Functional annotation of genomic data with metabolic inference.

Metabolomics is an appealing new approach in systems biology aimed at enabling an improved understanding of the dynamic biochemical composition of living systems. Biological systems are remarkably complex. Importantly, metabolites are the end products of cellular regulatory processes, and their concentrations reflect the ultimate response of a biological system to genetic or environmental changes. In this article, we describe the components of lipid metabolomics and then use them to investigate the metabolic basis for increased abdominal adiposity in 2 strains of divergently selected chickens. Lipid metabolomics were chosen due to the availability of well-developed analytical platforms and the pervasive physiological importance of lipids in metabolism. The analysis suggests that metabolic shifts that result in increased abdominal adiposity are not universal and vary with genetic background. Metabolomics can be used to reverse engineer selection programs through superior metabolic descriptions that can then be associated with specific gene networks and transcriptional profiles.

Nutritional and hormonal regulation of expression of the gene for malic enzyme.

We have provided a historical and personal description of the analysis of physiological and molecular mechanisms by which diet and hormones regulate the activity of hepatic malic enzyme. For the most part, our analyses have been reductionist in approach, striving for increasingly simpler systems in which we can ask more direct questions about the molecular nature of the signaling pathways that regulate the activity of malic enzyme. The reductionist approaches that were so successful at analyzing molecular mechanisms in cells in culture may now provide the means to analyze more definitively questions about the physiological mechanisms involved in nutritional regulation of gene expression. In addition to physiological questions, however, there are still many aspects of the molecular mechanisms that have not been elucidated. Despite considerable effort from many laboratories, the molecular mechanisms by which T3 regulates transcription are not clear. Similarly, the molecular details for the mechanisms by which glucagon, insulin, glucocorticoids, and fatty acids regulate gene expression remain to be determined. The role of fatty acids is particularly interesting because it may provide a model for mechanisms by which genes are regulated by metabolic intermediates; this is a form of transcriptional regulation widely used by prokaryotic organisms and extensively analyzed in prokaryotic systems, but poorly understood in higher eukaryotes. At any specific time, there is, of course, only one rate of transcription for each copy of the malic-enzyme gene in a cell. Our long-term objective is to understand how signals from all of the relevant regulatory pathways are integrated to bring about that rate.

Peroxisome proliferator-activated receptors: a family of lipid-activated transcription factors.

Peroxisome proliferator-activated receptors (PPARs) are a family of nuclear transcription factors that belong to the steroid receptor superfamily. This family of PPARs includes PPARalpha, PPARdelta, PPARgamma1, and PPARgamma2. These PPARs are related to the T3 and vitamin D(3) receptors and bind to a hexameric direct repeat as a heterodimeric complex with retinoid receptor Xalpha. PPARs regulate the expression of a wide array of genes that encode proteins involved in lipid metabolism, energy balance, eicosanoid signaling, cell differentiation, and tumorigenesis. A unique feature of these steroid-like receptors is that the physiologic ligands for PPARs appear to be fatty acids from the n-6 and n-3 families of fatty acids and their respective eicosanoid products. This review describes the characteristics, regulation, and gene targets for PPARs and relates their effects on gene expression to physiologic outcomes that affect lipid and glucose metabolism, thermogenesis, atherosclerosis, and cell differentiation.

Coordinate induction of peroxisomal acyl-CoA oxidase and UCP-3 by dietary fish oil: a mechanism for decreased body fat deposition.

Rats fed dietary fats rich in 20- and 22-carbon polyenoic fatty acids deposit less fat and expend more energy at rest than rats fed other types of fats. We hypothesized that this decrease in energetic efficiency was the product of: (a) enhanced peroxisomal fatty acid oxidation and/or (b) the up-regulation of genes encoding proteins that were involved with enhanced heat production, i.e. mitochondrial uncoupling proteins (UCP-2, UCP-3) and peroxisomal fatty acid oxidation proteins. Two groups of male Fisher 344 rats 3-4 week old (n=5 per group) were pair fed for 6 weeks a diet containing 40% of its energy fat derived from either fish oil or corn oil. Epididymal fat pads from rats fed the fish oil diet weighed 25% (P < 0.05) less than those found in rats fed corn oil. The decrease in fat deposition associated with fish oil ingestion was accompanied by a significant increase in the abundance of skeletal muscle UCP-3 mRNA. The level of UCP-2 mRNA skeletal muscle was unaffected by the type of dietary oil, but the abundance of UCP-2 mRNA in the liver and heart were significantly lower (P < 0.05) in rats fed fish oil than in rats fed corn oil. In addition to inducing UCP-3 expression, dietary fish oil induced peroxisomal acyl-CoA oxidase gene expression 2-3 fold in liver, skeletal muscle and heart. These data support the hypothesis that dietary fish oil reduces fat deposition by increasing the expression of mitochondrial uncoupling proteins and increasing fatty acid oxidation by the less efficient peroxisomal pathway.

