytes via epigenetic imprinting on a key histone mark.

Diets low in carbohydrates and proteins and enriched in fat stimulate the hepatic synthesis of ketone bodies (KB). These molecules are used as alternative fuel for energy production in target tissues. The synthesis and utilization of KB are tightly regulated both at transcriptional and hormonal levels. The nuclear receptor peroxisome proliferator activated receptor α (PPARα), currently recognized as one of the master regulators of ketogenesis, integrates nutritional signals to the activation of transcriptional networks regulating fatty acid β-oxidation and ketogenesis. New factors, such as circadian rhythms and paracrine signals, are emerging as important aspects of this metabolic regulation. However, KB are currently considered not only as energy substrates but also as signaling molecules. β-hydroxybutyrate has been identified as class I histone deacetylase inhibitor, thus establishing a connection between products of hepatic lipid metabolism and epigenetics. Ketogenic diets (KD) are currently used to treat different forms of infantile epilepsy, also caused by genetic defects such as Glut1 and Pyruvate Dehydrogenase Deficiency Syndromes. However, several researchers are now focusing on the possibility to use KD in other diseases, such as cancer, neurological and metabolic disorders. Nonetheless, clear-cut evidence of the efficacy of KD in other disorders remains to be provided in order to suggest the adoption of such diets to metabolic-related pathologies.

The metabolic transition from anaerobic glycolysis and fatty acid β-oxidation to glycolysis coupled to oxidative phosphorylation is a key process for the transition of quiescent neural stem cells to proliferative neural progenitor cells. However, a full characterization of the metabolic shift and the involvement of mitochondria during the last step of neurogenesis, from neuroblasts to neuron maturation, is still elusive. Here, we describe a model of neuroblasts, Neuro2a cells, with impaired differentiation capacity due to mitochondrial dysfunction. Using a detailed biochemical characterization consisting of steady-state metabolomics and metabolic flux analysis, we find increased fatty acid β-oxidation as a peculiar feature of neuroblasts with altered mitochondria. The consequent metabolic switch favors neuroblast proliferation at the expense of neuron maturation.

In the present study we chemically profiled tannin-enriched extracts from strawberries and tested their biological properties in a cell model of gastric inflammation. The chemical and biological features of strawberry tannins after in vitro simulated gastric digestion were investigated as well. The anti-inflammatory activities of pure strawberry tannins were assayed to get mechanistic insights. Tannin-enriched extracts from strawberries inhibit IL-8 secretion in TNFα-treated human gastric epithelial cells by dampening the NF-κB signaling. In vitro simulated gastric digestion slightly affected the chemical composition and the biological properties of strawberry tannins. By using pure compounds, we found that casuarictin may act as a pure NF-κB inhibitor while agrimoniin inhibits IL-8 secretion also acting on other biological targets; in our system procyanidin B1 prevents the TNFα-induced effects without interfering with the NF-κB pathway. We conclude that strawberry tannins, even after in vitro simulated gastric digestion, exert anti-inflammatory activities at nutritionally relevant concentrations.

Mitochondria represent the energy hub of cells and their function is under the constant influence of their tethering with other subcellular organelles. Mitochondria interact with the endoplasmic reticulum, lysosomes, cytoskeleton, peroxisomes, and nucleus in several ways, ranging from signal transduction, vesicle transport, and membrane contact sites, to regulate energy metabolism, biosynthetic processes, apoptosis, and cell turnover. Tumorigenesis is often associated with mitochondrial dysfunction, which could likely be the result of an altered interaction with different cell organelles or structures. The purpose of the present review is to provide an updated overview of the links between inter-organellar communications and interactions and metabolism in cancer cells, with a focus on mitochondria. The very recent publication of several reviews on these aspects testifies the great interest in the area. Here, we aim at (1) summarizing recent evidence supporting that the metabolic rewiring and adaptation observed in tumors deeply affect organelle dynamics and cellular functions and vice versa; (2) discussing insights on the underlying mechanisms, when available; and (3) critically presenting the gaps in the field that need to be filled, for a comprehensive understanding of tumor cells’ biology. Chemo-resistance and druggable vulnerabilities of cancer cells related to the aspects mentioned above is also outlined.

Gastritis is a widely spread inflammatory disease, mostly caused by Helicobacter pylori infection. Release of IL-8 by the stomach epithelium is a hallmark of gastritis and contributes to the amplification of the inflammatory state. Pharmacological modulation of IL-8 release is a strategy to relieve gastric inflammation and prevent more severe clinical outcomes. In search of nutraceuticals with potential anti-gastritis properties we used a bio-guided approach based on IL-8 secretion by gastric cells to characterize extracts from the fruits of different chestnut varieties. We found that the ability to inhibit IL-8 secretion correlated with the amount of proanthocyanidins and was associated to the not edible parts of chestnut in all the tested varieties. We also found that the anti-inflammatory activity is preserved upon mild thermal treatment and after in vitro simulated gastric digestion. By combining a robust bio-guided approach with a comprehensive analysis of the tannin fraction of chestnut extracts, we provide evidence for the potential use of chestnut-based nutraceuticals in human gastritis. The bioactive components of chestnut fruits inhibit IL-8 secretion by impairing NF-κB signaling and by other mechanisms, thus opening new applications of proanthocyanidins for inflammation-based diseases.

