Mammalian Energy Metabolism Research Articles

Purification and fermentation characteristics of exopolysaccharide from Fomitopsis castaneus Imaz

Short-chain fatty acids (SCFAs), which are the end products of carbohydrate fermentation in the gut, mainly contribute to energy metabolism in mammals. The amount of SCFAs produced during fermentation is an important parameter that characterizes the fermentation capacity of a system. This paper reports on the fermentation characteristics of exopolysaccharides (EPS) isolated from Fomitopsis castaneus Imaz, a wood-rot fungal species. We isolated and purified the main EPS fraction by freeze drying and DEAE-Sepharose fast flow chromatography. We then analyzed the monosaccharide composition of EPS. The isolated EPS was mainly composed of glucose, galactose, rhamnose, mannose, and arabinose. The characteristic absorption peaks of sugar esters were also detected. Fresh fecal extracts from healthy adults and children were used as fermentation substrate to simulate the human intestinal environment (anaerobic conditions at 37°C) and study the fermentation characteristics of the purified EPS. Adding the isolated EPS to the fermentation system of the simulated intestinal environment increased the SCFAs content in the fecal extract of adults and children. However, the yield of SCFAs, particularly butyric acid, in the fermentation system of fecal extract in children was higher than that in adults. Furthermore, adding exogenous lactic acid bacteria, such as Enterococcus fecalis and Enterococcus fecium, to the fermentation system effectively increased the SCFAs concentration in the model intestinal system of the children. By contrast, adding E. fecalis, Lactobacillus rhamnosus, and E. fecium increased the content of the produced SCFAs in the system of adults. Those results indicate that EPS isolated from F. castaneus Imaz was effectively fermented in the simulated intestinal environments, and the fermentation capability was enhanced by adding microbial flora.

Activation of SF1 Neurons in the Ventromedial Hypothalamus by DREADD Technology Increases Insulin Sensitivity in Peripheral Tissues.

The ventromedial hypothalamus (VMH) regulates glucose and energy metabolism in mammals. Optogenetic stimulation of VMH neurons that express steroidogenic factor 1 (SF1) induces hyperglycemia. However, leptin acting via the VMH stimulates whole-body glucose utilization and insulin sensitivity in some peripheral tissues, and this effect of leptin appears to be mediated by SF1 neurons. We examined the effects of activation of SF1 neurons with DREADD (designer receptors exclusively activated by designer drugs) technology. Activation of SF1 neurons by an intraperitoneal injection of clozapine-N-oxide (CNO), a specific hM3Dq ligand, reduced food intake and increased energy expenditure in mice expressing hM3Dq in SF1 neurons. It also increased whole-body glucose utilization and glucose uptake in red-type skeletal muscle, heart, and interscapular brown adipose tissue, as well as glucose production and glycogen phosphorylase a activity in the liver, thereby maintaining blood glucose levels. During hyperinsulinemic-euglycemic clamp, such activation of SF1 neurons increased insulin-induced glucose uptake in the same peripheral tissues and tended to enhance insulin-induced suppression of glucose production by suppressing gluconeogenic gene expression and glycogen phosphorylase a activity in the liver. DREADD technology is thus an important tool for studies of the role of the brain in the regulation of insulin sensitivity in peripheral tissues.

The promiscuous enzyme medium-chain 3-keto-acyl-CoA thiolase triggers a vicious cycle in fatty-acid beta-oxidation.