Metabolomic mapping of atypical antipsychotic effects in schizophrenia.

Schizophrenia is associated with impairments in neurotransmitter systems and changes in neuronal membrane phospholipids. Several atypical antipsychotic drugs induce weight gain and hypertriglyceridemia. To date, there has not been a comprehensive evaluation and mapping of global lipid changes in schizophrenia, and upon treatment with antipsychotics. Such mapping could provide novel insights about disease mechanisms and metabolic side effects of therapies used for its treatment. We used a specialized metabolomics platform 'lipidomics' that quantifies over 300 polar and nonpolar lipid metabolites (across seven lipid classes) to evaluate global lipid changes in schizophrenia and upon treatment with three commonly used atypical antipsychotics. Lipid profiles were derived for 50 patients with schizophrenia before and after treatment for 2-3 weeks with olanzapine (n=20), risperidone (n=14) or aripiprazole (n=16). Patients were recruited in two cohorts (study I, n=27 and study II, n=23) to permit an internal replication analyses. The change from baseline to post-treatment was then compared among the three drugs. Olanzapine and risperidone affected a much broader range of lipid classes than aripiprazole. Approximately 50 lipids tended to be increased with both risperidone and olanzapine and concentrations of triacylglycerols increased and free fatty acids decreased with both drugs but not with aripiprazole. Phosphatidylethanolamine concentrations that were suppressed in patients with schizophrenia were raised by all three drugs. Drug specific differences were also detected. A principal component analysis (PCA) identified baseline lipid alterations, which correlated with acute treatment response. A more definitive long-term randomized study of these drugs correlating global lipid changes with clinical outcomes could yield biomarkers that define drug-response phenotypes.

An improved technique for processing an overdenture on implants.

This article shows an alternate procedure for making an overdenture on implants. This method helps to eliminate a chairside reline for the dentist by directly incorporating the clips into the heat-cured acrylic resin. The procedure is divided into three main sections. The first section deals with the impressions, casts, and records. The second section relates to the making of the bar, and the third section shows the assembling of the clips.

[Prostheses on osteointegrated implants: restoration of a missing tooth (4)]. / Les prothèses sur implants ostéointégrés: restauration d'une dent manquante (4ième partie).

This article is the last of a series of four. This article describes in detail the fabrication of a crown connected to a Brånemark implant.

Regulation of the action of steroid/thyroid hormone receptors by medium-chain fatty acids.

Triiodothyronine (T3) causes a 30-fold increase in transcription of the malic enzyme gene in chick embryo hepatocytes; medium-chain fatty acids (MCFAs) inhibit this increase. T3 action is mediated by T3 receptors (TRs) that bind to T3 response elements (T3REs) in this gene's 5'-flanking DNA. In transiently transfected hepatocytes, fragments of 5'-flanking DNA of the malic enzyme gene or artificial T3REs that conferred T3 stimulation also conferred MCFA inhibition to linked reporter genes. Thus, MCFA inhibition may be mediated through cis-acting T3REs and trans-acting TRs, distinguishing MCFA action from that of other fatty acids which act through unique sequence elements. Using binding assays and overexpression of TR, we showed that MCFAs inhibited the transactivating but not the silencing function of TR and did not alter binding of T3 to TR or of TR to T3RE. The C-terminal ligand-binding domain of TR was sufficient to confer stimulation by T3, but not inhibition by MCFA. Inhibition of transactivation by MCFA was specific: ligand-stimulated transcription from T3 or estrogen response elements was inhibited, but that from glucocorticoid or cyclic AMP response elements was not. We propose that MCFAs or metabolites thereof influence the activity of a factor(s) that interacts with the T3 and estrogen receptors to inhibit ligand-stimulated transcription.