In this study, we investigated the ability of a phenolic extract from extra virgin olive oil (OPE) to modulate the inflammatory response in intestinal epithelial cells. Undifferentiated and differentiated Caco-2 cells were challenged with LPS (50 μg/mL) or IL-1β (5 ng/mL) to mimic the early and intermediate phase of intestinal inflammation, respectively. The effects of OPE on nuclear factor-κB-driven transcription and IL-8 promoter activity were evaluated in transfection assays, coupled to p65 nuclear translocation. Modulation of IL-8 mRNA levels by OPE was measured by quantitative RT-PCR while effects on protein levels by ELISA. Specific mitogen activated protein kinases inhibitors were used to investigate mRNA stability and the involvement of related signaling pathways. OPE prevented IL-8 expression and secretion in LPS-treated Caco-2 cells. In the presence of IL-1β OPE exhibited opposing effects on IL-8 gene transcription and mRNA/protein levels. While in IL-1β-treated cells IL-8 promoter activity was inhibited by treatment with OPE, IL-8 mRNA stability was strongly enhanced, leading to increased protein expression. Inhibitors of p38 and extracellular signal-regulated kinases partly prevented OPE effect on IL-8 mRNA levels. Intestinal epithelial cells represent a direct target of the action of olive oil phenols where they regulate IL-8 expression by transcriptional and posttranscriptional mechanisms.

Energy metabolism and mitochondrial function hold a core position in cellular homeostasis. Oxidative metabolism is regulated at multiple levels, ranging from gene transcription to allosteric modulation. To accomplish the fine tuning of these multiple regulatory circuits, the nuclear and mitochondrial compartments are tightly and reciprocally controlled. The fact that nuclear encoded factors, PPARγ coactivator 1α and mitochondrial transcription factor A, play pivotal roles in the regulation of oxidative metabolism and mitochondrial biogenesis is paradigmatic of this crosstalk. Here we provide an updated survey of the genetic and epigenetic mechanisms involved in the control of energy metabolism and mitochondrial function. Chromatin dynamics highly depends on post-translational modifications occurring at specific amino acids in histone proteins and other factors associated to nuclear DNA. In addition to the well characterized enzymes responsible for histone methylation/demethylation and acetylation/deacetylation, other factors have gone on the “metabolic stage”. This is the case of the new class of α-ketoglutarate-regulated demethylases (Jumonji C domain containing demethylases) and of the NAD+-dependent deacetylases, also known as sirtuins. Moreover, unexpected features of the machineries involved in mitochondrial DNA (mtDNA) replication and transcription, mitochondrial RNA processing and maturation have recently emerged. Mutations or defects of any component of these machineries profoundly affect mitochondrial activity and oxidative metabolism. Finally, recent evidences support the importance of mtDNA packaging in replication and transcription. These observations, along with the discovery that non-classical CpG islands present in mtDNA undergo methylation, indicate that epigenetics also plays a role in the regulation of the mitochondrial genome function.

Lipids in the nervous system accomplish a great number of key functions, from synaptogenesis to impulse conduction, and more. Most of the lipids of the nervous system are localized in myelin sheaths. It has long been known that myelin structure and brain homeostasis rely on specific lipid-protein interactions and on specific cell-to-cell signaling. In more recent years, the growing advances in large-scale technologies and genetically modified animal models have provided valuable insights into the role of lipids in the nervous system. Key findings recently emerged in these areas are here summarized. In addition, we briefly discuss how this new knowledge can open novel approaches for the treatment of diseases associated with alteration of lipid metabolism/homeostasis in the nervous system. This article is part of a Special Issue entitled Linking transcription to physiology in lipidomics.

Mitochondria are the energy-generating hubs of the cell. In spite of considerable advances, our understanding of the factors that regulate the molecular circuits that govern mitochondrial function remains incomplete. Using a genome-wide functional screen, we identify the poorly characterized protein Zinc finger CCCH-type containing 10 (Zc3h10) as regulator of mitochondrial physiology. We show that Zc3h10 is upregulated during physiological mitochondriogenesis as it occurs during the differentiation of myoblasts into myotubes. Zc3h10 overexpression boosts mitochondrial function and promotes myoblast differentiation, while the depletion of Zc3h10 results in impaired myoblast differentiation, mitochondrial dysfunction, reduced expression of electron transport chain (ETC) subunits, and blunted TCA cycle flux. Notably, we have identified a loss-of-function mutation of Zc3h10 in humans (Tyr105 to Cys105) that is associated with increased body mass index, fat mass, fasting glucose, and triglycerides. Isolated peripheral blood mononuclear cells from individuals homozygotic for Cys105 display reduced oxygen consumption rate, diminished expression of some ETC subunits, and decreased levels of some TCA cycle metabolites, which all together derive in mitochondrial dysfunction. Taken together, our study identifies Zc3h10 as a novel mitochondrial regulator.

Lipidomics and metabolomics have emerged as important tools for the characterization of specific physiological and pathological traits. The selection of the analytical approaches and the choice of a targeted rather than an untargeted strategy in metabolomics find their reasons in the driving hypothesis of the study, sample features and instrumental availability. Moreover, in the last years, -omics approaches have found their application in the study of sex-related dimorphism. In this review, lipidomic and metabolomic analyses are presented in a biomedical perspective. Here, we provide an updated overview covering recent applications of metabolomics and lipidomics in the area of sex-related differences in human and preclinical models. Experimental evidence underlines that sex is one of the most relevant biological variables significantly influencing metabolomic and lipidomic profiles. This knowledge can be exploited for the identification of novel sex-specific biomarkers and innovative targets relevant for gender medicine.