Mitochondrial fatty-acid beta-oxidation (mFAO) plays a central role in mammalian energy metabolism. Multiple severe diseases are associated with defects in this pathway. Its kinetic structure is characterized by a complex wiring of which the functional implications have hardly been explored. Repetitive cycles of reversible reactions, each cycle shortening the fatty acid by two carbon atoms, evoke competition between intermediates of different chain lengths for a common set of ‘promiscuous’ enzymes (enzymes with activity towards multiple substrates). In our validated kinetic model of the pathway, substrate overload causes a steep and detrimental flux decline. Here, we unravel the underlying mechanism and the role of enzyme promiscuity in it. Comparison of alternative model versions elucidated the role of promiscuity of individual enzymes. Promiscuity of the last enzyme of the pathway, medium-chain ketoacyl-CoA thiolase (MCKAT), was both necessary and sufficient to elicit the flux decline. Subsequently, Metabolic Control Analysis revealed that MCKAT had insufficient capacity to cope with high substrate influx. Next, we quantified the internal metabolic regulation, revealing a vicious cycle around MCKAT. Upon substrate overload, MCKAT’s ketoacyl-CoA substrates started to accumulate. The unfavourable equilibrium constant of the preceding enzyme, medium/short-chain hydroxyacyl-CoA dehydrogenase, worked as an amplifier, leading to accumulation of upstream CoA esters, including acyl-CoA esters. These acyl-CoA esters are at the same time products of MCKAT and inhibited its already low activity further. Finally, the accumulation of CoA esters led to a sequestration of free CoA. CoA being a cofactor for MCKAT, its sequestration limited the MCKAT activity even further, thus completing the vicious cycle. Since CoA is also a substrate for distant enzymes, it efficiently communicated the ‘traffic jam’ at MCKAT to the entire pathway. This novel mechanism provides a basis to explore the role of mFAO in disease and elucidate similar principles in other pathways of lipid metabolism.

Identification of Rare Variants in Metabolites of the Carnitine Pathway by Whole Genome Sequencing Analysis.

We use whole genome sequence data and rare variant analysis methods to investigate a subset of the human serum metabolome, including 16 carnitine-related metabolites that are important components of mammalian energy metabolism. Medium pass sequence data consisting of 12,820,347 rare variants and serum metabolomics data were available on 1,456 individuals. By applying a penalization method, we identified two genes FGF8 and MDGA2 with significant effects on lysine and cis-4-decenoylcarnitine, respectively, using Δ-AIC and likelihood ratio test statistics. Single variant analyses in these regions did not identify a single low-frequency variant (minor allele count > 3) responsible for the underlying signal. The results demonstrate the utility of whole genome sequence and innovative analyses for identifying candidate regions influencing complex phenotypes.

Response of AMP-activated protein kinase and energy metabolism to acute nitrite exposure in the Nile tilapia Oreochromis niloticus

Adenosine monophosphate-activated protein kinase (AMPK) is a prevalent mammalian energy metabolism sensor, but little is known about its role as an energy sensor in fish experiencing stress. We aimed to study AMPK in Oreochromis niloticus on both the molecular and the physical level. We found that the cDNAs encoding the AMPKα1 and AMPKα2 variants of the O. niloticus catalytic α subunit were 1753bp and 2563 bp long and encoded 571 and 557 amino acids, respectively. Both the AMPKα1 and the AMPKα2 isoform possess structural features similar to mammalian AMPKα, including a phosphorylation site at Thr172 in the N-terminus, and exhibit high homology with other fish and vertebrate AMPKα sequences (81.3%–98.1%). mRNA encoding the AMPKα isoforms was widely expressed in various tissues with distinctive patterns. AMPKα1 and AMPKα2 were primarily expressed in the intestines and brain, respectively. Under acute nitrite challenge, the mRNA encoding the AMPKα isoforms, as well as AMPK activity, changed over time. Its recovery period in freshwater, combined with the fact that it is highly conserved, suggests that fish AMPK, like its mammalian orthologues, acts as an energy metabolism sensor. Furthermore, subsequent decreases in AMPK mRNA levels and activity suggested that its action was transient but efficient. Physically, glucose, lactic acid and TGs in plasma, as well as energy materials in the hepatopancreas and muscle, were significantly altered over time, indicating changes in energy metabolism during the experimental period. These data have enabled us to characterize energy utilization in O. niloticus and further illustrate the role of fish AMPK as an energy sensor. This study provides new insight into energy metabolism and sensing by AMPK in teleost and necessitates further study of the multiple physiologic roles of AMPK in fish.

