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  • Cross-talk between the RcsCDB and RstAB systems to control STM1485 gene expression in Salmonella Typhimurium during acid-resistance response
    Biochimie (IF 3.188) Pub Date : 2019-02-11
    Mónica F. Torrez Lamberti, Juan V. Farizano, Fabián E. López, Martin G. Martínez Zamora, María M. Pescaretti, Mónica A. Delgado

    Bacterial survive and respond to adverse changes in the environment by regulating gene transcription through two-component regulatory systems. In Salmonella Typhimurium the STM1485 gene expression is induced under low pH (4.5) during replication inside the epithelial host cell, but it is not involved in sensing or resisting to this condition. Since the RcsCDB system is activated under acidic conditions, we investigated whether this system is able to modulate STM1485 expression. We demonstrated that acid-induced activation of the RcsB represses STM1485 transcription by directly binding to the promoter. Under the same condition, the RstA regulator activates the expression of this gene. Physiologically, we observed that RcsB-dependent repression is required for the survival of bacteria when they are exposed to pancreatic fluids. We hypothesized that STM1485 plays an important role in Salmonella adaptation to pH changes, during transition in the gastrointestinal tract. We suggest that bacteria surviving the gastrointestinal environment invade the epithelial cells, where they can remain in vacuoles. In this new environment, acidity and magnesium starvation activate the expression of the RstA regulator in a PhoPQ-dependent manner, which in turn induces STM1485 expression. These levels of STM1485 allow increased bacterial replication within vacuoles to continue the course of infection.

  • The C7-aminomethylpyrrolidine group rescues the activity of a thio-fluoroquinolone
    Biochimie (IF 3.188) Pub Date : 2019-02-11
    Sarah R.C. Lentz, Pratik R. Chheda, Lisa M. Oppegard, Tyrell R. Towle, Robert J. Kerns, Hiroshi Hiasa

    A Mg2+-water bridge between the C-3, C-4 diketo moiety of fluoroquinolones and the conserved amino acid residues in the GyrA/ParC subunit is critical for the binding of a fluoroquinolone to a topoisomerase-DNA covalent complex. The fluoroquinolone UING-5-249 (249) can bind to the GyrB subunit through its C7-aminomethylpyrrolidine group. This interaction is responsible for enhanced activities of 249 against the wild type and quinolone-resistant mutant topoisomerases. To further evaluate the effects of the 249-GyrB interaction on fluoroquinolone activity, we examined the activities of decarboxy- and thio-249 against DNA gyrase and conducted docking studies using the structure of a gyrase-ciprofloxacin-DNA ternary complex. We found that the 249-GyrB interaction rescued the activity of thio-249 but not that of decarboxy-249. A C7-group that binds more strongly to the GyrB subunit may allow for modifications at the C-4 position, leading to a novel compound that is active against the wild type and quinolone-resistant pathogens.

  • Morphine, but not methadone, inhibits microsomal prostaglandin E synthase-1 and prostaglandin-endoperoxide synthase 2 in lipopolysaccharide-stimulated horse synoviocytes
    Biochimie (IF 3.188) Pub Date : 2019-02-11
    S.V. Schwarzbach, C.F. Melo, P.L.P. Xavier, K.C. Roballo, Y.G. Cordeiro, C.E. Ambrósio, H. Fukumasu, A.B. Carregaro

    Osteoarthritis (OA) is one of the main locomotor disorders in horses. Although nonsteroidal anti-inflammatory drugs are the first-line treatment for OA, opioids could also be used. In previous studies, opioids showed promising anti-inflammatory and analgesic effects. In this study, we aimed to investigate the effects of two opioids (morphine and methadone) against inflammation in lipopolysaccharide (LPS)-stimulated synoviocytes by analyzing microsomal prostaglandin E synthase-1 (mPGES-1) and prostaglandin-endoperoxide synthase 2 (PTGS2) expression. Synoviocytes were obtained from the joints at the distal limbs of dead animals. The cytotoxic effects of LPS, morphine, and methadone were investigated by using a cell viability assay with crystal violet dye. Synoviocytes were treated with LPS, LPS plus morphine, or LPS plus methadone for 3, 6, and 12 h, and mPGES-1 and PTGS2 expression was measured using real-time polymerase chain reaction. LPS, morphine, and methadone did not affect the viability of synoviocytes, even at high concentrations. LPS treatment increased mPGES-1 and PTGS2 expression, whereas morphine inhibited the increase in mPGES-1 and PTGS2 expression in LPS-stimulated synoviocytes. Methadone did not inhibit mPGES-1 or PTGS2 expression. These results suggest that morphine may exhibit anti-inflammatory effect; therefore, it might be beneficial for the treatment of OA.

  • Thyroid Peroxidase as a dual active site enzyme: Focus on biosynthesis, hormonogenesis and thyroid disorders of autoimmunity and cancer
    Biochimie (IF 3.188) Pub Date : 2019-02-10
    Marlena Godlewska, Paul J. Banga

    Thyroid peroxidase (TPO) is the key enzyme involved in thyroid hormone synthesis. Autoantibodies to TPO (TPOAbs) are a hallmark of autoimmune thyroid disease (AITD). Here, we highlight recent progress over several years in understanding TPO biochemistry and function in various pathologies. TPO undergoes complex post-translational modifications as a dimer in endoplasmic reticulum during secretory pathway to apical membrane of thyrocytes. In silico modelling of TPO dimer has provided new information into the two enzyme active site regions and autoantigenic determinants. TPO and hydrogen peroxide generating DUOX and caveolin-1 form a complex known as thyroxisome to bring together in close proximity the components of hormone synthesis in apical membrane. Autoimmunity to TPO is characterised by autoantibodies and T cell reactivity in Hashimoto’s disease and Graves’ disease. TPOAbs are directed predominantly to two immunodominant determinants (IDR) termed IDR-A and IDR-B regions, with the latter antibodies more predominant in autoimmune disease. Strong genetic risk has been shown to be associated with TPOAbs for AITD development. A different antibody with unusual features of bispecificity for both TPO and thyroglobulin may play protective role in Hashimoto’s disease. In the context of TPO biology in human cancer, thyroid tumour tissue and breast cancer differ in TPO expression and isoform composition. In thyroid cancer, TPO expression is decreased partly by the BRAF (V600E) mutation, with direct impact on significant hormone production. TPOAbs may play a protective role in breast cancer development. An understanding of TPO and its unique two enzymatic active sites and autoantigenic determinants continues to add new knowledge on the biochemistry and immunology of this enzyme.

  • 更新日期:2019-02-05
  • The n-3 docosapentaenoic acid (DPA): a new player in the n-3 long chain polyunsaturated fatty acid family
    Biochimie (IF 3.188) Pub Date : 2019-02-01
    Drouin Gaetan, Rioux Vincent, Legrand Philippe
  • Role of epoxy-fatty acids and epoxide hydrolases in the pathology of neuro-inflammation
    Biochimie (IF 3.188) Pub Date : 2019-02-01
    Sean D. Kodani, Christophe Morisseau

    Neuroinflammation is a physiologic response aimed at protecting the central nervous system during injury. However, unresolved and chronic neuroinflammation can lead to long term damage and eventually neurologic disease including Parkinson’s disease, Alzheimer’s disease and dementia. Recently, enhancing the concentration of epoxyeicosatrienoic acids (EETs) through blocking their hydrolytic degradation by soluble epoxide hydrolase (sEH) has been applied towards reducing the long-term damage associated with central neurologic insults. Evidence suggests this protective effect is mediated, at least in part, through polarization of microglia to an anti-inflammatory phenotype that blocks the inflammatory actions of prostaglandins and promotes wound repair. This mini-review overviews the epidemiologic basis for using sEH inhibition towards neuroinflammatory disease and pharmacologic studies testing sEH inhibition in several neurologic diseases. Additionally, the combination of sEH inhibition with other eicosanoid signaling pathways is considered as an enhanced approach for developing potent neuroprotectants.