White adipose tissue (WAT) can undergo a phenotypic switch, known as browning, in response to environmental stimuli such as cold. Post-translational modifications of histones have been shown to regulate cellular energy metabolism, but their role in white adipose tissue physiology remains incompletely understood. Here we show that histone deacetylase 3 (HDAC3) regulates WAT metabolism and function. Selective ablation of Hdac3 in fat switches the metabolic signature of WAT by activating a futile cycle of de novo fatty acid synthesis and β-oxidation that potentiates WAT oxidative capacity and ultimately supports browning. Specific ablation of Hdac3 in adipose tissue increases acetylation of enhancers in Pparg and Ucp1 genes, and of putative regulatory regions of the Ppara gene. Our results unveil HDAC3 as a regulator of WAT physiology, which acts as a molecular brake that inhibits fatty acid metabolism and WAT browning., Histone deacetylases, such as HDAC3, have been shown to alter cellular metabolism in various tissues. Here the authors show that HDAC3 regulates WAT metabolism by activating a futile cycle of fatty acid synthesis and oxidation, which supports WAT browning.

Diabetic peripheral neuropathy causes a decrease in the levels of dihydroprogesterone and 5α-androstane-3α,17β-diol (3α-diol) in the peripheral nerves. These two neuroactive steroids exert protective effects, by mechanisms that still remain elusive. We have previously shown that the activation of Liver X Receptors improves the peripheral neuropathic phenotype in diabetic rats. This protective effect is accompanied by the restoration to control values of the levels of dihydroprogesterone and 3α-diol in peripheral nerves. In addition, activation of these receptors decreases peripheral myelin abnormalities by improving the lipid desaturation capacity, which is strongly blunted by diabetes, and ultimately restores the myelin lipid profile to non-diabetic values. On this basis, we here investigate whether dihydroprogesterone or 3α-diol may exert their protective effects by modulating the myelin lipid profile. We report that both neuroactive steroids act on the lipogenic gene expression profile in the sciatic nerve of diabetic rats, reducing the accumulation of myelin saturated fatty acids and promoting desaturation. These changes were associated with a reduction in myelin structural alterations. These findings provide evidence that dihydroprogesterone and 3α-diol are protective agents against diabetic peripheral neuropathy by regulating the de novo lipogenesis pathway, which positively influences myelin lipid profile.

After the completion of the human genome sequence and that from many other organisms, last decade has witnessed a spectacular gain of knowledge on gene functions. These studies provided new insights on the roles of genes in physiology and disease. Nonetheless, the availability of genetically modified models and of "omics" technologies such as next generation sequencing unveiled clear evidences on epigenetic regulation of many cellular functions. At this regard, sirtuins, belonging to class III histone deacetylase family, have emerged as regulators of metabolism as well as other cellular processes and seem ideally suited as targets of future therapeutical interventions. This review deals on general aspects of the biology of sirtuins and focuses on their relevance in lipid metabolism in different tissues, pointing to their exploitation as potential pharmacological targets of compounds that could be used as new therapeutic alternatives in several disorders ranging from type 2 diabetes and obesity to age-related cardiovascular and neurodegenerative diseases.

Chromatin modifications are sensitive to environmental and nutritional stimuli. Abnormalities in epigenetic regulation are associated with metabolic disorders such as obesity and diabetes that are often linked with defects in oxidative metabolism. Here, we evaluated the potential of class-specific synthetic inhibitors of histone deacetylases (HDACs), central chromatin-remodeling enzymes, to ameliorate metabolic dysfunction. Cultured myotubes and primary brown adipocytes treated with a class I–specific HDAC inhibitor showed higher expression of Pgc-1α, increased mitochondrial biogenesis, and augmented oxygen consumption. Treatment of obese diabetic mice with a class I– but not a class II–selective HDAC inhibitor enhanced oxidative metabolism in skeletal muscle and adipose tissue and promoted energy expenditure, thus reducing body weight and glucose and insulin levels. These effects can be ascribed to increased Pgc-1α action in skeletal muscle and enhanced PPARγ/PGC-1α signaling in adipose tissue. In vivo ChIP experiments indicated that inhibition of HDAC3 may account for the beneficial effect of the class I–selective HDAC inhibitor. These results suggest that class I HDAC inhibitors may provide a pharmacologic approach to treating type 2 diabetes.