Autonomic nervous system-mediated effects of galanin-like peptide on lipid metabolism in liver and adipose tissue.

Galanin-like peptide (GALP) is a neuropeptide involved in the regulation of feeding behavior and energy metabolism in mammals. While a weight loss effect of GALP has been reported, its effects on lipid metabolism have not been investigated. The aim of this study was to determine if GALP regulates lipid metabolism in liver and adipose tissue via an action on the sympathetic nervous system. The respiratory exchange ratio of mice administered GALP intracerebroventricularly was lower than that of saline-treated animals, and fatty acid oxidation-related gene mRNA levels were increased in the liver. Even though the respiratory exchange ratio was reduced by GALP, this change was not significant when mice were treated with the sympatholytic drug, guanethidine. Lipolysis-related gene mRNA levels were increased in the adipose tissue of GALP-treated mice compared with saline-treated animals. These results show that GALP stimulates fatty acid β-oxidation in liver and lipolysis in adipose tissue, and suggest that the anti-obesity effect of GALP may be due to anorexigenic actions and improvement of lipid metabolism in peripheral tissues via the sympathetic nervous system.

THE IMPACT OF TOXIC STRESS ON THE MORPHOFUNCTIONAL STATE OF THE HYPOTHALAMIC ARCUATE NUCLEUS IN THE CONDITIONS OF THE CHANGED MODE OF LIGHT EXPOSURE

Aim . Arcuate nucleus (AN) of the hypothalamus is one of the integrative centers responsible for energy metabolism in mammals. Reaction of the arcuatus nucleus of white rats on the toxic stress caused by introduction of cadmium chloride in the conditions of natural illumination, light and dark deprivations was studied. Methods. The toxicant was entered per os at a dose of 2 mg on 100 g of body weight a daily for 15 days after one month of adaptation to an the artificial photomode. Synthetic activity was estimated a method nucleoli volumes’ measurement on hematoxylin and eosin sections of the hypothalamus. Results. As light and dark deprivations led to the reduction of the nucleoli volumes of AN at animals of both sexes. Salt of cadmium in the conditions of natural illumination has caused reduction of the nucleoli sizes, but only in males. On the contrary, on the background of dark deprivation cadmium chloride led to decrease in activity of AN only in females. Light deprivation promoted increase in synthetic activity of AN neurocytes under the influence of cadmium salt in males, but did not change that in females. Conclusion. The impact of toxic stress induced by administration of cadmium chloride on the hypothalamic centre responsible for energy metabolism of the organism depends on light intensity and sex of the animal. Inhibition of the synthetic activity of AN by cadmium chloride in natural lighting is more pronounced in males, and in the dark of deprivation it is more pronounced in females.

An essential role for Ewing sarcoma gene (EWS) in early white adipogenesis.

White adipose tissue is important for mammalian energy homeostasis and metabolism. It was previously demonstrated that Ewing sarcoma gene (EWS) is essential for early classical brown fat lineage determination, but its role in white adipocyte differentiation is not known. Mouse embryonic fibroblasts (MEFs) lacking Ews and shRNA-mediated silencing of Ews in 3T3L1 preadipocytes were used to investigate the role of EWS in adipogenesis. White fat differentiation was determined by analyzing the expression of key adipogenic genes and by Oil red O staining. Following adipogenic stimulation, Ews expression arose rapidly in 3T3L1 cells during early induction period. Ews-null MEFs and 3T3L1 cells with reduced Ews expression failed to undergo adipogenesis. This was accompanied by significant reduction in the expression of critical early adipogenic regulators, Bmp2, Bmp4 (bone morphogenic protein 2 and 4), Cebpβ, and Cebpδ (CCAAT/enhancer binding protein β and δ). Complementation of recombinant BMP2 or BMP4 partially rescued adipogenesis in Ews-depleted 3T3L1 cells. These results demonstrate that EWS is essential during the early steps of white adipocyte differentiation, at least in part through its regulation of BMP2 and BMP4 expression.

