Peroxisome proliferator-activated receptor mediates cross-talk with thyroid hormone receptor by competition for retinoid X receptor. Possible role of a leucine zipper-like heptad repeat.

The peroxisome proliferator-activated receptors (PPAR) and thyroid hormone receptors (TR) are members of the nuclear receptor superfamily, which regulate lipid metabolism and tissue differentiation. In order to bind to DNA and activate transcription, PPAR requires the formation of heterodimers with the retinoid X receptor (RXR). In addition to activating transcription through its own response elements, PPAR is able to selectively down-regulate the transcriptional activity of TR, but not vitamin D receptor. The molecular basis of this functional interaction has not been fully elucidated. By means of site-directed mutagenesis of hPPAR alpha we mapped its inhibitory action on TR to a leucine zipper-like motif in the ligand binding domain of PPAR, which is highly conserved among all subtypes of this receptor and mediates heterodimerization with RXR. Replacement of a single leucine by arginine at position 433 of hPPAR alpha (L433R) abolished heterodimerization of PPAR with RXR and consequently its trans-activating capacity. However, a similar mutation of a leucine residue to arginine at position 422 showed no alteration of heterodimerization, DNA binding, or transcriptional activation. The dimerization deficient mutant L433R was no longer able to inhibit TR action, demonstrating that the selective inhibitory effect of PPAR results from the competition for RXR as well as possibly for other TR-auxiliary proteins. In contrast, abolition of DNA binding by a mutation in the P-box of PPAR (C122S) did not eliminate the inhibition of TR trans-activation, indicating that competition for DNA binding is not involved. Additionally, no evidence for the formation of PPAR:TR heterodimers was found in co-immunoprecipitation experiments. In summary, we have demonstrated that PPAR selectively inhibits the transcriptional activity of TRs by competition for RXR and possibly non-RXR TR-auxiliary proteins. In contrast, this functional interaction is independent of the formation of PPAR:TR heterodimers or competition for DNA binding.

The Gene Ontology in 2010: extensions and refinements.

The Gene Ontology (GO) Consortium (http://www.geneontology.org) (GOC) continues to develop, maintain and use a set of structured, controlled vocabularies for the annotation of genes, gene products and sequences. The GO ontologies are expanding both in content and in structure. Several new relationship types have been introduced and used, along with existing relationships, to create links between and within the GO domains. These improve the representation of biology, facilitate querying, and allow GO developers to systematically check for and correct inconsistencies within the GO. Gene product annotation using GO continues to increase both in the number of total annotations and in species coverage. GO tools, such as OBO-Edit, an ontology-editing tool, and AmiGO, the GOC ontology browser, have seen major improvements in functionality, speed and ease of use.

Novel targets of the CbrAB/Crc carbon catabolite control system revealed by transcript abundance in Pseudomonas aeruginosa.

The opportunistic human pathogen Pseudomonas aeruginosa is able to utilize a wide range of carbon and nitrogen compounds, allowing it to grow in vastly different environments. The uptake and catabolism of growth substrates are organized hierarchically by a mechanism termed catabolite repression control (Crc) whereby the Crc protein establishes translational repression of target mRNAs at CA (catabolite activity) motifs present in target mRNAs near ribosome binding sites. Poor carbon sources lead to activation of the CbrAB two-component system, which induces transcription of the small RNA (sRNA) CrcZ. This sRNA relieves Crc-mediated repression of target mRNAs. In this study, we have identified novel targets of the CbrAB/Crc system in P. aeruginosa using transcriptome analysis in combination with a search for CA motifs. We characterized four target genes involved in the uptake and utilization of less preferred carbon sources: estA (secreted esterase), acsA (acetyl-CoA synthetase), bkdR (regulator of branched-chain amino acid catabolism) and aroP2 (aromatic amino acid uptake protein). Evidence for regulation by CbrAB, CrcZ and Crc was obtained in vivo using appropriate reporter fusions, in which mutation of the CA motif resulted in loss of catabolite repression. CbrB and CrcZ were important for growth of P. aeruginosa in cystic fibrosis (CF) sputum medium, suggesting that the CbrAB/Crc system may act as an important regulator during chronic infection of the CF lung.

Structural motifs of biomolecules.

Biomolecular structures are assemblies of emergent anisotropic building modules such as uniaxial helices or biaxial strands. We provide an approach to understanding a marginally compact phase of matter that is occupied by proteins and DNA. This phase, which is in some respects analogous to the liquid crystal phase for chain molecules, stabilizes a range of shapes that can be obtained by sequence-independent interactions occurring intra- and intermolecularly between polymeric molecules. We present a singularity-free self-interaction for a tube in the continuum limit and show that this results in the tube being positioned in the marginally compact phase. Our work provides a unified framework for understanding the building blocks of biomolecules.