Tm7sf2 gene encodes 3beta-hydroxysterol Delta(14)-reductase (C14SR, DHCR14), an endoplasmic reticulum enzyme acting on Delta(14)-unsaturated sterol intermediates during the conversion of lanosterol to cholesterol. The C-terminal domain of lamin B receptor, a protein of the inner nuclear membrane mainly involved in heterochromatin organization, also possesses sterol Delta(14)-reductase activity. The subcellular localization suggests a primary role of C14SR in cholesterol biosynthesis. To investigate the role of C14SR and lamin B receptor as 3beta-hydroxysterol Delta(14)-reductases, Tm7sf2 knockout mice were generated and their biochemical characterization was performed. No Tm7sf2 mRNA was detected in the liver of knockout mice. Neither C14SR protein nor 3beta-hydroxysterol Delta(14)-reductase activity were detectable in liver microsomes of Tm7sf2((-/-)) mice, confirming the effectiveness of gene inactivation. C14SR protein and its enzymatic activity were about half of control levels in the liver of heterozygous mice. Normal cholesterol levels in liver membranes and in plasma indicated that, despite the lack of C14SR, Tm7sf2((-/-)) mice are able to perform cholesterol biosynthesis. Lamin B receptor 3beta-hydroxysterol Delta(14)-reductase activity determined in liver nuclei showed comparable values in wild-type and knockout mice. These results suggest that lamin B receptor, although residing in nuclear membranes, may contribute to cholesterol biosynthesis in Tm7sf2((-/-)) mice. Affymetrix microarray analysis of gene expression revealed that several genes involved in cell-cycle progression are downregulated in the liver of Tm7sf2((-/-)) mice, whereas genes involved in xenobiotic metabolism are upregulated.

Mammals dispose of cholesterol mainly through 7alpha-hydroxylated bile acids, and the enzyme catalyzing the 7alpha-hydroxylation, cholesterol 7alpha-hydroxylase (CYP7A1), has a deep impact on cholesterol homeostasis. In this review, we present the study of regulation of CYP7A1 as a good exemplification of the extraordinary contribution of molecular biology to the advancement of our understanding of metabolic pathways that has taken place in the last 2 decades. Since the cloning of the gene from different species, experimental evidence has accumulated, indicating that the enzyme is mainly regulated at the transcriptional level and that bile acids are the most important physiological inhibitors of CYP7A1 transcription. Multiple mechanisms are involved in the control of CYP7A1 transcription and a variety of transcription factors and nuclear receptors participate in sophisticated regulatory networks. A higher order of transcriptional regulation, stemming from the so-called histone code, also applies to CYP7A1, and recent findings clearly indicate that chromatin remodelling events have profound effects on its expression. CYP7A1 also acts as a sensor of signals coming from the gut, thus representing another line of defence against the toxic effects of bile acids and a downstream target of agents acting at the intestinal level. From the pharmacological point of view, bile acid binding resins were the first primitive approach targeting the negative feed-back regulation of CYP7A1 to reduce plasma cholesterol. In recent years, new drugs have been designed based on recent discoveries of the regulatory network, thus confirming the position of CYP7A1 as a focus for innovative pharmacological intervention.

Background Cholesterol elimination occurs through bile acid synthesis that starts within the liver from 7α-hydroxylation or in extrahepatic tissues from 27-hydroxylation. This study was aimed at investigating in vivo these two pathways in patients with chronic liver disease. Methods Serum concentrations of 7α- and 27-hydroxycholesterol were measured in 54 patients (29 with primary biliary cirrhosis and 25 with chronic hepatitis C) and 18 controls. The rate of oxysterol plasma appearance was calculated after intravenous infusions of deuterated 7α- and 27-hydroxycholesterol in patients ( n = 8) and control subjects ( n = 8) who gave consent. The expression of sterol 27-hydroxylase was evaluated in macrophages isolated from 20 subjects. Results In patients with liver disease, the rate of plasma appearance of 7α-hydroxycholesterol was significantly reduced (1.44 ± 0.96 vs . 2.75 ± 1.43 mg/hour, p = 0.03), the degree of reduction being related with the severity of the disease ( p = 0.01) whereas that of 27-hydroxycholesterol was unaffected. The rate of plasma appearance of 27-hydroxycholesterol was significantly related to its serum concentrations ( r = 0.54, p = 0.03) and to its release from cultured macrophages ( r = 0.85, p = 0.03). Conclusions In liver disease 7α-hydroxylation of cholesterol seems to be impaired while 27-hydroxylation is unaffected. Serum concentrations of 27-hydroxycholesterol are useful to obtain information on the activity of this alternative pathway.

Myelin is a membrane characterized by high lipid content to facilitate impulse propagation. Changes in myelin fatty acid (FA) composition have been associated with peripheral neuropathy [1], but the specific role of peripheral nerve FA synthesis in myelin formation and function is poorly understood. We explored the extent to which lack of the key regulator of FA synthesis as Sterol Regulatory Element Binding Factor-1c (Srebf-1c) could result in the development of peripheral neuropathy. We found that Srebf-1c null mice display a neuropathic phenotype consisting in hypermyelinated small caliber fibers, the result of changes in myelin periodicity. Unexpectedly, transcriptomics and metabolomics revealed activation of peroxisome proliferator activated receptor α (Pparα) signaling in Srebf-1c null peripheral nerve as a result of increased levels of two distinct phosphatidylcholine-based Pparα ligands, PC-C16:0/C18:1 and PC-C18:0/C18:1 [2, 3]. Pparα is a nuclear receptor that directs uptake, utilization and catabolism of FAs [4]. As a consequence of abnormal local Pparα activation, Srebf-1c null peripheral nerve exhibit increased fatty acid utilization, a detrimental condition leading to peripheral neuropathy. Treatment with a Pparα antagonist rescues the neuropathy of Srebf-1c null mice. These findings reveal the importance of FA synthesis to sustain peripheral nerve structure and function.