Molecular regulation of adiposity in a migrant, the gray catbird (1100.1)

Prior to their migration, birds make physiological and behavioral adjustments to quickly accumulate fat stores to fuel their journey. The majority of these fat stores are utilized during their long‐distance flights. However, birds often complete vernal migrations with residual fat, which may be detrimental due to impaired flight behavior and increased predation risk, thus return to lean body composition is also rapid. This need for swift and substantial changes in a bird’s physiology marks migration as a unique life history stage emphasizing phenotypic flexibility, defined as intra‐individual and reversible phenotypic transformations. Specific mechanisms directing stage transitions of adiposity in migrating birds are largely unknown. This study aims to characterize the role of peroxisome proliferator‐activated receptors (PPARs) in migratory adiposity of the gray catbird (Dumetella carolinensis). PPARs are a family of nuclear receptors involved in mammalian energy metabolism and are highly conserved in our study species. We find evidence of PPAR involvement in migratory adiposity with a coordinated increased rate of glycerol release from adipose tissue, a downstream PPAR process, during periods of migration compared to non‐migratory stages. These data support further investigation into PPAR target gene expression experiments, and advance our understanding of the regulation of stage transitions associated with migratory adiposity.

Epigallocatechin gallate counteracts oxidative stress in docosahexaenoxic acid-treated myocytes

Skeletal muscle is a key organ of mammalian energy metabolism, and its mitochondria are multifunction organelles that are targets of dietary bioactive compounds. The goal of this work was to examine the regulation of mitochondrial dynamics, functionality and cell energy parameters using docosahexaenoic acid (DHA), epigallocatechin gallate (EGCG) and a combination of both in L6 myocytes. Compounds (at 25μM) were incubated for 4h. Cells cultured with DHA displayed less oxygen consumption with higher ADP/ATP ratio levels concomitant with downregulation of Cox and Ant1 gene expression. The disruption of energetic homeostasis by DHA, increases intracellular reactive oxygen species (ROS) levels and decreases mitochondrial membrane potential. The defence mechanism to counteract the excess of ROS production was by the upregulation of Ucp2, Ucp3 and MnSod gene expression. Moreover myocytes cultured with DHA had a higher mitochondrial mass with a higher proportion of large and elongated mitochondria, whereas the fission genes Drp1 and Fiss1 and the fusion gene Mfn2 were downregulated. In myocytes co-incubated with DHA and EGCG, ROS levels and the adenosine diphosphate (ADP)/adenosine triphosphate (ATP) ratio were similar to untreated myocytes and the decrease of oxygen consumption, higher mitochondrial mass and the overexpression of Ucp2 and Ucp3 genes were similar to the DHA-treated cells with also a higher amount of mitochondrial deoxyribonucleic acid (DNA), and reduced Drp1 and Fiss1 gene expression levels. In conclusion the addition of EGCG to DHA returned the cells to the control conditions in terms of mitochondrial morphology, energy and redox status, which were unbalanced in the DHA-treated myocytes.

PPARs and Metabolic Syndrome.