  • Elucidation of the mechanism of disulfide exchange between staphylococcal thioredoxin2 and thioredoxin reductase2: a structural insight
    Biochimie (IF 3.188) Pub Date : 2019-01-31
    Madhuparna Bose, Sudipta Bhattacharyya, Rupam Biswas, Amlan Roychowdhury, Atanu Bhattacharjee, Ananta Kumar Ghosh, Amit Kumar Das
  • Structure function relationships in three lipids A from the Ralstonia genus rising in obese patients
    Biochimie (IF 3.188) Pub Date : 2019-01-28
    Wei Zhang-Sun, François Tercé, Remy Burcelin, Alexey Novikov, Matteo Serino, Martine Caroff

    The identification of a functional molecular moiety relating the lipopolysaccharides (LPSs) to their capacity to induce inflammation-mediated metabolic diseases needed to be performed. We previously described a proportional increase in the relative abundance of the 16SrDNA bacterial gene from the genus Ralstonia, within the microbiota from the adipose tissue stroma vascular fraction of obese patients, suggesting a causal role of the bacteria. Therefore, we first characterized the structures of the lipids A, the inflammatory inducing moieties of LPSs, of three Ralstonia species: Ralstonia eutropha, R. mannitolilytica and R. pickettii, and then compared each, in terms of in vitro inflammatory capacities. R. pickettii lipid A displaying only 5 Fatty Acids (FA) was a weaker inducer of inflammation, compared to the two other species harboring hexa-acylated lipids A, despite the presence of 2 AraN substituents on the phosphate groups. With regard to in vitro pro-inflammatory activities, TNF-α and IL-6 inducing capacities were compared on THP-1 cells treated with LPSs isolated from the three Ralstonia. R. pickettii, with low inflammatory capacities, and recently involved in nosocomial outcomes, could explain the low inflammatory level reported in previous studies on diabetic patients and animals. In addition, transmission electron microscopy was performed on the three Ralstonia species. It showed that the R. pickettii under-acylated LPSs, with a higher level of phosphate substitution had the capacity of producing more outer membrane vesicles (OMVs). The latter could facilitate transfer of LPSs to the blood and explain the increased low-grade inflammation observed in obese/diabetic patients.

  • Development of conformational antibodies targeting Cripto-1 with neutralizing effects in vitro
    Biochimie (IF 3.188) Pub Date : 2019-01-28
    Giuseppina Focà, Emanuela Iaccarino, Annalia Focà, Luca Sanguigno, Gustavo Untiveros, Maria Cuevas-Nunez, Luigi Strizzi, Antonio Leonardi, Menotti Ruvo, Annamaria Sandomenico

    Human Cripto-1 (Cripto-1), the founding member of the EGF-CFC superfamily, is a key regulator of many processes during embryonic development and oncogenesis. Cripto-1 is barely present or even absent in normal adult tissues while it is aberrantly re-expressed in various tumors. Blockade of the CFC domain-mediated Cripto-1 functions is acknowledged as a promising therapeutic intervention point to inhibit the tumorigenic activity of the protein. In this work, we report the generation and characterization of murine monoclonal antibodies raised against the synthetic folded CFC [112-150] domain of the human protein. Through subtractive ELISA assays clones were screened for the ability to specifically recognize “hot spot” residues on the CFC domain, which are crucial for the interaction with Activin Type I receptor (ALK4) and GRP78. On selected antibodies, SPR and epitope mapping studies have confirmed their specificity and have revealed that recognition occurs only on a conformational epitope. Furthermore, FACS analyses have confirmed the ability of 1B4 antibody to recognize the membrane-anchored and soluble native Cripto-1 protein in a panel of human cancer cells. Finally, we have evaluated its functional effects through in vitro cellular signaling assays and cell cycle analysis. These findings suggest that the selected anti-CFC mAbs have the potential to neutralize the protein oncogenic activity and may be used as theranostic molecules suitable as tumor homing agents for Cripto-1-overexpressing cancer cells and tissues and to overcome drug-resistance in routine cancer therapies.

  • Emergence of membrane sphingolipids as a potential therapeutic target
    Biochimie (IF 3.188) Pub Date : 2019-01-28
    Sunil Kumar Sahu, Yusuf A. Hannun, Nan Yao

    Background Though sphingolipids are ubiquitously present in eukaryotic cells, but until the last decade, they were merely considered as a structural component of the plasma membrane with limited function. However, over the last decade, numerous functions have been ascribed to sphingolipids after the seminal discoveries on the bioactivities of several sphingolipids. Scope of Review Sphingolipids are now well-recognized signals for fundamental cellular processes. Here we discussed about the advent of several sphingolipids components as potential therapeutic target for both human and plants. Major Conclusions Sphingolipid contents and/or sphingolipid-metabolizing enzyme expression/activity often get impaired during pathophysiological conditions, and hence manipulation of this signaling pathway may be beneficial in disease diagnosis, and the plasma concentrations can serve as an important prognostic and diagnostic marker for the disease. General Significance Sphingolipids are emerging as a goldmine for new therapeutic drug targets with promising new applications (cosmeceutical and nutraceutical), thereby opening new avenues for pharmaceuticals and nutraceutical industries.

  • New Pieces to the Carbon Metabolism Puzzle of Nitrosomonas europaea: Kinetic characterization of glyceraldehyde-3phosphate and succinate semialdehyde dehydrogenases
    Biochimie (IF 3.188) Pub Date : 2019-01-26
    María Cecilia Corregido, Matías Damián Asención Diez, Alberto Álvaro Iglesias, Claudia Vanesa Piattoni

    Nitrosomonas europaea is a chemolithotroph that obtains energy through the oxidation of ammonia to hydroxylamine while assimilates atmospheric CO2 to cover the cell carbon demands for growth. This microorganism plays a relevant role in the nitrogen biogeochemical cycle on Earth but its carbon metabolism remains poorly characterized. Based on sequence homology, we identified two genes (cbbG and gabD) coding for redox enzymes in N. europaea. Cloning and expression of the genes in Escherichia coli, allowed the production of recombinant enzymes purified to determine their biochemical properties. The protein CbbG is a glyceraldehyde-3-phosphate (Ga3P) dehydrogenase (Ga3PDHase) catalyzing the reversible oxidation of Ga3P to 1,3-bis-phospho-glycerate (1,3bisPGA), using specifically NAD+/NADH as cofactor. CbbG showed ∼6-fold higher Km value for 1,3bisPGA but ∼5-fold higher kcat for the oxidation of Ga3P. The protein GabD irreversibly oxidizes Ga3P to 3Pglycerate using NAD+ or NADP+, thus resembling a non-phosphorylating Ga3PDHase. However, the enzyme showed ∼6-fold higher Km value and three orders of magnitude higher catalytic efficiency with succinate semialdehyde (SSA) and NADP+. Indeed, the GabD protein identity corresponds to an SSA dehydrogenase (SSADHase). CbbG seems to be the only Ga3PDHase present in N. europaea; which would be involved in reducing triose-P during autotrophic carbon fixation. Otherwise, in cells grown under conditions deprived of ammonia and oxygen, the enzyme could catalyze the glycolytic step of Ga3P oxidation producing NADH. As an SSADHase, GabD would physiologically act producing succinate and preferentially NADPH over NADH; thus being part of an alternative pathway of the tricarboxylic acid cycle converting α-ketoglutarate to succinate. The properties determined for these enzymes contribute to better identify metabolic steps in CO2 assimilation, glycolysis and the tricarboxylic acid cycle in N. europaea. Results are discussed in the framework of metabolic pathways that launch biosynthetic intermediates relevant in the microorganism to develop and fulfill its role in nature.

  • N-3 polyunsaturated fatty acids: an innovative strategy against obesity and related metabolic disorders, intestinal alteration and gut microbiota dysbiosis
    Biochimie (IF 3.188) Pub Date : 2019-01-25
    Jérôme Bellenger, Sandrine Bellenger, Quentin Escoula, Célia Bidu, Michel Narce

    Obesity is now widely recognized to be associated with low-grade systemic inflammation. It has been shown that high-fat feeding modulates gut microbiota which strongly increased intestinal permeability leading to lipopolysaccharide absorption causing metabolic endotoxemia that triggers inflammation and metabolic disorders. N-3 polyunsaturated fatty acids (PUFAs) have been shown associated with anti-obesity properties, but results still remain heterogeneous and very few studies underlined the metabolic pathways involved. Thus, the use of Fat-1 transgenic mice allows to better understanding whether endogenous n-3 PUFAs enrichment contributes to obesity and associated metabolic disorders prevention. It specially evidence that such effects occur through modulations of gut microbiota and intestinal permeability. Then, by remodeling gut microbiota, endogenous n-3 PUFAs improve HF/HS-diet induced features of the metabolic syndrome which in turn affects host metabolism. Thus, increasing anti-obesogenic microbial species in the gut microbiota population (i.e Akkermansia) by appropriate n-3 PUFAs may represent a promising strategy to control or prevent metabolic diseases.