In its 46th year, the International Conference on the Bioscience of Lipids (ICBL) landed in Corsica for the first time, at the end of September 2005. The first ICBL meeting dates back to 1953; since then, conferences have been held in different venues, normally chosen from among European countries. Despite their European roots, ICBL conferences are conceived to offer scientists interested in the field of lipid bioscience the opportunity to gather together from all over the world. In fact, ICBL meetings attract scientists not only from Europe, but also from the USA, South America, in particular Argentina, and Japan. In recent years, the conference has adopted a 4-day format consisting of in 1-day or halfday scientific sessions dedicated to specific topics. They range from classic themes in lipid biochemistry to innovative aspects, combining lipid science with medical sciences, biotechnology and ‘omics’ approaches. According to the usual format, each session includes contributions in the form of plenary lectures delivered by invited speakers and oral communications selected from submitted abstracts. The 2005 edition, the first in the ICBL history to be organized by two countries, was co-chaired by Maurizio Crestani from Italy and Paul Grimaldi and Michele Lagarde from France, with the support of a joint French–Italian organizing committee. The conference opened on the evening of September 20th with the 9th LLM Van Deenen Lecture, a special section introduced in 1997 to commemorate the distinguished Dutch lipidologist who passed away in 1994. The lipid scientist selected for the lecture this year was Alan A Spector, distinguished Professor at the University of Iowa, IA, USA, who delivered a lecture entitled ‘Fatty acids and derivatives in cell function’. In his presentation, Spector revisited the biochemistry and biology of the bioactive metabolites derived from arachidonic acid. He provided an excellent example of how a scientific topic is developed, applying typical biochemical tools and novel concepts borrowed from molecular biology. In fact, he described the multiple steps that led him to the isolation and structural identification of arachidonic acid metabolites, especially those produced by cytochrome P450-dependent monooxygenases, along with the identification of the enzymatic source and metabolic fate. He concluded the description of these metabolites by describing their multifaceted biological properties, from vasocontraction/relaxation to the modulation of the activity of nuclear receptors. This year the conference program also included a special lecture dedicated to Giovanni Galli, Professor of Biochemistry at the Department of Pharmacological Sciences, University of Milano, Italy, who served as secretary of ICBL for over 20 years and passed away suddenly in 2003. Alberico L Catapano, a colleague from the same department at the University of Milano and a friend of Galli, presented the major contributions of his research group in the lipoprotein field with a speech entitled ‘Plasma lipoproteins and endothelial function: from gene expression to biological relevance’. Catapano focused in particular on triglyceride-rich and highdensity lipoproteins whose effects are definitely less well elucidated than those exerted by low-density lipoproteins. By combining global approaches, such as cDNA microarray analysis and molecular approaches that allow pathways modulating gene expression to be dissected, Catapano presented evidence that lipoproteins may affect endothelial function, including cross-talk with proinflammatory and prothrombotic pathways. These studies significantly advance our understanding of the role that lipoproteins play in atherosclerosis.

Cellular homeostasis is maintained through a complicated network of signaling, transport and enzymatic events that take place in different compartments of the cell. The result of this composite network determines the cellular behavior in response to environmental challenges. Within this network, many cellular functions are regulated at the transcriptional level and consequently the comprehension of the molecular mechanisms of gene regulation is fundamental to fully understand how the cell reacts to environmental changes. Moreover, it offers an opportunity to find novel targets for the design of better strategies to improve healthy human nutrition and to treat metabolic diseases. In this framework, nuclear receptors have emerged as key regulators of many cellular functions in response to lipid action. In this review, we will discuss the new concepts on the biology of the recently “adopted” nuclear receptors sensing oxysterols (LXR), bile acids (FXR) and fatty acids (PPARs) and their function in the integrated regulation of lipid and glucose metabolism. We will also address the role played by other orphan nuclear receptors, such as FTF, SHP and HNF-4, acting in concert with these lipid-sensing nuclear receptors in the regulation of cellular metabolism.

Myelin is a membrane characterized by high lipid content to facilitate impulse propagation. Changes in myelin fatty acid (FA) composition have been associated with peripheral neuropathy, but the specific role of peripheral nerve FA synthesis in myelin formation and function is poorly understood. We have found that mice lacking sterol regulatory element-binding factor-1c (Srebf1c) have blunted peripheral nerve FA synthesis that results in development of peripheral neuropathy. Srebf1c-null mice develop Remak bundle alterations and hypermyelination of small-caliber fibers that impair nerve function. Peripheral nerves lacking Srebf1c show decreased FA synthesis and glycolytic flux, but increased FA catabolism and mitochondrial function. These metabolic alterations are the result of local accumulation of two endogenous peroxisome proliferator-activated receptor-α (Pparα) ligands, 1-palmitoyl-2-oleyl-sn-glycerol-3-phosphatidylcholine and 1-stearoyl-2-oleyl-sn-glycerol-3-phosphatidylcholine. Treatment with a Pparα antagonist rescues the neuropathy of Srebf1c-null mice. These findings reveal the importance of peripheral nerve FA synthesis to sustain myelin structure and function., These studies were supported by funding from Giovanni Armenise-Harvard Foundation Career Development Grant (N.M.), Fondazione CARIPLO 2014-0991 (N.M.), Fondazione CARIPLO 2012-0547 (R.C.M.), Italian Ministry of Health GR-2011-02346791 (M.D. and N.M.) and Research Center for the Characterization and Safe Use of Natural Compounds—“Giovanni Galli” directed by D.C. S.S. is an employee and founder of DASP s.r.l.; all other authors declare no competing financial interests.