Peroxisome proliferator-activated receptors (PPARs) exert versatile biological effects, notably in energy metabolism. During the last two decades, numerous studies have demonstrated that PPARs act as pivotal regulators of metabolic syndrome, a series of disorders in energy utilization and storage that are implicated with type 2 diabetes, diabetic nephropathy, and cardiovascular diseases, to mention a few. PPARα and PPARγ are the molecular targets of a number of marketed drugs for the treatment of these diseases, and accumulating evidence suggested PPARβ/δ as a potential therapeutic drug target as well. Although energy metabolism and metabolic syndrome are the most intensively studied domain of PPARs, it has not been addressed specifically in any issue of PPAR Research ever since its launch. Here, we gathered 3 reviews and 5 research articles that encompass metabolic syndrome and its complications. M. Aprile et al. tackled the subject of PPARγ and human adipogenesis in their research article. Rather than focusing on canonical PPARγ transcripts, authors largely emphasized on the critical contribution of PPARγ dominant negative isoforms to adipogenesis and their implied potential role in pathological conditions. In addition, a novel of PPARγ dominant negative transcript, γ1ORF4, was first identified in this study. In regard to nonalcoholic fatty liver diseases, the hepatic expression of the metabolic syndrome, M. Sharif et al. conducted a thorough analysis of previously published data about the steatogenic role of PPARγ and summarized two probable PPARγ ligand-dependent toxicological modes of action: (i) activation of PPARγ in hepatocytes and (ii) inhibition in adipocytes. Two papers, one review and a research article, by Z. Jia and Y. Sun et al., appraised the role of PPARγ in diabetic nephropathy (DN). Their comprehensive review summarized the limitations of traditional PPARγ agonists, addressed the advantages of newly developed PPARγ agonists, and rendered new insights into the therapeutic potential of PPARγ agonists in the treatment of DN, while the research article suggested that a combination of PPARγ agonists with COX-2/PGE2 inhibitors may be an alternative way of dealing with DN. In another research article, J. Jin et al. analyzed the correlation between PPAR gene polymorphisms and pediatric primary nephrotic syndrome (PNS) by comparing children with PNS against healthy subjects. They found that PPARγ (Pro12Ala) and PGC-1α (Gly482Ser) polymorphisms are associated with abnormal insulin and triglyceride metabolism in pediatric PNS patients, suggesting that these polymorphisms may be relevant to the prognosis of this chronic disease. The knowledge of the role of PPARα in metabolic disorder-associated cardiovascular diseases was well recognized in this special issue. Z. Jia et al.'s research article asserted the involvement of HMGB1 (high mobility group box 1) in the protective effect of PPARα in cardiac hypertrophy and provided a novel approach to study the pathogenesis of cardiac hypertrophy. Although most studies showed that PPARα activation confers protection against atherogenesis, the intriguing possibility that PPARα might foster atherogenesis is also considered. In this current issue, M. Vechoropoulos et al. found that PPARα mediates the proatherogenic effect of chronic nitric oxide synthesis inhibition and this effect is independent of blood pressure and serum lipids alterations. These data further shaped the view that the role of PPARα in atherosclerosis needs to be reevaluated. Lastly, in the review article “PPARs Integrate the Mammalian Clock and Energy Metabolism,” we collected recent findings about the role of PPARs in biological clocks. This brand new function of PPARs bridges energy metabolism with circadian rhythm whose relationship has been known for long time, but not well understood. We summarized the circadian function of three PPAR subtypes one by one and concluded that the abnormality of PPARs and circadian rhythm could impinge on each other and thus leads to metabolic disorders. Further investigation of PPARs in this field will give us a new perspective on the therapeutic advances in the treatment of metabolic ailments. In conclusion, this special issue is packed with intriguing novel breakthroughs and insights into PPARs and metabolic syndrome. We hope that these advances will generate more interest from the scientific community in better understanding of the role of PPARs in metabolic syndrome and associated complications. Lihong Chen Zhanjun Jia Guangrui Yang