  • An aptamer based thermofluorimetric assay for ethanolamine
    Biochimie (IF 3.188) Pub Date : 2019-01-25
    Mostafa Mahmoud, Stefan Laufer, Hans-Peter Deigner

    There is a great need for fast, simple and precise diagnostic assays capable of direct quantification of biomarkers in complex biological matrices. Yet, the commonly used techniques such as ELISA/Immunoassays are tedious and involve various steps e.g. blocking, washing and signal development. Moreover, most of these assays have very limited ability of detecting small molecules and have hardly any multiplexing capabilities. The gold standard and alternative, mass-spectrometry, however, depends upon expensive hardware and is incompatible with point of care (POC) diagnostics. As opposed to POC assays for proteins or larger targets where variable formats are readily available. Here, we present a simple, versatile and fast one-step assay for detecting a small molecule, ethanolamine as example. The assay makes use of commonly available qPCR machines to detect target-concentration dependent shifts in the melting temperatures of aptamer beacons. The method allows detection of ethanolamine in the low nM range without requiring tedious elaboration of assay conditions as required for molecular beacons at room temperature. If generalizable, it may change the situation of small molecule assays significantly.

  • Dimers of serotonin receptors: impact on ligand affinity and signaling
    Biochimie (IF 3.188) Pub Date : 2019-01-24
    Luc Maroteaux, Catherine Béchade, Anne Roumier

    Membrane receptors often form complexes with other membrane proteins that directly interact with different effectors of the signal transduction machinery. G-protein-coupled receptors (GPCRs) were for long time considered as single pharmacological entities. However, evidence for oligomerization appeared for various classes and subtypes of GPCRs. This review focuses on metabotropic serotonin (5-hydroxytryptamine, 5-HT) receptors, which belong to the rhodopsin-like class A of GPCRs, and will summarize the convergent evidence that homo- and hetero-dimers containing 5-HT receptors may exist in transfected cells and in-vivo. We will show that complexes involving 5-HT receptors may acquire new signal transduction pathways and new physiological roles. In some cases, these complexes participate in disease-specific deregulations, that can be differentially affected by various drugs. Hence, selecting receptor complex-specific responses of these heterodimers may constitute an emerging strategy likely to improve beneficial therapeutic effects.

  • Kinetic mechanism of Enterococcus faecium D-aspartate ligase
    Biochimie (IF 3.188) Pub Date : 2019-01-22
    Veronika Škedelj, Urša Pečar Fonović, Peter Molek, Sophie Magnet, Jean-Luc Mainardi, Didier Blanot, Stanislav Gobec, Jure Stojan, Anamarija Zega

    Enterococcus faecium D-aspartate ligase (Aslfm) is a peptide bond-forming enzyme that is involved in the peptidoglycan assembly pathway. It catalyzes the ATP-dependent ligation of the β-carboxylate of D-Asp to the ε-amino group of L-Lys in the nucleotide precursor UDP- MurNAc-pentapeptide. The enzyme is of interest as a target of new, potential, narrow-spectrum antibiotics directed against multiresistant E. faecium. The kinetic mechanism of Aslfm has not been fully characterized. To determine it, a progress curve analysis of Aslfm catalytic process using pyruvate kinase/lactate dehydrogenase ATPase detection assay was performed. With an inspection of the shape of measured progress curves and the results of specific qualitative experiments, the Aslfm reaction mechanism was singled out. The proposed Aslfm kinetics reaction scheme was evaluated by fitting the parameters of the corresponding differential equations to progress curves using the computer program ENZO. The complete kinetic analysis result is consistent with the substrate binding order 1) ATP, 2) D-Asp, and 3) UDP-MurNAc-pentapeptide. The analysis suggests that slowly establishing non-productive equilibria between the free and ATP-bound enzyme with the participating pentapeptide are responsible for initial reaction burst followed by a steady-state period before the complete depletion of the reactant added in the lowest concentration.

  • 更新日期:2019-01-23
  • Vasopressin receptors in islets enhance glucose tolerance, pancreatic beta-cell secretory function, proliferation and survival
    Biochimie (IF 3.188) Pub Date : 2019-01-21
    Shruti Mohan, R. Charlotte Moffett, Keith G. Thomas, Nigel Irwin, Peter R. Flatt

    Arginine vasopressin (AVP), a peptide secreted from the posterior pituitary, is chiefly regarded as a hormone involved in the regulation of body fluid balance and osmolality. However, recent evidence has revealed that posterior pituitary hormones can exert important actions on endocrine pancreatic function. In the present study, the presence of AVP receptors, namely Avpr1a (V1a), Avpr1b (V1b) and Avpr2 (V2) was demonstrated in murine islets as well as rodent BRIN BD11 and human 1.1B4 beta-cells. Further to this, AVP was shown to induce significant concentration-dependent (10-12 – 10-6 M) increases of insulin release from both rodent and human beta-cells, as well as mouse islets. Insulinotropic actions of AVP were completely annulled by specific V1a or V1b receptor antagonists, and partially abolished by an oxytocin receptor antagonist. In addition, beta-cell insulin secretory actions of AVP were augmented by both IBMX (200 μM) and KCl (30 mM) and linked to significantly increased cAMP production and [Ca2+]i. AVP substantially increased proliferation of rodent and human beta-cells. Moreover, AVP fully protected against cytokine-induced beta-cell apoptosis. AVP had no effect on glucagon secretion. Immunohistochemical examination of beta- and alpha-cells revealed co-expression of AVP with glucagon, and particularly insulin. Finally, administration of AVP in combination with glucose to mice significantly reduced blood glucose, which was associated with increased plasma insulin. These data indicate that AVP possesses novel and potentially important effects on pancreatic endocrine function. Understanding disturbances in islet AVP receptor signalling could reveal insight into the beta-cell defects associated with diabetes.

  • Properties of L-amino acid deaminase: en route to optimize bioconversion reactions
    Biochimie (IF 3.188) Pub Date : 2019-01-20
    Paolo Motta, Loredano Pollegioni, Gianluca Molla
  • Structural basis for the design of selective inhibitors for Schistosoma mansoni dihydroorotate dehydrogenase
    Biochimie (IF 3.188) Pub Date : 2019-01-19
    M.Cristina Nonato, Ricardo A.P. de Pádua, Juliana S. David, Renata A.G. Reis, Giovani P. Tomaleri, Humberto D’Muniz Pereira, Felipe A. Calil
  • M17 aminopeptidases diversify function by moderating their macromolecular assemblies and active site environment
    Biochimie (IF 3.188) Pub Date : 2019-01-14
    Nyssa Drinkwater, Tess R. Malcolm, Sheena McGowan

    The family of M17 aminopeptidases (alias ‘leucine aminopeptidases’, M17-LAPs) utilizes a highly conserved hexameric structure and a binuclear metal center to selectively remove N-terminal amino acids from short peptides. However, M17-LAPs are responsible for a wide variety of functions that are seemingly unrelated to proteolysis. Herein, we aimed to investigate the myriad of functions attributed to M17. Further, we attempted to differentiate between the different molecular mechanisms that allow the conserved hexameric structure of an M17-LAP to mediate such diverse functions. We have provided an overview of research that identifies precise physiological roles of M17-LAPs, and the distinct mechanisms by which the enzymes moderates those roles. The review shows that the conserved hexameric structure of the M17-LAPs has an extraordinary capability to moderate different molecular mechanisms. We have broadly categorized these mechanisms as ‘aminopeptidase-based’, which include the characteristic proteolysis reactions, and ‘association-driven’, which involves moderation of the molecule’s macromolecular assembly and higher order complexation events. The different molecular mechanisms are capable of eliciting very different cellular outcomes, and must be regarded as distinct when the physiological roles of this large and important family are considered.