After the completion of the human genome sequence and that from many other organisms, last decade has witnessed a spectacular gain of knowledge on gene functions. These studies provided new insights on the roles of genes in physiology and disease. Nonetheless, the availability of genetically modified models and of "omics" technologies such as next generation sequencing unveiled clear evidences on epigenetic regulation of many cellular functions. At this regard, sirtuins, belonging to class III histone deacetylase family, have emerged as regulators of metabolism as well as other cellular processes and seem ideally suited as targets of future therapeutical interventions. This review deals on general aspects of the biology of sirtuins and focuses on their relevance in lipid metabolism in different tissues, pointing to their exploitation as potential pharmacological targets of compounds that could be used as new therapeutic alternatives in several disorders ranging from type 2 diabetes and obesity to age-related cardiovascular and neurodegenerative diseases.

Peroxisome proliferator-activated receptors (PPARs) are ligand-dependent transcription factors regulating lipid and glucose metabolism. Ongoing drug discovery programs aim to develop dual PPARα/γ agonists devoid of the side effects of the marketed antidiabetic agents thiazolidinediones and the dual agonists glitazars. Recently, we described a new dual PPARα/γ ligand, LT175, with a partial agonist profile against PPARγ and interacting with a newly identified region of the PPARγ-ligand binding domain (1). Here we show that LT175 differentially activated PPARγ target genes involved in fatty acid esterification and storage in 3T3-L1-derived adipocytes. This resulted in a less severe lipid accumulation compared with that triggered by rosiglitazone, suggesting that LT175 may have a lower adipogenic activity. Consistent with this hypothesis, in vivo administration of LT175 to mice fed a high-fat diet decreased body weight, adipocyte size, and white adipose tissue mass, as assessed by magnetic resonance imaging. Furthermore, LT175 significantly reduced plasma glucose, insulin, non-esterified fatty acids, triglycerides, and cholesterol and increased circulating adiponectin and fibroblast growth factor 21 levels. Oral glucose and insulin tolerance tests showed that the compound improves glucose homeostasis and insulin sensitivity. Moreover, we demonstrate that the peculiar interaction of LT175 with PPARγ affected the recruitment of the coregulators cyclic-AMP response element-binding protein-binding protein and nuclear corepressor 1 (NCoR1), fundamentals for the PPARγ-mediated adipogenic program. In conclusion, our results describe a new PPAR ligand, modulating lipid and glucose metabolism with reduced adipogenic activity, that may be used as a model for a series of novel molecules with an improved pharmacological profile for the treatment of dyslipidemia and type 2 diabetes.

Bile acids regulate the cholesterol 7alpha-hydroxylase gene (CYP7A1), which encodes the rate-limiting enzyme in the classical pathway of bile acid synthesis. Here we report a novel mechanism whereby bile acid feedback regulates CYP7A1 transcription through the nuclear receptor hepatocyte nuclear factor-4 (HNF-4), which binds to the bile acid response element (BARE) at nt -149/-118 relative to the transcription start site. Using transient transfection assays of HepG2 cells with Gal4-HNF-4 fusion proteins, we show that chenodeoxycholic acid (CDCA) dampened the transactivation potential of HNF-4. Overexpression of a constitutive active form of MEKK1, an upstream mitogen-activated protein kinase (MAPK) module triggered by stress signals, strongly repressed the promoter activity of CYP7A1 via the consensus sequence for HNF-4 embedded in the BARE. Similarly, MEKK1 inhibited the activity of HNF-4 in the Gal4-based assay. The involvement of the MEKK1-dependent pathway in the bile acid-mediated repression of CYP7A1 was confirmed by co-transfecting a dominant negative form of the stress-activated protein kinase kinase, SEK, which abolished the effect of CDCA upon CYP7A1 transcription. Treatment of transfected HepG2 cells with tumor necrosis factor alpha (TNF-alpha), an activator of the MEKK1 pathway, led to the repression of CYP7A1 via the HNF-4 site in the BARE. TNF-alpha also inhibited the transactivation potential of HNF-4. Collectively, our results demonstrate for the first time that HNF-4, in combination with a MAPK signaling pathway, acts as a bile acid sensor in the liver. Furthermore, the effects of CDCA and TNF-alpha converge to HNF-4, which binds to the BARE of CYP7A1, suggesting a link between the cascades elicited by bile acids and pro-inflammatory stimuli in the liver.

Regulation of bile acid synthesis, a key determinant of cholesterol homeostasis, is still incompletely understood. To elucidate the feedback control exerted on bile acid biosynthesis in humans with obstructive cholestasis, 16 patients with bile duct obstruction were studied. In vivo 7alpha-hydroxylation, reflecting bile acid synthesis, was assayed in 13 of them by tritium release analysis. Serum 27-hydroxycholesterol was determined by gas chromatography-mass spectrometry. In a subgroup, hepatic cholesterol 7alpha-hydroxylase mRNA was assayed by real-time polymerase chain reaction (PCR), enzyme activity was determined by isotope incorporation, and microsomal cholesterol content was assayed by gas chromatography-mass spectrometry. Age-matched control subjects were studied in parallel. Hydroxylation rates were lower in cholestatic patients (108 +/- 33 mg of cholesterol per day, mean +/- SEM; controls: 297 +/- 40 mg/d; P

