Reduction of atherosclerosis by the peroxisome proliferator-activated receptor alpha agonist fenofibrate in mice

Several clinical and angiographic intervention trials have shown that fibrate treatment leads to a reduction of the coronary events associated to atherosclerosis. Fibrates are ligands for peroxisome proliferator-activated receptor alpha (PPARalpha) that modulate risk factors related to atherosclerosis by acting at both systemic and vascular levels. Here, we investigated the effect of treatment with the PPARalpha agonist fenofibrate (FF) on the development of atherosclerotic lesions in apolipoprotein (apo) E-deficient mice and human apoA-I transgenic apoE-deficient (hapoA-I Tg x apoE-deficient) mice fed a Western diet. In apoE-deficient mice, plasma lipid levels were increased by FF treatment with no alteration in the cholesterol distribution profile. FF treatment did not reduce atherosclerotic lesion surface area in the aortic sinus of 5-month-old apoE-deficient mice. By contrast, FF treatment decreased total cholesterol and esterified cholesterol contents in descending aortas of these mice, an effect that was more pronounced in older mice exhibiting more advanced lesions. Furthermore, FF treatment reduced MCP-1 mRNA levels in the descending aortas of apoE-deficient mice, whereas ABCA-1 expression levels were maintained despite a significant reduction of aortic cholesterol content. In apoE-deficient mice expressing a human apoA-I transgene, FF increased human apoA-I plasma and hepatic mRNA levels without affecting plasma lipid levels. This increase in human apoA-I expression was accompanied by a significant reduction in the lesion surface area in the aortic sinus. These data indicate that the PPARalpha agonist fenofibrate reduces atherosclerosis in these animal models of atherosclerosis.     

Discovery and Verification of Osteopontin and Beta-2-microglobulin as Promising Markers for Staging Human African Trypanosomiasis