  • Biomarkers Mapping of Neuropathic Pain in a Nerve Chronic Constriction Injury Mice Model
    Biochimie (IF 3.188) Pub Date : 2019-01-11
    S. Vincenzetti, S. Pucciarelli, Y. Huang, M. Ricciutelli, C. Lambertucci, R. Volpini, G. Scuppa, L. Soverchia, M. Ubaldi, V. Polzonetti
  • Molecular cloning and characterization of porcine Na⁺/K⁺-ATPase isoform α4
    Biochimie (IF 3.188) Pub Date : 2019-01-08
    Knud Larsen, Carina Henriksen, Kaja Kjær Kristensen, Jamal Momeni, Leila Farajzadeh

    Na+/K+-ATPase is responsible for maintaining electrochemical gradients of Na+ and K+, which is essential for a variety of cellular functions including neuronal activity. The α-subunit of the Na+/K+-ATPase is composed of four different polypeptides (α1–α4) encoded by different genes. Na,K-ATPase α4, encoded by the ATP1A4 gene, is expressed in testis and in male germ cells of humans, rats and mice. The α4 polypeptide has an important role in sperm motility, and is essential for male fertility. Here we present the RT-PCR cloning and characterization of the porcine ATP1A4 cDNA coding for Na⁺/K⁺-ATPase polypeptide α4. The Na⁺/K⁺-ATPase polypeptide α4, consisting of 1030 amino acids, displays a high homology with its human counterpart (86 %). Phylogenetic analysis demonstrated that porcine Na⁺/K⁺-ATPase polypeptide α4 is closely related to other mammalian counterparts. In addition, the genomic structure of the porcine ATP1A4 gene was determined, and the intron-exon organization was found to be similar to that of the human ATP1A4 gene. The promoter sequence for the porcine ATP1A4 gene was also identified. Investigation of the genetic variation in the porcine ATP1A4 gene revealed a missense A/G SNP in exon 18. This A/G polymorphism results in a substitution of a methionine to a glycine residue (M888G). A very high overall DNA methylation rate of the ATP1A4 gene, 70-80 %, was observed in both brain and liver. Expression analysis demonstrated that the porcine ATP1A4 gene is predominantly expressed in testis. The sequence of the porcine ATP1A4 cDNA encoding the Na⁺/K⁺-ATPase α4 protein has been submitted to GenBank under the accession number GenBank Accession No. MG587082.

  • Oxalomalate reduces tumor progression in melanoma via ROS-dependent proapoptotic and antiangiogenic effects
    Biochimie (IF 3.188) Pub Date : 2019-01-09
    Sung Hwan Kim, In Sup Kil, Oh-Shin Kwon, Boem Sik Kang, Dong-Seok Lee, Hyun-Shik Lee, Jin Hyup Lee, Jeen-Woo Park

    The potent cytotoxicity of reactive oxygen species (ROS) can cause various diseases, however, it may also serve as a powerful chemotherapeutic strategy capable of killing cancer cells. Oxalomalate (OMA, α-hydroxy-β-oxalosuccinic acid), a tricarboxylic acid intermediate, is a well-known competitive inhibitor of two classes of NADP+-dependent isocitrate dehydrogenase (IDH) isoenzymes, which serve as the major antioxidants and redox regulators in the mitochondria and cytosol. In this study, we investigated the therapeutic effects of OMA in melanoma and elucidated the associated underlying mechanisms of action using in vitro and in vivo models. OMA targeting IDH enzymes suppressed melanoma growth through activation of apoptosis and inhibition of angiogenesis. Mechanistically, our findings showed that OMA activated p53-mediated apoptosis through ROS-dependent ATM-Chk2 signaling and reduced the expression of vascular endothelial growth factor through ROS-dependent E2F1-mediated hypoxia inducible factor-1α degradation. In particular, OMA-induced suppression of IDH activity resulted in induction of ROS stress response, ultimately leading to apoptotic cell death and antiangiogenic effects in melanoma cells. Thus, OMA might be a potential candidate drug for melanoma skin cancer therapy.

  • Osmolytes resist against harsh osmolarity: Something old something new
    Biochimie (IF 3.188) Pub Date : 2019-01-07
    Seyed Mahdi Hosseiniyan Khatibi, Fatemeh Zununi Vahed, Simin Sharifi, Mohammadreza Ardalan, Mohammadali Mohajel Shoja, Sepideh Zununi Vahed

    From the halophilic bacteria to human, cells have to survive under the stresses of harsh environments. Hyperosmotic stress is a process that triggers cell shrinkage, oxidative stress, DNA damage, and apoptosis and it potentially contributes to a number of human diseases. Remarkably, by high salts and organic solutes concentrations, a variety of organisms struggle with these conditions. Different strategies have been developed for cellular osmotic adaptations among which organic osmolyte synthesis/accumulation is a conserved once. Osmolytes are naturally occurring solutes used by cells of several halophilic (micro) organisms to preserve cell volume and function. In this review, the osmolytes diversity and their protective roles in harsh hyperosmolar environments from bacteria to human cells are highlighted. Moreover, it provides a close look at mammalian kidney osmoregulation at a molecular level. This review provides a concise view on the recent developments and advancements on the applications of osmolytes. Identification of disease-related osmolytes and their targeted-delivery may be used as a therapeutic measurement for treatment of the pathological conditions and the inherited diseases related to protein misfolding and aggregation. The molecular and cellular aspects of cell adaptation against harsh environmental osmolarity will benefit the development of effective drugs for many diseases.

  • FNDC5: a novel player in metabolism and metabolic syndrome
    Biochimie (IF 3.188) Pub Date : 2019-01-03
    Richard Y. Cao, Hongchao Zheng, Damian Redfearn, Jian Yang

    Half a decade ago, transmembrane protein fibronectin type III domain-containing protein 5 (FNDC5) was found to be cleaved as a novel myokine irisin, which burst into prominence for browning of white adipose tissue during exercise. However, FNDC5, the precursor of irisin, has been paid relatively little attention compared with irisin despite evidence that FNDC5 is associated with the metabolic syndrome, which accounts for one-fourth of the world’s adult population and contributes to diabetes, cardiovascular disease and all-cause mortality. Besides N-terminal and C-terminal sequences, the FNDC5 protein contains an irisin domain and a short transmembrane region. FNDC5 has shown to be widely distribute in different tissues and is highly expressed in heart, brain, liver, and skeletal muscle. Clinical studies have demonstrated that FNDC5 is essential for maintaining metabolic homeostasis and dysregulation of FNDC5 will lead to systemic metabolism imbalance and the onset of metabolic disorders. Growing evidence has suggested that FNDC5 gene polymorphisms are related to health and disease in different human populations. Additionally, FNDC5 has been found relevant to the regulation of metabolism and metabolic syndrome through diverse upstream and downstream signaling pathways in experimental studies. The present review summarizes the characteristics, clinical significance, and molecular mechanisms of FNDC5 in metabolic syndrome and proposes a novel concept that FNDC5 is activated by forming a putative ligand-receptor complex. Knowledge about the role of FNDC5 may be translated into drug development and clinical applications for the treatment of metabolic disorders.

  • Amelioration of sepsis-induced acute kidney injury through inhibition of inflammatory cytokines and oxidative stress in dendritic cells and neutrophils respectively in mice: Role of spleen tyrosine kinase signaling
    Biochimie (IF 3.188) Pub Date : 2018-12-29
    Naif O. Al-Harbi, Ahmed Nadeem, Sheikh F. Ahmad, Mohammed M. Alanazi, Abdullah A. Aldossari, Fawaz Alasmari

    Sepsis often leads to complications such as acute kidney injury (AKI) which is reported to range from 30-50% in critically ill patients. Dendritic (DCs) and neutrophils play a decisive role in the advancement of AKI through release of inflammatory cytokines and reactive oxygen species (ROS) respectively. Both of these processes are assumed to be controlled by spleen tyrosine kinase (Syk) signaling in DCs and neutrophils. However, the role of Syk signaling in these immune cells in sepsis-induced AKI has not been investigated. Therefore, the purpose of this study was to evaluate the effect of a Syk inhibitor, R406 on sepsis-induced AKI in a mouse model. Renal function (creatinine/blood urea nitrogen), inflammatory cytokines (IL-6/MCP-1) in CD11c+DCs and oxidant parameters in neutrophils [inducible nitric oxide synthase (iNOS), NADPH oxidase (NOX2), nitrotyrosine] were assessed. Our results showed elevated expression of Syk in neutrophils and CD11c+DC which was linked with increased IL-6/MCP-1 in CD11c+ DCs, and iNOS, NOX2 and nitrotyrosine in neutrophils during sepsis-induced AKI. Inhibitor of Syk signaling, R406 led to improvement of sepsis-induced AKI as depicted by an attenuation of creatinine/blood urea nitrogen in serum, renal myeloperoxidase activity, and repair of tubular structures in kidney. Further, R406 led to a decrease in IL-6/MCP-1 in CD11c+ DCs, and iNOS, NOX2 and nitrotyrosine in neutrophils during sepsis-induced AKI. In conclusion, our study proposes that Syk signaling in DCs and neutrophils plays a critical role during sepsis-induced AKI. Therefore, Syk inhibition in innate immune cells might serve as an effective strategy to limit inflammatory cascade during AKI.