Eczéma chronique des mains: traitements conventionnels et nouveautés [Chronic hand eczema, conventional and new treatments].

Chronic hand eczema is a frequent cause of consultation. In Europe and Switzerland, it's one of the main reasons for patients to interrupt their profession. The etiology is pluri-factorial. Atopic patients are more likely predisposed. Pruritus, associated to pain and bleeding, is intense. Psychosocial consequences are huge, making this illness to an important public health problem. Topical treatment and UV-light are the main therapeutical strategy but the results are often disappointing. Recently, alitretinoine (9-cis retinoic acid) became the treatment of second choice with good response, allowing patients to preserve a good quality of life and their job.

Peroxisome proliferator-activated receptors: a nuclear receptor signaling pathway in lipid physiology.

Peroxisome proliferator-activated receptors (PPARs) are lipid-activated transcription factors that belong to the steroid/thyroid/retinoic acid receptor superfamily. All their characterized target genes encode proteins that participate in lipid homeostasis. The recent finding that antidiabetic thiazolidinediones and adipogenic prostanoids are ligands of one of the PPARs reveals a novel signaling pathway that directly links these compounds to processes involved in glucose homeostasis and lipid metabolism including adipocyte differentiation. A detailed understanding of this pathway could designate PPARs as targets for the development of novel efficient treatments for several metabolic disorders.

Fluorodeoxyuridine mediated cell cycle synchronization in S-phase increases the Auger radiation cell killing with 125I-iododeoxyuridine.

Aim: 125I-iododeoxyuridine is a potential Auger radiation therapy agent. Its incorporation in DNA of proliferating cells is enhanced by fluorodeoxyuridine. Here, we evaluated therapeutic activities of 125I-iododeoxyuridine in an optimized fluorodeoxyuridine pre-treatment inducing S-phase synchronization. Methods: After S-phase synchronization by fluorodeoxyuridine, cells were treated with 125I-iododeoxyuridine. Apoptosis analysis and S-phase synchronization were studied by flow cytometry. Cell survival was determined by colony-forming assay. Based on measured growth parameters, the number of decays per cell that induced killing was extrapolated. Results: Treatment experiments showed that 72 to 91% of synchronized cells were killed after 0.8 and 8 kBq/ml 125I-iododeoxyuridine incubation, respectively. In controls, only 8 to 38% of cells were killed by corresponding 125I-iododeoxyuridine activities alone and even increasing the activity to 80 kBq/ml gave only 42 % killing. Duplicated treatment cycles or repeated fluorodeoxyuridine pre-treatment allowed enhancing cell killing to >95 % at 8 kBq/ml 125I-iododeoxyuridine. About 50 and 160 decays per S-phase cells in controls and S-phase synchronization, respectively, were responsible for the observed cell killing at 0.8 kBq/ml radio-iododeoxyuridine. Conclusion: These data show the successful application of fluorodeoxyuridine that provided increased 125I-iododeoxyuridine Auger radiation cell killing efficacy through S-phase synchronization and high DNA incorporation of radio-iododeoxyuridine.

Study of inner ear and lateral line hair cell regeneration

Death of sensory hair cells in the inner ear results in two global health problems that millions of people around the world suffer: hearing loss and balance disorders. Hair cells convert sound vibrations and head movements into electrical signals that are conveyed to the brain, and as a result of aging, exposure to noise, modern drugs or genetic predisposition, hair cells die. In mammals, the great majority of hair cells are produced during embryogenesis, and hair cells that are lost after birth are not replaceable. However, in the last decades, researches have shown some model organisms that retain the ability to regenerate hair cells damaged after embryogenesis, such as Zebrafish and chicken, providing clues as to the cellular and molecular mechanisms that may block hair cell regeneration in mammals. This discovery initiated a search for methods to stimulate regeneration or replacement of hair cells in mammals, a search that, if fruitful, will revolutionize the treatment of hearing loss and balance disorders. One aim of my project is to study the role of retinoic acid in adult Zebrafish and in mice, which is a metabolite of vitamin A known as an essential molecule to activate hair cell regeneration after cells damaged in Zebrafish embryo. We want to study important genes involved in retinoic acid pathway, such as Aldh1a3 and RARs genes, to check what their role is in the inner ear of adult Zebrafish and compare result obtained in the inner ear of mice. On the other hand, Zebrafish lateral line contains neuromast, which are formed by the same structure than the inner ear: hair cells surrounded by supporting cells and neurons. The lateral line is a structure below the skin's surface that makes easier to damage hair cells to study their regeneration. For that reason, another aim of my project is to study how Sox2 and Atoh1, essential genes during the inner ear development, change their expression during hair cell regeneration in the lateral line. In my project, the most important concepts related to Zebrafish world are explained in order to understand why we have studied this animal and these essential genes. Then, techniques that we used are explained, with their protocol attached in the annexes. Finally, results of my project are shown, but many of them were not expected and they would be needed to follow studying.