I'm not Dead Yet: Flies and Mice

A decrease in the activity of the Indy gene (I'm not dead yet) in flies extends life span without a decline in fertility, physical activity, flight velocity or metabolic rate. Naturally occurring variants in Indy, worldwide, modify Indy expression and increase reproduction and longevity, indicating that alterations in Indy expression in the wild imparts significant improvement in fitness and longevity. The Indy gene codes for a high-affinity dicarboxylate/citrate plasma membrane transporter that is found most abundantly at the plasma membrane of adult fat body, oenocytes and midgut cells, the primary sites of intermediary metabolism in the fly. Decreases in Indy affect metabolism similarly to calorie restriction (CR). CR down-regulates Indy in normal flies, and Indy long-lived flies share phenotypes with long-lived CR flies that include decreased insulin-like signaling, lipid storage, and weight, as well as resistance to starvation and increased spontaneous physical activity.Knock-out of a mammalian homolog of Indy (mINDY; SLC13A5) in the mouse protects mice from age-related or high-fat-feeding-related insulin resistance and adiposity, and promotes phenotypes similar to those seen in CR. mINDY-deleted mice show increased hepatic mitochondrial biogenesis, lipid oxidation, and energy expenditure, as well as decreased hepatic de novo lipogenesis, activation of hepatic AMPK, induction of PGC-1alpha, inhibition of ACC-2, and reduction of SREBP-1c. mINDY-deleted mice have reduced body weight, and preserve normal insulin signaling during aging or on high-fat diets, indicating that some of the positive effects of Indy in flies can be extended to mammals. Indy's effect on mammalian energy metabolism suggests that Indy is a potential target for treatment of obesity and type 2 diabetes, and that understanding of its regulation could lead to discovery of new agents for extending healthy life span and provide valuable insight into the genetic mechanisms of normal aging.

English

Glucose transporter type-1 (glut1) and citrate synthase plays crucial role in glucose transport and regulation of tricarboxylic acid cycle (TCA) cycle in mammalian energy metabolism. The present study was aimed to clone and characterize glut1 and citrate synthase cDNA in water buffalo (Bubalus bubalis). Total of 90 cumulus oocyte complexes (COCs) were used for mRNA isolation and reverse transcribed to cDNA, which was further used in polymerase chain reaction (PCR) amplification of glut1 and citrate synthase. PCR products of glut1 and citrate synthase were cloned by T/A cloning using pGEM-T easy vector and further sequenced. Gene sequence analysis of glut1 and citrate synthase revealed that they have open reading frame of 1479 (encoding 492 aa) and 1401 bp (encoding 466 aa), respectively. Further phylogenic analysis of gene and deduced amino acid sequences suggests that bubaline glut1 shares ∼ 89 to 98% and ∼ 97 to 99% similarity at nucleotide and amino acid level respectively whereas citrate synthase shared ∼ 89 to 99% at nucleotide and ∼ 96 to 99% at amino acid level respectively with other domestic species and human. Predicted protein structures of buffalo glut1 protein accentuate the presence of crucial amino acids involved in glucose transport moreover the essential catalytic residues are highly conserved in buffalo citrate synthase. Key words: Buffalo, cloning, characterization, Glut1, citrate synthase.

MTORC1 Controls Mitochondrial Activity and Biogenesis through 4E-BP-Dependent Translational Regulation

mRNA translation is thought to be the most energy-consuming process in the cell. Translation and energy metabolism are dysregulated in a variety of diseases including cancer, diabetes, and heart disease. However, the mechanisms that coordinate translation and energy metabolism in mammals remain largely unknown. The mechanistic/mammalian target of rapamycin complex 1 (mTORC1) stimulates mRNA translation and other anabolic processes. We demonstrate that mTORC1 controls mitochondrial activity and biogenesis by selectively promoting translation of nucleus-encoded mitochondria-related mRNAs via inhibition of the eukaryotic translation initiation factor 4E (eIF4E)-binding proteins (4E-BPs). Stimulating the translation of nucleus-encoded mitochondria-related mRNAs engenders an increase in ATP production capacity, a required energy source for translation. These findings establish a feed-forward loop that links mRNA translation to oxidative phosphorylation, thereby providing a key mechanism linking aberrant mTOR signaling to conditions of abnormal cellular energy metabolism such as neoplasia and insulin resistance.