  • Cloning and expression of ATP N-glycosidase from the freshwater sponge Ephydatia muelleri
    Biochimie (IF 3.188) Pub Date : 2018-12-29
    Tõnu Reintamm, Kerli Vallmann, Kaidi Kolk, Mailis Päri, Annika Lopp, Nele Aas-Valleriani, Merike Kelve

    Previously we had discovered unusual enzymatic activity in the marine sponge Axinella polypoides, ATP N-glycosidase (Reintamm et al, 2003). We show here that the Ephydatia muelleri mRNA encoding protein with PNP_UDP_1 (phosphorylase superfamily) signature is the secreted ATP N-glycosidase. The functionality of the protein was established by recombinant expression in Pichia pastoris. In addition to the enzymatic domain, the full-length protein contains the N-terminal cysteine-rich domain belonging to the subfamily SCP_HrTT-1 (cd05559) of the SCP (sperm coating protein) superfamily (cl00133).

  • Biochemical and structural insights into PLP fold type IV transaminase from Thermobaculum terrenum
    Biochimie (IF 3.188) Pub Date : 2018-12-29
    Ekaterina Yu. Bezsudnova, Konstantin M. Boyko, Alena Yu. Nikolaeva, Yulia S. Zeifman, Tatiana V. Rakitina, Dmitry A. Suplatov, Vladimir O. Popov

    The high catalytic efficiency of enzymes under reaction conditions is one of the main goals in biocatalysis. Despite the dramatic progress in the development of more efficient biocatalysts by protein design, the search for natural enzymes with useful properties remains a promising strategy. The PLP-dependent transaminases represent a group of industrially important enzymes due to their ability to stereoselectively transfer amino groups between diverse substrates; however, the complex mechanism of substrate recognition and conversion makes the design of transaminases a challenging task. Here we report a detailed structural and kinetic study of thermostable transaminase from the bacterium Thermobaculum terrenum (TaTT) using the methods of enzyme kinetics, X-ray crystallography and molecular modeling. TaTT can convert L-branched-chain and L-aromatic amino acids as well as (R)-(+)-1-phenylethylamine at a high rate and with high enantioselectivity. The structures of TaTT in complex with the cofactor pyridoxal 5’-phosphate covalently bound to enzyme and in complex with its reduced form, pyridoxamine 5’-phosphate, were determined at resolutions of 2.19 Å and 1.5 Å, and deposited in the Protein Data Bank as entries 6GKR and 6Q8E, respectively. TaTT is a fold-type IV pyridoxal 5’-phosphate-dependent enzyme. In terms of structural similarity, the enzyme is close to known branched-chain amino acid aminotransferases, but differences in characteristic sequence motifs in the active site were observed in TaTT compared to canonical branched-chain amino acid aminotransferases, which can explain the improved binding of aromatic amino acids and (R)-(+)-1-phenylethylamine. This study has shown for the first time that high substrate specificity towards both various L-amino acids and (R)-primary amines can be implemented within one pyridoxal 5’-phosphate-dependent active site of fold-type IV. These results complement our knowledge of the catalytic diversity of transaminases and indicate the need for further biochemical and bioinformatic studies to understand the sequence-structure-function relationship in these enzymes.

  • The Relationship Between the Placental Serotonin Pathway and Fetal Growth Restriction
    Biochimie (IF 3.188) Pub Date : 2018-12-31
    Suveena Ranzil, David W. Walker, Anthony J. Borg, Euan M. Wallace, Peter R. Ebeling, Padma Murthi

    Fetal growth restriction (FGR) is a complex disorder of human pregnancy that leads to poor health outcomes in offspring. These range from immediate risks such as perinatal morbidity and stillbirths, to long-term complications including severe neurodevelopmental problems. Despite its relatively high global prevalence, the aetiology of FGR and its complications is not currently well understood. We now know that serotonin (5-HT) is synthesised in the placenta and is crucial for early fetal forebrain development in mice. However, the contribution of a disrupted placental 5-HT synthetic pathway to the pathophysiology of placental insufficiency in FGR and its significant fetal neurodevelopmental complications are unclear.

  • The human ribosome can interact with the abasic site in mRNA via a specific peptide of the uS3 protein located near the mRNA entry channel
    Biochimie (IF 3.188) Pub Date : 2018-12-27
    Anastasia S. Ochkasova, Maria I. Meschaninova, Aliya G. Venyaminova, Anton V. Ivanov, Dmitri M. Graifer, Galina G. Karpova

    The small subunit ribosomal protein uS3 is a critically important player in the ribosome-mRNA interactions during translation and has numerous functions not directly related to protein synthesis in eukaryotes. A peculiar feature of the human uS3 protein is the ability of its fragment 55-64 exposed on the 40S subunit surface near the mRNA entry channel to form cross-links with 3’-terminal dialdehyde derivatives of various unstructured RNAs and with abasic sites in single-stranded DNAs. Here we showed that the ability of the above uS3 fragment to cross-link to abasic sites in DNAs is inherent only in mature cytoplasmic 40S subunits, but not nuclear pre-40S particles, which implies that it may be relevant to the ribosome-mRNA interplay. To clarify this issue, we investigated interactions of human ribosomes with synthetic mRNA analogues bearing an abasic site protected by a photocleavable group at the 3’-termini. We found that these mRNA analogues can form specific complexes with 80S ribosomes and 40S subunits, where the undamaged upstream part of the analogue is fixed in the mRNA binding channel by interaction with the P-site tRNA, and the downstream part located outside the ribosome is cross-linked to the uS3 fragment 55-64. The yield of cross-links of the mRNA analogues was rather high when their undamaged parts were bound to the mRNA channel prior to deprotection of the abasic site enabling its covalent attachment to the 40S subunit via the uS3 protein, but not vice versa. Based on our findings, one can assume that abasic sites, which can occur in mRNAs due to oxidative stress and ageing, directly interact with the uS3 fragment exposed on the 40S subunit surface near the mRNA entry channel during translation. Consequently, the 40S subunit can be considered as a potential mRNA quality controller.

  • ORP2 interacts with phosphoinositides and controls the subcellular distribution of cholesterol
    Biochimie (IF 3.188) Pub Date : 2018-12-24
    Annika Koponen, Amita Arora, Kohta Takahashi, Henriikka Kentala, Annukka Kivelä, Eeva Jääskeläinen, Johan Peränen, Pentti Somerharju, Elina Ikonen, Tapani Viitala, Vesa M. Olkkonen

    ORP2 is a sterol-binding protein with documented functions in lipid and glucose metabolism, Akt signaling, steroidogenesis, cell adhesion, migration and proliferation. Here we investigate the interactions of ORP2 with phosphoinositides (PIPs) by surface plasmon resonance (SPR), its affinity for cholesterol with a pull-down assay, and its capacity to transfer sterol in vitro. Moreover, we determine the effects of wild-type (wt) ORP2 and a mutant with attenuated PIP binding, ORP2(mHHK), on the subcellular distribution of cholesterol, and analyze the interaction of ORP2 with the related cholesterol transporter ORP1L. ORP2 showed specific affinity for PI(4,5)P2, PI(3,4,5)P3 and PI(4)P, with suggestive Kd values in the μM range. Also binding of cholesterol by ORP2 was detectable, but a Kd could not be determined. Wt ORP2 was in HeLa cells mainly detected in the cytosol, ER, late endosomes, and occasionally on lipid droplets (LDs), while ORP2(mHHK) displayed an enhanced LD localization. Overexpression of wt ORP2 shifted the D4H cholesterol probe away from endosomes, while ORP2(mHHK) caused endosomal accumulation of the probe. Although ORP2 failed to transfer dehydroergosterol in an in vitro assay where OSBP is active, its knock-down resulted in the accumulation of cholesterol in late endocytic compartments, as detected by both D4H and filipin probes. Interestingly, ORP2 was shown to interact and partially co-localize on late endosomes with ORP1L, a cholesterol transporter/sensor at ER-late endosome junctions. Our data demonstrates that ORP2 binds several phosphoinositides, both PI(4)P and multiply phosphorylated species. ORP2 regulates the subcellular distribution of cholesterol dependent on its PIP-binding capacity. The interaction of ORP2 with ORP1L suggests a concerted action of the two ORPs.