Glut2-dependent glucose-sensing controls thermoregulation by enhancing the leptin sensitivity of NPY and POMC neurons.

The physiological contribution of glucose in thermoregulation is not completely established nor whether this control may involve a regulation of the melanocortin pathway. Here, we assessed thermoregulation and leptin sensitivity of hypothalamic arcuate neurons in mice with inactivation of glucose transporter type 2 (Glut2)-dependent glucose sensing. Mice with inactivation of Glut2-dependent glucose sensors are cold intolerant and show increased susceptibility to food deprivation-induced torpor and abnormal hypothermic response to intracerebroventricular administration of 2-deoxy-d-glucose compared to control mice. This is associated with a defect in regulated expression of brown adipose tissue uncoupling protein I and iodothyronine deiodinase II and with a decreased leptin sensitivity of neuropeptide Y (NPY) and proopiomelanocortin (POMC) neurons, as observed during the unfed-to-refed transition or following i.p. leptin injection. Sites of central Glut-2 expression were identified by a genetic tagging approach and revealed that glucose-sensitive neurons were present in the lateral hypothalamus, the dorsal vagal complex, and the basal medulla but not in the arcuate nucleus. NPY and POMC neurons were, however, connected to nerve terminals from Glut2-expressing neurons. Thus, our data suggest that glucose controls thermoregulation and the leptin sensitivity of NPY and POMC neurons through activation of Glut2-dependent glucose-sensing neurons located outside of the arcuate nucleus.

Guiding ligands to nuclear receptors.

Retinoic acid-the active metabolite of vitamin A-influences biological processes by activating the retinoic acid receptor (RAR). In this issue, Schug et al. (2007) demonstrate that retinoic acid also activates the peroxisome proliferator-activated receptor beta/delta (PPARbeta/delta). Remarkably, retinoic acid signaling through RAR or PPARbeta/delta-which depends on cytoplasmic retinoic acid transporters-commits the cell to opposite fates, apoptosis or survival, respectively.

Establishment of an In Vitro Photoallergy Test Using NCTC2544 Cells and IL-18 Production

Differentiation between photoallergenic and phototoxic reactions induced by low molecular weight compounds represents a current problem. The use of eratinocytes as a potential tool for the detection of photoallergens as opposed to photoirritants is considered an interesting strategy for developing in vitro methods. We have previously demonstrated the possibility to use the human keratinocyte cell line NCTC2455 and the production of interleukin-18 (IL-18) to screen low molecular weight sensitizers. The purpose of this work was to explore the possibility to use the NCTC2544 assay to identify photoallergens and discriminate from phototoxic chemicals. First, we identified suitable condition of UV-irradiation (3.5 J/cm2) by investigating the effect of UVAirradiation on intracellular IL-18 on untreated or chloropromazine (a representative phototoxic compound)- treated NCTC2544 cells. Then, the effect of UVA-irradiation over NCTC2544 cells treated with increasing concentrations of 15 compounds including photoallergens (benzophenone, 4-ter-butyl-4-methoxydibenzoylmethane, 2-ethylexyl-p-methoxycinnamate, ketoprofen, 6-methylcumarin); photoirritant and photoallergen (4-aminobenzoic acid, chlorpromazine, promethazine); photoirritants (acridine, ibuprofen, 8-methoxypsoralen, retinoic acid); and negative compounds (lactic acid, SDS and p-phenilendiamine) was investigated. Twenty-four hours after exposure, cytotoxicity was evaluated by the MTT assay or LDH leakage, while ELISA was used to measure the production of IL-18. At the maximal concentration assayed with non-cytotoxic effects (CV80 under irradiated condition), all tested photoallergens induced a significant and a dose-dependent increase of intracellular IL-18 following UVA irratiation, whereas photoirritants failed. We suggest that this system may be useful for the in vitro evaluation of the photoallergic potential of chemicals.

Circulating FABP4 Is a prognostic biomarker in patients with acute coronary syndrome but not in asymptomatic individuals.