L-Carnitine Exposure and Mitochondrial Function in Human Neuronal Cells

L-Carnitine is a naturally occurring substance required in mammalian energy metabolism that functions by facilitating long-chain fatty acid entry into cellular mitochondria, thereby delivering substrate for oxidation and subsequent energy production. It has been purposed that L-carnitine may improve and preserve cognitive performance, and may lead to better cognitive aging through the life span, and several controlled human clinical trials with L-carnitine support the hypothesis that this substance has the ability to improve cognitive function. We further hypothesized that, since L-carnitine is an important co-factor of mammalian mitochondrial energy metabolism, acute administration of L-carnitine to human tissue culture cells should result in detectable increases in mitochondrial function. Cultures of SH-SY-5Y human neuroblastoma and 1321N1 human astrocytoma cells grown in 96-well cell culture plates were acutely administered L-carnitine hydrochloride, and then, mitochondrial function was assayed using the colorimetric 2,3-bis[2-methoxy-4-nitro-5-sulfophenyl]-2H-tetrazolium-5-carboxyanilide inner salt cell assay kit in a VERSAmax tunable microplate reader. Significant increases in mitochondrial function were observed when human neuroblastoma or human astrocytoma cells were exposed to 100 nM (20 μg L-carnitine hydrochloride/L) to 100 μM (20 mg L-carnitine hydrochloride/L) concentrations of L-carnitine hydrochloride in comparison to unexposed cells, whereas no significant positive effects were observed at lower or higher concentrations of L-carnitine hydrochloride. The results of the present study provide insights for how L-carnitine therapy may significantly improve human neuronal function, but we recommend that future studies further explore different derivatives of L-carnitine compounds in different in vitro cell-based systems using different markers of mitochondrial function.

The role of short-chain fatty acids in the interplay between diet, gut microbiota, and host energy metabolism

Short-chain fatty acids (SCFAs), the end products of fermentation of dietary fibers by the anaerobic intestinal microbiota, have been shown to exert multiple beneficial effects on mammalian energy metabolism. The mechanisms underlying these effects are the subject of intensive research and encompass the complex interplay between diet, gut microbiota, and host energy metabolism. This review summarizes the role of SCFAs in host energy metabolism, starting from the production by the gut microbiota to the uptake by the host and ending with the effects on host metabolism. There are interesting leads on the underlying molecular mechanisms, but there are also many apparently contradictory results. A coherent understanding of the multilevel network in which SCFAs exert their effects is hampered by the lack of quantitative data on actual fluxes of SCFAs and metabolic processes regulated by SCFAs. In this review we address questions that, when answered, will bring us a great step forward in elucidating the role of SCFAs in mammalian energy metabolism.

Nitric oxide and energy metabolism in mammals

Nitric oxide (NO) is a signaling molecule synthesized from L-arginine by NO synthase in animals. Increasing evidence shows that NO regulates the mammalian metabolism of energy substrates and that these effects of NO critically depend on its concentrations at the reaction site and the period of exposure. High concentrations of NO (in the micromolar range) irreversibly inhibit complexes I, II, III, IV, and V in the mitochondrial respiratory chain, whereas physiological levels of NO (in the nanomolar range) reversibly reduce cytochomrome oxidase. Thus, NO reduces oxygen consumption by isolated mitochondria to various extents. In intact cells, through cGMP and AMP-activated protein kinase signaling, physiological levels of NO acutely stimulate uptake and oxidation of glucose and fatty acids by skeletal muscle, heart, liver, and adipose tissue, while inhibiting the synthesis of glucose, glycogen and fat in the insulin-sensitive tissues, and enhancing lipolysis in white adipocytes. Chronic effects of physiological levels of NO in vivo include stimulation of angiogenesis, blood flow, mitochondrial biogenesis, and brown adipocyte development. Modulation of NO-mediated pathways through dietary supplementation with L-arginine or its precursor L-citrulline may provide an effective, practical strategy to prevent and treat metabolic syndrome, including obesity, diabetes, and dyslipidemia in mammals, including humans.