  • A comparative study of the complete lipopolysaccharide structures and biosynthesis loci of Bordetella avium, B. hinzii, and B. trematum
    Biochimie (IF 3.188) Pub Date : 2018-12-19
    Alexey Novikov, Nico Marr, Martine Caroff

    A dozen species of human and animal pathogens have been described to date in the Bordetella genus, with the majority being respiratory tract pathogens. Bordetella avium lipopolysaccharides have been shown to be important virulence factors for this bird pathogen. B. hinzii is closely related to the B. avium species, but has also been isolated from humans. B. trematum is associated to ear and blood infections in humans. Its lipid A structure, the biological active moiety of LPS, was found to be closely related to those of B. avium and B. hinzii. It is important to unveil the subtle structural modifications orchestrated during the LPS biosynthetic pathway to better understand host adaptation. The present data are also important in the context of deciphering the virulence pathways of this important genus containing the major pathogens B. pertussis and B. parapertussis, responsible for whooping cough. We recently reported the isolated lipid A structures of the three presented species, following the previously identified O-chain structures. In the present study, we provide details on the free and O-chain-linked core oligosaccharide which was required to characterize the complete LPS structures. Data are presented here in relation to relevant biosynthesis genes. The present characterization of the three species is well illustrated by Matrix Assisted Laser Desorption Mass Spectrometry experiments, and data were obtained mainly on native LPS molecules for the first time.

  • Genetic depletion of p53 attenuates cocaine-induced hepatotoxicity in mice
    Biochimie (IF 3.188) Pub Date : 2018-12-18
    Huynh Nhu Mai, Garima Sharma, Naveen Sharma, Eun-Joo Shin, Dae-Joong Kim, Duc Toan Pham, Quynh Dieu Trinh, Choon-Gon Jang, Seung Yeol Nah, Ji Hoon Jeong, Hyoung-Chun Kim

    Cocaine, an addictive drug, is known to induce hepatotoxicity via oxidative damage and proapoptosis. Since p53, a tumor suppressor gene, plays a major role in inducing oxidative stress and apoptosis, we examined the role of p53 inhibition against cocaine-induced hepatotoxicity. Cocaine treatment significantly increased oxidative parameters (i.e., reactive oxygen species, 4-hydroxylnonenal, and protein carbonyl) in the liver of wild type (WT) mice. We found that the pharmacological (i.e. pifithrin-α) and genetic (i.e. p53 knockout) inhibition of p53 significantly attenuates cocaine-induced hepatotoxicity. Cocaine treatment increased alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels in the serum of mice, signifying hepatic damage. Consistently, these increases were attenuated by inhibition of p53, implying protection against cocaine-induced hepatic damage. In addition, cocaine treatment significantly increased PKCδ, cleaved PKCδ and p53 levels in the liver of WT mice. These increases were followed by the interaction between p53 and PKCδ, and pro-apoptotic consequences (i.e., cytosolic release of cytochrome c, activation of caspase-3, increase in Bax level and decreases in Bcl-2 and Bcl-xL levels). These changes were attenuated by p53 depletion, reflecting that the critical role of PKCδ in p53-mediated apoptotic potentials. Combined, our results suggest that the inhibition of p53 is important for protection against oxidative burdens, pro-apoptotic events, and hepatic degeneration induced by cocaine.

  • Role of human Keap1 S53 and S293 residues in modulating the binding of Keap1 to Nrf2
    Biochimie (IF 3.188) Pub Date : 2018-12-18
    Shuangshuang Wei, Yechun Pei, Yuerong Wang, Huai Guan, Yonglin Huang, Tian Xing, Rodney W. Johnson, Dayong Wang

    Keap1 is deemed as a suppressor of Nrf2 in cytoplasm by sequestrating Nrf2 to proteolysis as an adapter of the Cul3-Rbx1 E3 ubiquitin ligase complex. In the study, it was proposed that post-translational modification might affect the interaction between Nrf2 and Keap1, and the profiles of the phosphorylation of amino acid residues of Keap1 and its effects on the binding of Keap1 to Nrf2 was investigated. A mass spectrometry analysis revealed that S53 and S293 were phosphorylated upon an oxidative stress. Using Keap1 proteins with amino acid residues mutated to glutamate to simulate the introduction of a negative charge by phosphorylation, it was found that a potential phosphorylation of S53 affected Keap1-Nrf2 binding in the pull-down assay, and induced nuclear translocation of Nrf2 in the electrophoretic mobility shift assay. Sequence homology analysis showed that S53 was highly conserved. Structural modeling around BTB domain of wild type and S53E-mutant Keap1 showed that the negative charge introduced by S53E mutation generates a salt bridge between E53 and ionized guanidine group of Arg50. Real-time qRT-PCR for transcription levels of antioxidant genes that are modulated by Nrf2 further proved the effects of the potential phosphorylation of S53 under an oxidative stress condition. In summary, S53 is a potential phosphorylation site of Keap1, and the phosphorylation could enhance the antioxidative capacity of cells in response to an oxidative stress.

  • L-arginine inhibited apoptosis of fish leukocytes via regulation of NF-κB-mediated inflammation, NO synthesis, and anti-oxidant capacity
    Biochimie (IF 3.188) Pub Date : 2018-12-19
    Haiou Zheng, Qian Guo, Xuzhuo Duan, Zhen Xu, Qingchao Wang

    The increased apoptosis plays an important role in bacterial invasion. In addition, LPS can induce inflammation and apoptosis of leukocytes via the production of reactive oxygen and nitrogen species. In the present study, we investigated the potential protective role of L-arginine (L-Arg) against the apoptosis of fish leukocytes in vitro. The results of Annexin V-FITC/PI staining and TUNEL assay indicated that L-Arg significantly alleviated the apoptosis of fish leukocytes induced by LPS at 24 h and 72 h post incubation (hpi). High caspase-3 activities induced by LPS at 72 hpi were significantly inhibited by L-Arg. Moreover, L-Arg supplementation also significantly decreased the mRNA expression levels of caspases at most time points, which contributed to the anti-apoptotic roles of L-Arg. Further analysis showed that L-Arg significantly inhibited the expression of several pro-inflammatory cytokines including IL-8 and TNF-α, partially via the down-regulation of the genes involved in NF-κB/MyD88 including NF-κB, IKKα and IKKγ. The down-regulation of these pro-inflammatory cytokines by L-Arg supplementation led to the further decrease in the expression of death receptor FasL, contributing to the anti-apoptotic effect of L-Arg. In addition, L-Arg supplementation increased both iNOS mRNA expression and NO production. The mRNA expressions of several anti-oxidant enzymes including SOD, CAT and GSHPx were also significantly increased after L-Arg supplementation, which accelerated the clearance of reactive oxygen species. In all, L-Arg inhibited apoptosis of fish leukocytes both via the increased NO production and antioxidant capacity and via the inhibition of inflammation mediated by NF-κB/MyD88 pathway.

  • Inhibitor Of Polyamine Catabolism Mdl72.527 Restores The Sensitivity To Doxorubicin Of Monocytic Leukemia Thp-1 Cells Infected With Human Cytomegalovirus
    Biochimie (IF 3.188) Pub Date : 2018-12-19
    Natalia E. Fedorova, Yana Yu. Chernoryzh, Galina R. Vinogradskaya, Svetlana S. Emelyanova, Larisa E. Zavalyshina, Kirill I. Yurlov, Natalia F. Zakirova, Valery N. Verbenko, Sergey N. Kochetkov, Alla A. Kushch, Alexander V. Ivanov

    Leukemic cells from different patients exhibit different sensitivity to anticancer drugs including doxorubicin (DOX). Resistance to chemotherapy decreases efficacy of the treatment and promotes cancer recurrence and metastases. One of the approaches to overcome drug resistance includes E2F1-mediated regulation of the р73 protein that belongs to the р53 family. Its ΔNp73 isoform exhibits pro-oncogenic effects, and TAp73 – anti-oncogenic effects. Human cytomegalovirus (HCMV), often found in tumors, suppresses pro-apoptotic pathways and E2F1/p73 in particular. The activity of E2F1 and p73 transcription factors is linked to metabolism of biogenic polyamines. Therefore, it could be suggested that compounds that target polyamine-metabolizing enzymes can sensitize HCMV-infected hematological malignancies to doxorubicin. Here we report that HCMV infection of ТНР-1 monocytic leukemic cells considerably elevates E2F1 levels and shifts the balance between the р73 isoforms towards ΔNp73 leading to survival of DOX-treated leukemic cells. In contrast, MDL72.527, an inhibitor of polyamine catabolism, decreases ΔNp73/ТАр73 ratio and thus restores sensitivity of the cells to DOX. Our findings indicate the combination of doxorubicin and MDL72.527 may present a novel strategy for therapy of leukemia in patients with and without HCMV infection.