A number of recent studies revealed that epigenetic modifications play a central role in the regulation of lipid and of other metabolic pathways such as cholesterol homeostasis, bile acid synthesis, glucose and energy metabolism. Epigenetics refers to aspects of genome functions regulated in a DNA sequence-independent fashion. Chromatin structure is controlled by epigenetic mechanisms through DNA methylation and histone modifications. The main modifications are histone acetylation and deacetylation on specific lysine residues operated by two different classes of enzymes: Histone acetyltransferases (HATs) and histone deacetylases (HDACs), respectively. The interaction between these enzymes and histones can activate or repress gene transcription: Histone acetylation opens and activates chromatin, while deacetylation of histones and DNA methylation compact chromatin making it transcriptionally silent. The new evidences on the importance of HDACs in the regulation of lipid and other metabolic pathways will open new perspectives in the comprehension of the pathophysiology of metabolic disorders.

Diabetic peripheral neuropathy (DPN) is characterized by myelin abnormalities; however, the molecular mechanisms underlying such deficits remain obscure. To uncover the effects of diabetes on myelin alterations, we have analyzed myelin composition. In a streptozotocintreated rat model of diabetic neuropathy, analysis of sciatic nerve myelin lipids revealed that diabetes alters myelin's phospholipid, FA, and cholesterol content in a pattern that can modify membrane fluidity. Reduced expression of relevant genes in the FA biosynthetic pathway and decreased levels of the transcriptionally active form of the lipogenic factor sterol-regulatory element binding factor-1c (SREBF-1c) were found in diabetic sciatic nerve. Expression of myelin's major protein, myelin protein zero (P0), was also suppressed by diabetes. In addition, we confirmed that diabetes induces sciatic nerve myelin abnormalities, primarily infoldings that have previously been associated with altered membrane fluidity. In a diabetic setting, synthetic activator of the nuclear receptor liver X receptor (LXR) increased SREBF-1c function and restored myelin lipid species and P0 expression levels to normal. These LXR-modulated improvements were associated with restored myelin structure in sciatic nerve and enhanced performance in functional tests such as thermal nociceptive threshold and nerve conduction velocity. These findings demonstrate an important role for the LXR-SREBF-1c axis in protection from diabetes-induced myelin abnormalities. Copyright © 2012 by the American Society for Biochemistry and Molecular Biology, Inc.

Cholesterol and fatty acid moieties are the most abundant lipid species in the brain. The blood brain barrier (BBB), formed by tightly juxtaposed endothelial cells, regulates the accessibility of circulating lipids to brain cells. In physiological conditions lipoprotein cholesterol cannot enter the BBB. Cholesterol necessary for myelination and other functions (for example, synthesis of oxysterols and neuroactive steroids) is synthesized in situ by astrocytes and oligodendrocytes (not shown). In contrast, 24-hydroxycholesterol can cross the BBB, thus entering the systemic circulation. Fatty acids, the building blocks of many specialized lipids, derive from blood (albumin-associated free fatty acids) or are synthesized in situ. Notably, in brain cells the fraction of fatty acids subject to b-oxidation is rather small.

Lignan-rich whole-grain cereals, beans, berries, and nuts show protective effects against a variety of chronic diseases, including cancer. Lignans are converted by intestinal microflora to enterolactone (EL) and its oxidation product enterodiol (ED). To investigate the immunomodulatory effect of EL and ED in human cells, peripheral blood lymphocytes were treated with increasing physiologically relevant concentrations of EL and ED (0-1000 microM) and stimulated with lipopolysaccharide (LPS) and anti-CD3 plus anti-CD28 monoclonal antibodies. A dose-related inhibition of cell proliferation and cytokine production was observed, with EL being the most active. Molecular investigations in THP-1 cells showed that both EL and ED prevented inhibitory-kappaB (I-kappaB) degradation and nuclear factor-kappaB (NF-kappaB) activation, which in turn resulted in decreased tumor necrosis factor-alpha (TNF-alpha) production. EL and ED were also able to pass the intestinal barrier and modulate cytokine production. The findings of the present study reveal potential mechanisms that could explain some in vivo beneficial effects of lignans.

Cholesterol and other cholesterol related metabolites, oxysterols, and bile acids, establish specific interactions with enzymes and other proteins involved in cholesterol and bile acid homeostasis, triggering a variety of biological responses. The substrate-enzyme binding represents the best-characterized type of complementary interaction between proteins and small molecules. Key enzymes in the pathway that converts cholesterol to bile acids belong to the cytochrome P450 superfamily. In contrast to the majority of P450 enzymes, those acting on cholesterol and related metabolites exhibit higher stringency with respect to substrate molecules. This stringency, coupled with the specificity of the reactions, dictates the chemical features of intermediate metabolites (oxysterols) and end products (bile acids). Both oxysterols and bile acids have emerged in recent years as new signalling molecules due to their ability to interact and activate nuclear receptors, and consequently to regulate the transcription of genes involved in cholesterol and bile acid homeostasis and metabolism, but also in glucose and fatty acid metabolism. Interestingly, other proteins function as bile acid or sterol receptors. New findings indicate that bile acids also interact with a membrane G protein-coupled receptor, triggering a signalling cascade that ultimately promote energy expenditure. On the other end, cholesterol and side chain oxysterols establish specific interactions with different proteins residing in the endoplasmic reticulum that result in controlled protein degradation and/or trafficking to the Golgi and the nucleus. These regulatory pathways converge and contribute to adapt cholesterol uptake and synthesis to the cellular needs.