N-Carbamoylputrescine, a citrulline-derived polyamine, is not a significant citrulline metabolite in rats

Citrulline, a key amino acid of the urea cycle, has been shown to play a regulatory role in protein and energy metabolism in mammals. We questioned whether N-carbamoyl-putrescine (NCP), the decarboxylated derivative of citrulline, could play a role in the biological properties of this amino acid. To evidence the presence of NCP in mammalian tissues, we developed a sensitive reverse-phase high-performance liquid chromatography (HPLC) with fluorimetric detection method with precolumn dansyl derivatization and solid-phase extraction for the determination of NCP together with polyamines in biological samples. Dansyl NCP was identified with a 5.85-min retention time. Linearity was obtained in a concentration range of 0.125 to 12.5μM. Intraday and day-to-day relative coefficients of variation ranged from 8.9% to 12.3% and from 14% to 14.3%, respectively. Recovery rates in serum ranged from 75% to 83%. Thereafter, we used this method to search for the presence of NCP in serum, muscle, liver, jejunum, and ileum in rats after both short-term intraperitoneal injection and long-term oral citrulline supplementation. We failed to detect NCP in these animals. These data suggest that NCP is not a significant citrulline metabolite in rats.

Ghrelin potentiates TSH-induced expression of the thyroid tissue-specific genes thyroglobulin, thyroperoxidase and sodium-iodine symporter, in rat PC-Cl3 Cells

Ghrelin is a 28-amino-acid peptide that stimulates pituitary growth-hormone secretion and modulates food-intake and energy metabolism in mammals. It is mainly secreted by the stomach, but it is also expressed in many other tissues such as cartilage or the thyroid gland. In the present study we have analyzed by RT-PCR and using immunohistochemistry and immunofluorescence the expression and tissue distribution of ghrelin and its functional receptor (GHS-R type 1α) in thyroid cell-lines and in normal and pathological rat thyroid tissue. Additionally, by measuring the incorporation of BrdU, we have investigated if, as previously noted for FRTL-5 cells, ghrelin enhances the proliferation rate in the PC-Cl3 rat-thyrocyte cell-line. Finally, we have determined the stimulatory effect of ghrelin on TSH-induced expression of the tissue-specific key genes involved in the synthesis of thyroid hormone: thyroglobulin, thyroperoxidase and sodium–iodine symporter. Our data provide direct evidence that C-cell secreted ghrelin may be involved in the paracrine regulation of the thyroid follicular cell function.

Indy knockdown in mice mimics elements of dietary restriction

A new mouse knock-out of the Indy gene sheds light on the mechanisms by which a reduction in Indy may lead to a healthier life. Mutations in the Indy gene (for I'm not dead yet) were first described in flies where they dramatically extend life span without a loss in fertility, physical activity, flight velocity or metabolic rate [1, 2]. The Indy gene codes for a high-affinity dicarboxylate/citrate plasma membrane transporter found most abundantly at the plasma membrane of adult fat body, oenocytes and midgut cells, the primary sites of intermediary metabolism in the fly [3]. In support of the hypothesis that the Indy mutation might be altering metabolism in a manner similar to dietary restriction (DR), it has been shown that DR down-regulates Indy in normal flies and that Indy long-lived flies share several phenotypes with long-lived DR flies, including decreased insulin-like signaling, lipid storage, weight gain, and resistance to starvation, and an increase in spontaneous physical activity [4]. In a new report in Cell Metabolism [5], knock-out of a mammalian homolog of Indy (mINDY; SLC13A5) shows dramatic effects in protecting mice from the adiposity and insulin resistance that develop with high-fat feeding or aging, and leads to phenotypes similar to those seen in DR. mINDY-deleted mice exhibit increased hepatic mitochondrial biogenesis, lipid oxidation, and energy expenditure, with a concomitant decrease in hepatic de novo lipogenesis and a reduction in ATP/ADP ratio, leading to activation of hepatic AMPK, induction of PGC-1alpha, inhibition of ACC-2, and reduction of SREBP-1c. These mice have reduced body weight, and preserve normal insulin signaling in the face of a high-fat diet or aging, demonstrating that some of the positive effects of Indy in flies can be extended to mammals. The profound beneficial effects on mammalian energy metabolism suggest that Indy is a potential target for treatment of obesity and type 2 diabetes. By focusing on the potential role of mammalian INDY in prolonging healthy life span, we stand to obtain new agents for extending healthy life span and to gain valuable insight into the genetic underpinnings of normal aging.