  • Dioclea violacea lectin ameliorates inflammation in the temporomandibular joint of rats by suppressing intercellular adhesion molecule-1 expression
    Biochimie (IF 3.188) Pub Date : 2018-12-14
    Juliana T. Clemente-Napimoga, Maria A.S.M. Silva, Sylvia N.C. Peres, Alexandre H.P. Lopes, Claudia F. Lossio, Messias V. Oliveira, Vinicius J.S. Osterne, Kyria S. Nascimento, Henrique B. Abdalla, Juliana M. Teixeira, Benildo S. Cavada, Marcelo H. Napimoga

    Inflammation of temporomandibular joint (TMJ) tissues are the most common cause of pain conditions associated with temporomandibular disorders (TMDs). After a tissue and/or neural damage, the inflammatory response is characterized by plasma extravasation and leukocytes infiltration in the TMJ tissues, which in turn, release inflammatory cytokines cascades responsible for inflammatory pain. Lectins are glycoproteins widely distributed in nature that may exhibit anti-inflammatory properties. This study demonstrated by molecular docking and MM/PBSA that the lectin from Dioclea violacea (DVL) interacts favorably with α-methyl-D-mannoside, N-acetyl-D-glucosamine, and core1-sialyl-Lewis X which are associated with leukocytes migration during an inflammatory response. Wistar rats pretreated with intravenously injection of DVL demonstrated a significant inhibition of plasma extravasation induced by carrageenan (a non-neurogenic inflammatory inductor) and mustard oil (a neurogenic inflammatory inductor) in the TMJ periarticular tissues (p<0.05; ANOVA, Tukey’s test). In addition, DVL significantly reduced carrageenan-induced leukocyte migration in the TMJ periarticular tissues mediated by down-regulation of ICAM-1 expression. These results suggest a potential anti-inflammatory effect of DVL in inflammatory conditions of TMJ.

  • Parathyroid hormone-stimulation of Runx2 during osteoblast differentiation via the regulation of lnc-SUPT3H-1:16 (RUNX2-AS1:32) and miR-6797-5p
    Biochimie (IF 3.188) Pub Date : 2018-12-15
    B. Arumugam, M. Vishal, S. Shreya, D. Malavika, V. Rajpriya, Z. He, N.C. Partridge, N. Selvamurugan

    Parathyroid hormone (PTH) acts as a regulator of calcium homeostasis and bone remodeling. Runx2, an essential transcription factor in bone, is required for osteoblast differentiation. Noncoding RNAs such as long noncoding RNAs (lncRNAs) and microRNAs (miRNAs) play crucial roles in regulating gene expression in osteoblasts. In this study, we investigated the effects of PTH on osteoblast differentiation via Runx2, lncRNA, and miRNA expression in human bone marrow stromal cells (hBMSCs) and human osteoblastic cells (MG63). PTH-treatment of hBMSCs for 24 h, 7 days, and 14 days stimulated Runx2 mRNA expression. Using bioinformatics tools, we identified 17 lncRNAs originating from human Runx2 gene. Among these, lnc-SUPT3H-1:16 (RUNX2-AS1:32) expression was highly up-regulated by the 7 d PTH-treatment in hBMSCs. We also identified miR-6797-5p as the putative target of lnc-SUPT3H-1:16 and Runx2 using bioinformatics tools. PTH-treatment increased the expression of miR-6797-5p in hBMSCs, and overexpression of miR-6797-5p decreased osteoblast differentiation in MG63 cells, suggesting a role for lnc-SUPT3H-1:16 as sponge molecule. A luciferase gene reporter assay identified direct targeting of miR-6797-5p with lnc-SUPT3H-1:16 and 3´UTR Runx2 in MG63 cells. Thus, PTH stimulated the expression of lnc-SUPT3H-1:16, miR-6797-5p and Runx2, and due to the sponging mechanism of lnc- SUPT3H-1:16 towards miR-6797-5p, Runx2 was protected, resulting in the promotion of osteoblast differentiation.

  • Ribosomal protein eL42 contributes to the catalytic activity of the yeast ribosome at the elongation step of translation
    Biochimie (IF 3.188) Pub Date : 2018-12-11
    Codjo Hountondji, Jean-Bernard Créchet, Mayo Tanaka, Mieko Suzuki, Jun-ichi Nakayama, Blanche Aguida, Konstantin Bulygin, Jean Cognet, Galina Karpova, Soria Baouz

    The GGQ minidomain of the ribosomal protein eL42 was previously shown to contact the CCA-arm of P-site bound tRNA in human ribosome, indicating a possible involvement of the protein in the catalytic activity. Here, using Schizosaccharomyces pombe (S. pombe) cells, we demonstrate that the GGQ minidomain and neighboring region of eL42 is critical for the ribosomal function. Mutant eL42 proteins containing amino acid substitutions within or adjacent to the GGQ minidomain failed to complement the function of wild-type eL42, and expression of the mutant eL42 proteins led to severe growth defects. These results suggest that the mutations in eL42 interfere with the ribosomal function in vivo. Furthermore, we show that some of the mutations associated with the conserved GGQ region lead to reduced activities in the poly(Phe) synthesis and/or in the peptidyl transferase reaction with respect to puromycin, as compared with those of the wild-type ribosomes. A pK value of 6.95 was measured for the side chain of Lys-55/Arg-55, which is considerably less than that of a Lys or Arg residue. Altogether, our findings suggest that eL42 contributes to the 80S ribosome’s peptidyl transferase activity by promoting the course of the elongation cycle.

  • TRPC channels mediated calcium entry is required for proliferation of human airway smooth muscle cells induced by nicotine-nAChR
    Biochimie (IF 3.188) Pub Date : 2018-12-11
    Yongliang Jiang, Yumin Zhou, Gongyong Peng, Heshen Tian, Dan Pan, Lei Liu, Xing Yang, Chao Li, Wen Li, Ling Chen, Pixin Ran, Aiguo Dai

    The present study was designed to explore the role of transient receptor potential canonical 3 (TRPC3) in nicotine-induced chronic obstructive pulmonary disease (COPD) and its underlying mechanism. In this study, the expression and localization of α5 nicotinic acetylcholine receptor (α5-nAchR) in lung tissues were determined by western blotting and immunohistochemistry. The quantitative real-time PCR (qRT-PCR) analysis was performed to examine the mRNA expression levels of α5-nAchR and TRPC3 in human airway smooth muscle cells (HASMCs). Cell viability was assessed by CCK-8 assay. Proliferation was detected by cell counting and EdU immunofluorescent staining. Fluorescence calcium imaging was carried out to measure cytosolic Ca2+ ([Ca2+]cyt) concentration. The results showed that the α5-nAchR and TRPC3 expressions were significantly up-regulated in lung tissues of COPD smokers. Nicotine promoted HASMC proliferation, which was accompanied by elevated α5-nAchR and TRPC3 expressions, basal [Ca2+]cyt, store-operated calcium entry (SOCE) and the rate of Mn2+ quenching in HASMCs. Further investigation indicated that nicotine-induced Ca2+ response and TRPC3 up-regulation was reversibly blocked by small interfering RNA (siRNA) suppression of α5-nAChR. The knockdown of TRPC3 blunted Ca2+ response and HASMC proliferation induced by nicotine. In conclusion, nicotine-induced HASMC proliferation was mediated by TRPC3-dependent calcium entry via α5-nAchR, which provided a potential target for treatment of COPD.