In vivo studies suggest that the phenolic component contributes to the anti-inflammatory and antiatherosclerotic actions of olive oil; however, the effects in circulating cells are not fully characterized. Monocytes play a key role in inflammation-based diseases by expressing several molecules, including metalloproteinases (MMPs). In the present study, we investigated the effects of olive oil phenolic extract and individual compounds on MMP-9 in THP-1 cells, a human monocyte-like cell line. Olive oil extract prevented the stimulation of MMP-9 expression and secretion in tumor necrosis factor alpha-treated THP-1 cells. Oleuropein aglycone, a typical olive oil phenol, was active at concentrations found in the extract, although other compounds probably contribute to the biological activity. We also found that the effect of the extract and individual compounds on MMP-9 is due to impaired nuclear factor-kappaB signaling. Our findings provide further evidence on the mechanisms by which olive oil reduces the inflammatory burden associated with disorders, such as atherosclerosis.

Cholesterol and other cholesterol related metabolites, oxysterols, and bile acids, establish specific interactions with enzymes and other proteins involved in cholesterol and bile acid homeostasis, triggering a variety of biological responses. The substrate-enzyme binding represents the best-characterized type of complementary interaction between proteins and small molecules. Key enzymes in the pathway that converts cholesterol to bile acids belong to the cytochrome P450 superfamily. In contrast to the majority of P450 enzymes, those acting on cholesterol and related metabolites exhibit higher stringency with respect to substrate molecules. This stringency, coupled with the specificity of the reactions, dictates the chemical features of intermediate metabolites (oxysterols) and end products (bile acids). Both oxysterols and bile acids have emerged in recent years as new signalling molecules due to their ability to interact and activate nuclear receptors, and consequently to regulate the transcription of genes involved in cholesterol and bile acid homeostasis and metabolism, but also in glucose and fatty acid metabolism. Interestingly, other proteins function as bile acid or sterol receptors. New findings indicate that bile acids also interact with a membrane G protein-coupled receptor, triggering a signalling cascade that ultimately promote energy expenditure. On the other end, cholesterol and side chain oxysterols establish specific interactions with different proteins residing in the endoplasmic reticulum that result in controlled protein degradation and/or trafficking to the Golgi and the nucleus. These regulatory pathways converge and contribute to adapt cholesterol uptake and synthesis to the cellular needs.

NAD(+)-dependent sirtuin deacetylases have emerged as potential therapeutic targets for treatment of human illnesses such as cancer, metabolic, cardiovascular, and neurodegenerative diseases. The benefits of sirtuin modulation by small molecules have been demonstrated for these diseases. In contrast to the discovery of inhibitors of SIRT1, -2, and -3, only activators for SIRT1 are known. Here, we rationalized the potential of the previously unexplored dihydropyridine scaffold in developing sirtuin ligands, thus we prepared a series of 1,4-dihydropyridine-based derivatives 1-3. Assessment of their SIRT1-3 deacetylase activities revealed the importance of the substituent at the N1 position of the dihydropyridine structure on sirtuin activity. Placement of cyclopropyl, phenyl, or phenylethyl groups at N1 conferred nonselective SIRT1 and SIRT2 inhibition activity, while a benzyl group at N1 conferred potent SIRT1, -2, and -3 activation. Senescence assays performed on hMSC and mitochondrial function studies conducted with murine C2C12 myoblasts confirmed the compounds' novel and unique SIRT-activating properties.

Tm7sf2 gene encodes 3beta-hydroxysterol Delta(14)-reductase (C14SR, DHCR14), an endoplasmic reticulum enzyme acting on Delta(14)-unsaturated sterol intermediates during the conversion of lanosterol to cholesterol. The C-terminal domain of lamin B receptor, a protein of the inner nuclear membrane mainly involved in heterochromatin organization, also possesses sterol Delta(14)-reductase activity. The subcellular localization suggests a primary role of C14SR in cholesterol biosynthesis. To investigate the role of C14SR and lamin B receptor as 3beta-hydroxysterol Delta(14)-reductases, Tm7sf2 knockout mice were generated and their biochemical characterization was performed. No Tm7sf2 mRNA was detected in the liver of knockout mice. Neither C14SR protein nor 3beta-hydroxysterol Delta(14)-reductase activity were detectable in liver microsomes of Tm7sf2((-/-)) mice, confirming the effectiveness of gene inactivation. C14SR protein and its enzymatic activity were about half of control levels in the liver of heterozygous mice. Normal cholesterol levels in liver membranes and in plasma indicated that, despite the lack of C14SR, Tm7sf2((-/-)) mice are able to perform cholesterol biosynthesis. Lamin B receptor 3beta-hydroxysterol Delta(14)-reductase activity determined in liver nuclei showed comparable values in wild-type and knockout mice. These results suggest that lamin B receptor, although residing in nuclear membranes, may contribute to cholesterol biosynthesis in Tm7sf2((-/-)) mice. Affymetrix microarray analysis of gene expression revealed that several genes involved in cell-cycle progression are downregulated in the liver of Tm7sf2((-/-)) mice, whereas genes involved in xenobiotic metabolism are upregulated.