Human African trypanosomiasis, or sleeping sickness, is a parasitic disease endemic in sub-Saharan Africa, transmitted to humans through the bite of a tsetse fly. The first or hemolymphatic stage of the disease is associated with presence of parasites in the bloodstream, lymphatic system, and body tissues. If patients are left untreated, parasites cross the blood-brain barrier and invade the cerebrospinal fluid and the brain parenchyma, giving rise to the second or meningoencephalitic stage. Stage determination is a crucial step in guiding the choice of treatment, as drugs used for S2 are potentially dangerous. Current staging methods, based on counting white blood cells and demonstrating trypanosomes in cerebrospinal fluid, lack specificity and/or sensitivity. In the present study, we used several proteomic strategies to discover new markers with potential for staging human African trypanosomiasis. Cerebrospinal fluid (CSF) samples were collected from patients infected with Trypanosoma brucei gambiense in the Democratic Republic of Congo. The stage was determined following the guidelines of the national control program. The proteome of the samples was analyzed by two-dimensional gel electrophoresis (n = 9), and by sixplex tandem mass tag (TMT) isobaric labeling (n = 6) quantitative mass spectrometry. Overall, 73 proteins were overexpressed in patients presenting the second stage of the disease. Two of these, osteopontin and β-2-microglobulin, were confirmed to be potential markers for staging human African trypanosomiasis (HAT) by Western blot and ELISA. The two proteins significantly discriminated between S1 and S2 patients with high sensitivity (68% and 78%, respectively) for 100% specificity, and a combination of both improved the sensitivity to 91%. The levels of osteopontin and β-2-microglobulin in CSF of S2 patients (μg/ml range), as well as the fold increased concentration in S2 compared with S1 (3.8 and 5.5 respectively) make the two markers good candidates for the development of a test for staging HAT patients.Human African trypanosomiasis (HAT), or sleeping sickness, is caused by an extracellular protozoan parasite of the genus Trypanosoma, which is transmitted through the bite of a tsetse fly (genus Glossina). Two morphologically identical subspecies of the parasite, are responsible for the two geographically and clinically different forms of HAT: a chronic form, widespread in West and Central Africa, caused by T. b. gambiense, and an acute form, endemic in eastern Africa, caused by T. b. rhodesiense (1). In both forms of the disease, parasites are initially localized in the blood stream, lymph, and peripheral tissues; this is the first or hemolymphatic stage (S1). During this stage, patients present generic clinical features that are common to other infectious diseases such as human immunodeficiency virus (HIV), malaria, and tuberculosis (TB), which can coexist with HAT, thus making its early diagnosis difficult (2). If treatment is not carried out, the disease progresses to the second or meningoencephalitic stage (S2) after trypanosomes cross the blood-brain barrier (BBB) and invade the central nervous system (CNS). This phase is characterized by a broad range of neurological signs that are indicative of CNS involvement (1). Diagnosis of HAT is based on parasitological demonstration of parasites in blood or lymph-node aspirate (3). All positive or suspect patients have to undergo a lumbar puncture and cerebrospinal fluid (CSF)1 examination, to determine whether they have second stage disease (4). According to the World Health Organization (WHO) guidelines, the meningoencephalitic stage is defined by the presence of parasites in CSF and/or a white blood cell (WBC) count of more than 5 cells per μl (5). Other parameters, such as intrathecal IgM production could also provide additional information to determine whether the CNS is involved (6, 7).Treatment of HAT patients varies depending on the infecting parasite and the stage of disease (5, 8). S2 drugs in current use, including melarsoprol, eflornithine, and a combination of nifurtimox and eflornithine have several limitations, such as a high rate of toxicity (melarsoprol causes death to 5% of treated patients) (9), complex logistics, and mode of administration (6, 10). Consequently, staging is a vital step in the diagnosis and treatment of HAT. However, the poor specificity or sensitivity of WBC counting and of parasitological techniques for demonstration of parasites in CSF, highlight the need for discovery of better tools for staging the disease.Several attempts have been made during the last decade to identify potential biomarkers able to discriminate between the two stages of sleeping sickness. Most of the efforts focused on cytokines and chemokines, because the patient''s immune system plays a crucial role in the brain pathology (11–14).Proteomic approaches are increasingly being applied in biomedical research and clinical medicine to investigate body fluids as a source of biomarkers (15), including the diagnosis of neurological disorders such as Alzheimer''s disease (16), Parkinson''s disease (17), and multiple sclerosis (18, 19). The protein composition of CSF is strictly regulated and can reflect the physiological or pathological state of the CNS (15). Thus in the present study, we addressed the challenge of staging HAT by analyzing CSF from T. b. gambiense patients using two complementary proteomic strategies: a classical approach based on two-dimensional gel electrophoresis (2-DE), and quantitative mass spectrometry (MS) using isobaric tandem mass tag (TMT) technology (sixplex TMT® MS/MS) (20).     

Determination of the binding mode and interacting amino-acids for dibasic H3 receptor antagonists

Due to its involvement in major CNS functions, the histamine H3 receptor (H3R) is the subject of intensive medicinal chemistry investigation, supported by the range of modern drug discovery tools, such as receptor modeling and ligand docking. Although the receptor models described to date share a majority of common traits, they display discrete alternatives in amino-acid conformation, rendering ligand binding modes quite different. Such variations impede structure-based drug design in the H3R field. In the present study, we used a combination of medicinal chemistry, receptor-guided and ligand-based methods to elucidate the binding mode of antagonists. The approaches converged towards a ligand orientation perpendicular to the membrane plane, bridging Glu206 of the transmembrane helix 5 to acidic amino acids of the extracellular loops. This consensus will help future structure-based drug design for H3R ligands.     

Epistasis in iron metabolism: complex interactions between Cp, Mon1a, and Slc40a1 loci and tissue iron in mice