OBJECTIVE: Blood-borne biomarkers reflecting atherosclerotic plaque burden have great potential to improve clinical management of atherosclerotic coronary artery disease and acute coronary syndrome (ACS). APPROACH AND RESULTS: Using data integration from gene expression profiling of coronary thrombi versus peripheral blood mononuclear cells and proteomic analysis of atherosclerotic plaque-derived secretomes versus healthy tissue secretomes, we identified fatty acid-binding protein 4 (FABP4) as a biomarker candidate for coronary artery disease. Its diagnostic and prognostic performance was validated in 3 different clinical settings: (1) in a cross-sectional cohort of patients with stable coronary artery disease, ACS, and healthy individuals (n=820), (2) in a nested case-control cohort of patients with ACS with 30-day follow-up (n=200), and (3) in a population-based nested case-control cohort of asymptomatic individuals with 5-year follow-up (n=414). Circulating FABP4 was marginally higher in patients with ST-segment-elevation myocardial infarction (24.9 ng/mL) compared with controls (23.4 ng/mL; P=0.01). However, elevated FABP4 was associated with adverse secondary cerebrovascular or cardiovascular events during 30-day follow-up after index ACS, independent of age, sex, renal function, and body mass index (odds ratio, 1.7; 95% confidence interval, 1.1-2.5; P=0.02). Circulating FABP4 predicted adverse events with similar prognostic performance as the GRACE in-hospital risk score or N-terminal pro-brain natriuretic peptide. Finally, no significant difference between baseline FABP4 was found in asymptomatic individuals with or without coronary events during 5-year follow-up. CONCLUSIONS: Circulating FABP4 may prove useful as a prognostic biomarker in risk stratification of patients with ACS.

Broken colinearity of the amphioxus Hox cluster.

Background In most eumetazoans studied so far, Hox genes determine the identity of structures along the main body axis. They are usually linked in genomic clusters and, in the case of the vertebrate embryo, are expressed with spatial and temporal colinearity. Outside vertebrates, temporal colinearity has been reported in the cephalochordate amphioxus (the least derived living relative of the chordate ancestor) but only for anterior and central genes, namely Hox1 to Hox4 and Hox6. However, most of the Hox gene expression patterns in amphioxus have not been reported. To gain global insights into the evolution of Hox clusters in chordates, we investigated a more extended expression profile of amphioxus Hox genes. Results Here we report an extended expression profile of the European amphioxus Branchiostoma lanceolatum Hox genes and describe that all Hox genes, except Hox13, are expressed during development. Interestingly, we report the breaking of both spatial and temporal colinearity for at least Hox6 and Hox14, which thus have escaped from the classical Hox code concept. We show a previously unidentified Hox6 expression pattern and a faint expression for posterior Hox genes in structures such as the posterior mesoderm, notochord, and hindgut. Unexpectedly, we found that amphioxus Hox14 had the most divergent expression pattern. This gene is expressed in the anterior cerebral vesicle and pharyngeal endoderm. Amphioxus Hox14 expression represents the first report of Hox gene expression in the most anterior part of the central nervous system. Nevertheless, despite these divergent expression patterns, amphioxus Hox6 and Hox14 seem to be still regulated by retinoic acid. Conclusions Escape from colinearity by Hox genes is not unusual in either vertebrates or amphioxus and we suggest that those genes escaping from it are probably associated with the patterning of lineage-specific morphological traits, requiring the loss of those developmental constraints that kept them colinear.

Sensitive Troponin Assays: Diagnostic and Prognostic Use in Cardiology

Cardiac troponins (cTns) are the recommended biochemical markers in the diagnosis of myocardial infarction (MI). They are very sensitive and tissue-specific but are limited by their delayed appearance in the circulation. Biochemical markers with more rapid release kinetics, e.g. myoglobin and especially heart-type fatty acid-binding protein (H-FABP), have been used to enhance the early identification of MI. The implementation of cTns into clinical practice has shown that cardiomyocyte injury occurs in many other clinical conditions than MI. The aim of this study was to evaluate the impact of modern and highly sensitive cTnI assays on the early diagnosis of MI. In a patient cohort with suspected MI, such a sensitive cTnI assay enhanced the early diagnostic accuracy when compared to a less sensitive cTnI assay and to myoglobin. When compared to H-FABP during the early hours after symptom onset, the sensitive cTnI assay showed at least similar and, after 6 hours, superior diagnostic accuracy. A positive cTnI test result had superior prognostic value when compared to H-FABP, even among early presenters. The prognostic value of cTn in acute heart failure (AHF) was evaluated in 364 patients who participated in the FINN-AKVA study. The patients presented with AHF but no acute coronary syndrome (ACS). Up to half of the patients had elevated cTn levels which were associated with higher 6-month mortality. The magnitude of cTn elevation was directly proportional to mortality. Finally, the clinical spectrum of cTnI elevations was evaluated in 991 cTnI positive emergency department (ED) patients. 83% of the patients had MI and 17% had cTnI elevation due to other clinical conditions. The latter patient group was characterized by lower absolute cTnI levels and – importantly – higher in-hospital mortality when compared to the MI patients. In conclusion, the use of a highly sensitive cTnI assay enhances the early diagnostic accuracy and risk stratification in suspected MI patients. Cardiac troponin elevations are highly prevalent also in other acute clinical conditions and indicate an adverse outcome of these patients.