Specific effects of polyunsaturated fatty acids on gene expression.

Dietary polyunsaturated fatty acids modulate the transcription of numerous proteins including several hepatic lipogenic enzymes. Several lines of evidence indicate that polyunsaturated fatty acids do not regulate the transcription of hepatic lipogenic genes via a peroxisomal proliferator-activated receptor, but the mechanism may involve transcription factors related to the carbohydrate response region.

[Dentures on osseointegrated implants: the complete fixed-removable lower denture (2)]. / Les prosthèses sur implants ostéointegrés: la prothèse complète inférieure fixe-amovible. (2ième partie)

This article is the second of a series of four. This article describes in detail the fabrication of a complete fixed-removable prosthesis connected to a dolder bar.

[Dentures on osseointegrated implants: the complete lower fixed denture (1)]. / Les prosthèses sur implants ostéointegrés: la prothèse complète inférieure fixe. (1ère partie).

This article is the first of a series of four. Each one summarizes the clinical and technical prosthetic stages of different types of restorations. This article describes in detail the fabrication of a complete fixed prosthesis using screw type osseointegrated implants.

Nutritional and hormonal regulation of the gene for malic enzyme.

In vivo, refeeding starved chickens stimulates transcription of the avian gene for malic enzyme in liver; in hepatocytes in culture, triiodothyronine (T3) and insulin stimulate transcription of this gene. In vivo, starvation, and in hepatocytes in culture, glucagon, medium-chain fatty acids (MCFA) and long-chain fatty acids (LCFA) inhibit transcription of the malic enzyme gene. We have defined a T3-response unit in the 5'-flanking DNA of the malic enzyme gene; it contains one major T3 response element and several minor ones; maximum responsiveness is dependent on the presence of all of these elements. LCFA probably act by inhibiting binding of T3 to its nuclear receptor. MCFA appear to act by a different mechanism. Inhibitory MCFA have chain lengths of six, seven or eight carbons; a common feature of other inhibitory compounds is that they can be metabolized to MCFA. Eight-carbon fatty acids with a hydroxyl on the 2- or 3-carbon are more potent inhibitors than octanoate, whereas 2-bromo-fatty acids and 2-hydroxy hexanoate are not inhibitory. In transfection experiments with a large variety of constructs derived from the malic enzyme 5'-flanking DNA, the ability of fatty acids to inhibit promoter function localizes to regions of DNA that contain T3REs. Promoter function of artificial T3REs also is inhibited by MCFA. Inhibition of promoter function using malic enzyme DNA is relatively constant in magnitude irrespective of the size of the T3 response. We postulate that MCFA directly regulates one of the functions of the T3 receptor.

A novel 3T3-L1 preadipocyte variant that expresses PPARgamma2 and RXRalpha but does not undergo differentiation.

This report describes a novel adipocyte-like cell line termed 3T3-L1/RB1 that was derived from preadipocyte cell line, 3T3-L1. The 3T3-L1/RB1 cells continued to divide after reaching confluence, formed foci, and constitutively expressed a low level of adipose fatty acid binding protein (A-FABP) mRNA. However, 3T3L-1/RB cells did not undergo terminal differentiation as indicated by the failure of insulin and thiazolidendiones to induce the expression of A-FABP, lipoprotein lipase, and fatty acid synthase. We hypothesized that the 3T3-L1/RB1 variant did not respond to differentiation stimuli because it did not express either peroxisomal proliferator activated receptor gamma2 (PPARgamma2) or its heterodimer partner, retinoid X receptor alpha (RXRalpha). Surprisingly, Western blots revealed that 3T3-L1/ RB1 cells contained both PPARgamma2 and RXRalpha proteins at levels equal to or greater than that of the parent cell line. However, gel retardation assays using the adipose response element from A-FABP and nuclear protein extracts from 3T3-L1/RB1 cells treated with insulin or pioglitazone revealed that nuclear protein extracts from 3T3-L1/RB1 cells had very little ability to bind the PPARgamma2 recognition sequence of the A-FABP gene. These data suggest that the 3T3-L1/RB1 variant contains a mutation that may prevent ligand activation of PPARgamma2, and the subsequent conversion of 3T3-L1/RB1 cells to mature fat cells.