Disorders of iron metabolism are among the most common acquired and constitutive diseases. Hemochromatosis has a solid genetic basis and in Northern European populations it is usually associated with homozygosity for the C282Y mutation in the HFE protein. However, the penetrance of this mutation is incomplete and the clinical presentation is highly variable. The rare and common variants identified so far as genetic modifiers of HFE-related hemochromatosis are unable to account for the phenotypic heterogeneity of this disorder. There are wide variations in the basal iron status of common inbred mouse strains, and this diversity may reflect the genetic background of the phenotypic diversity under pathological conditions. We therefore examined the genetic basis of iron homeostasis using quantitative trait loci mapping applied to the HcB-15 recombinant congenic strains for tissue and serum iron indices. Two highly significant QTL containing either the N374S Mon1a mutation or the Ferroportin locus were found to be major determinants in spleen and liver iron loading. Interestingly, when considering possible epistatic interactions, the effects of Mon1a on macrophage iron export are conditioned by the genotype at the Slc40a1 locus. Only mice that are C57BL/10ScSnA homozygous at both loci display a lower spleen iron burden. Furthermore, the liver-iron lowering effect of the N374S Mon1a mutation is observed only in mice that display a nonsense mutation in the Ceruloplasmin (Cp) gene. This study highlights the existence of genetic interactions between Cp, Mon1a, and the Slc40a1 locus in iron metabolism, suggesting that epistasis may be a crucial determinant of the variable biological and clinical presentations in iron disorders.     

Microbiological Conversion of Lapachol by Various Microorganisms

Oxydative cyclization of lapachol was attempted by feeding this compound into the culture broth of Beauveria sulfurescens and two ionophore-producing strains of Streptomyces albus and Streptomyces griseus. As a result, three compounds were obtained, lomatiol, lomatic acid and lomatiol acetate, only with B. sulfurescens and S. albus. The structures of these products were determinated by NMR and mass spectrometry. The stereochemistry is given by a comparison with butene analogs.     

FXR-deficiency confers increased susceptibility to torpor

The role of the nuclear receptor FXR in adaptive thermogenesis was investigated using FXR-deficient mice. Despite elevated serum bile acid concentrations and increased mRNA expression profiles of thermogenic genes in brown adipose tissue, FXR-deficiency did not alter energy expenditure under basal conditions. However, FXR-deficiency accelerated the fasting-induced entry into torpor in a leptin-dependent manner. FXR-deficient mice were also extremely cold-intolerant. These altered responses may be linked to a more rapid decrease in plasma concentrations of metabolic fuels (glucose, triglycerides) thus impairing uncoupling protein 1-driven thermogenesis. These results identify FXR as a modulator of energy homeostasis.     

Conjugated linoleic acids: all the same or to everyone its own function?

Conjugated linoleic acid (CLA) is a generic term referring to a mixture of geometrical and positional isomers of linoleic acid in which up to 16 members have been identified. Many potentially beneficial health effects have been ascribed to these fatty acids when consumed as a mixture, and where generally 2 isomers dominate, e.g. the 9c, 11t-isomer, the so-called rumenic acid, and the 10t, 12c-isomer: anti-carcinogenic, immune modulator, anti-atherosclerotic, and anti-obesity among the most spectacular. The question arises as to whether the pleiotropic biological activity is supported by one or several of the isomers. Recent studies using pure individual isomers have started to elucidate this issue, but many others are required to ascribe a respective role to each CLA isomer (the main ones as well as the minor ones), such as those occurring in some complex mixtures already commercially available, or even in foodstuff. The aim of the present study was to focus on the CLA-isomer specific effects depicted in the literature up to now.     

Carbohydrate cycling in micro-organisms: what can (13)C-NMR tell us?

The extension of (13)C-nuclear magnetic resonance (NMR) techniques to study cellular metabolism over recent years has provided valuable data supporting the occurrence, diversity and extent of carbon cycling in the carbohydrate metabolism of micro-organisms. The occurrence of such cycles, resulting from the simultaneous operation of different and sometimes opposite individual steps, is inherently related to the network organisation of cellular metabolism. These cycles are tentatively classified here as 'reversibility', 'metabolic' and 'substrate' cycles on the basis of their balance in carbon and cofactors. Current hypotheses concerning the physiological relevance of carbohydrate cycles are discussed in light of the (13)C-NMR data. They most likely represent system-level mechanisms for coherent and timely partitioning of carbon resources to fit with the various biosynthetic, energetic or redox needs of cells and/or additional strategies in the adaptive capacity of micro-organisms to face variation in environmental conditions.