  • MiR-204-5p promotes apoptosis and inhibits migration of osteosarcoma via targeting EBF2
    Biochimie (IF 3.188) Pub Date : 2018-12-06
    Mao Li, Yajun Shen, Qin Wang, Xuefeng Zhou

    Osteosarcoma is one of the most malignant cancer adolescents and young adults and metastatic osteosarcoma is a huge life threat with a 5-year survival lower than 20%. However, the mechanisms through which localized osteosarcoma turned metastatic are not fully understood. Here, we studied the role of miR-204-5p in osteosarcoma and found that miR-204-5p is downregulated in both osteosarcoma patients and osteosarcoma cell lines.In addition, overexpression of miR-204-5p resulted in increase of osteosarcoma cell apoptosis and decrease of osteosarcoma cell migration and invasion. Besides, our in vivo xenograft data showed strong inhibitory role of miR-204-5p in tumor growth. Importantly, our data showed that miR-204-5p regulates the mRNA stability of Early B Cell Factor 2 (EBF2), a crucial regulator in osteosarcoma apoptosis, by directly binding to 3’ UTR of EBF2. Besides, our data further revealed that overexpressed EBF2 inhibited apoptosis and facilitated migration and invasion of osteosarcoma cells. Additionally, EBF2 overexpression rescued the phenotype caused by miR-204-5p.Our data indicated that miR-204-5p is an anti-oncogenic miRNA in osteosarcoma which functions through inhibiting oncogenic transcription factor EBF2.These results provided new therapeutic targets for metastatic osteosarcoma and insights into molecular regulation of EBF2.

  • Folate and choline absorption and uptake: Their role in fetal development
    Biochimie (IF 3.188) Pub Date : 2018-12-06
    Anna Radziejewska, Agata Chmurzynska

    Scope In this review, we attempt to assess how choline and folate transporters affect fetal development. We focus on how the expression of these transporters in response to choline and folate intake affects transport effectiveness. We additionally describe allelic variants of the genes encoding these transporters and their phenotypic effects. Methods and results We made an extensive review of recent articles describing role of choline and folate - with particularl emphasize on their transporters - in fetal development. Folate and choline are necessary for the proper functioning of the cell and body.During pregnancy, the requirements of these nutrients increase because of elevated maternal demand and the rapid division of fetal cells. The concentrations of folate and choline in cells depend on food intake, the absorption of nutrients, and the cellular transport system, which is tissue-specific and developmentally regulated. Relatively few studies have investigated the role of choline transporters in fetal development. Conclusions In this review we show relations between functioning of folate and choline transporters and fetal development.

  • SOS genes contribute to Bac8c induced apoptosis-like death in Escherichia coli
    Biochimie (IF 3.188) Pub Date : 2018-12-05
    Heejeong Lee, Dong Gun Lee

    Antimicrobial peptides are one of the promising substitutes for conventional antibiotics. To examine their potential usefulness for controlling bacterial infections, the present study aimed to determine the induction of the self-destruction of Escherichia coli mediated by bac8c, which was modified from template peptide bactenecin. The potential of bac8c (RIWVIWRR) was reflected by the change in physiological following exposure, which led to the accumulation of reactive oxygen species (ROS) with an imbalance in the antioxidant system and damage to the intracellular lipids. In addition, bac8c-mediated death exhibited typical features to bacterial apoptosis-like death (ALD). The pathway was not enhanced in the absence of autocleavage of RecA and LexA, the SOS response proteins. We observed that elevated levels of intracellular ROS induced ALD, but this effect was insufficient before preceding RecA activation and LexA autocleavage. Furthermore, dinF, which encodes a member of the toxic compound extrusion (MATE) family, was evaluated to examine the role of the ALD pathway. dinF protein was required to accelerate the ALD-mediated bac8c in combination with RecA. Taken together, bac8c-mediated cell death underlies the severe SOS response, leading to RecA activation, LexA proteolysis, and dinF overexpression. Since then, overproduction of intracellular ROS facilitated the cell death.

  • Endonuclease G modulates the alternative splicing of deoxyribonuclease 1 mRNA in human CD4+ T lymphocytes and prevents the progression of apoptosis
    Biochimie (IF 3.188) Pub Date : 2018-12-03
    Dmitry D. Zhdanov, Yulia A. Gladilina, Vadim S. Pokrovsky, Dmitry V. Grishin, Vladimir A. Grachev, Valentina S. Orlova, Marina V. Pokrovskaya, Svetlana S. Alexandrova, Anna A. Plyasova, Nikolay N. Sokolov

    Apoptotic endonucleases act cooperatively to fragment DNA and ensure the irreversibility of apoptosis. However, very little is known regarding the potential regulatory links between endonucleases. Deoxyribonuclease 1 (DNase I) inactivation is caused by alternative splicing (AS) of DNase I pre-mRNA skipping exon 4, which occurs in response to EndoG overexpression in cells. The current study aimed to determine the role of EndoG in the regulation of DNase I mRNA AS and the modulation of its enzymatic activity. A strong correlation was identified between the EndoG expression levels and DNase I splice variants in human lymphocytes. EndoG overexpression in CD4+ T cells down-regulated the mRNA levels of the active full-length DNase I variant and up-regulated the levels of the non-active spliced variant, which acts in a dominant-negative fashion. DNase I AS was induced by the translocation of EndoG from mitochondria into nuclei during the development of apoptosis. The DNase I spliced variant was induced by recombinant EndoG or by incubation with EndoG-digested cellular RNA in an in vitro system with isolated cell nuclei. Using antisense DNA oligonucleotides, we identified a 72-base segment that spans the adjacent segments of exon 4 and intron 4 and appears to be responsible for the AS. DNase I-positive CD4+ T cells overexpressing EndoG demonstrated decreased progression towards bleomycin-induced apoptosis. Therefore, EndoG is an endonuclease with the unique ability to inactivate another endonuclease, DNase I, and to modulate the development of apoptosis.

  • Serotonergic neurotransmission manipulation for the understanding of brain development and function: learning from Tph2 genetic models
    Biochimie (IF 3.188) Pub Date : 2018-12-01
    Marta Pratelli, Massimo Pasqualetti

    Serotonin (5-hydroxytriptamine; 5-HT) is a fascinating neurotransmitter that thanks to an extensive axonal network is released throughout the entire central nervous system (CNS) and exerts its action on the modulation of a countless number of physiological, behavioral and cognitive processes. In addition, cumulating evidences have linked alteration in 5-HT neurotransmission with the onset of psychiatric and neurodevelopmental disorders, such as depression, autisms and schizophrenia. Nevertheless only 5% of the total body content of serotonin exerts its action in the CNS, while the rest is synthetized and stored in peripheral tissues where it acts as an autacoid. In 2003 it became evident that two distinct isoforms of tryptophan hydroxylase (Tph), the rate-limiting enzyme for the synthesis of serotonin, are selectively expressed in peripheral tissues and in the CNS, with Tph2 as the brain specific isoform. In the present review we describe how the discovery of Tph2 has improved our understanding on the role of serotonergic neurotransmission. We mainly focus on the analysis of animal models generated by genetic manipulation of Tph2, in which the synthesis of brain serotonin was either reduced or disrupted. The consequences of an altered serotonergic neurotransmission on brain development, as well as on physiological and behavioral processes will be assessed. Finally, we report on several association studies that have linked single nucleotide polymorphisms (SNPs) in the human TPH2 gene with behavioral disturbances and neuropsychiatric disorders.

  • Oleate increase neutral lipid accumulation, cellular respiration and rescues palmitate-exposed GLP-1 secreting cells by reducing ceramide-induced ROS
    Biochimie (IF 3.188) Pub Date : 2018-12-01
    Stelia Ntika, Ketan Thombare, Masood Aryapoor, Hjalti Kristinsson, Peter Bergsten, Camilla Krizhanovskii

    Background Fatty acids (FAs), and especially monounsaturated FAs (MUFAs) stimulate GLP-1 release. However, lipotoxicity is indicated in GLP-1 secreting cells following long-term exposure to elevated levels of saturated FAs (SFAs) in vivo and in vitro, where in vitro studies indicate that cosupplementation with MUFAs confers lipoprotection. SFAs and MUFAs differentially affect the fate of cells in ways that depend on the cell type, concentration and ratio of the FAs. The present study was designed to further elucidate the mechanisms underlying the effects of SFAs/MUFAs on GLP-1-producing cells in terms of lipotoxicity/lipoprotection and GLP-1 secretion. Methods Cultured GLP-1 secreting cells were exposed to hyperlipidemia simulated by SFA-albumin complexes where the molar ratio was 2:1. The cellular response to simulated hyperlipidemia was assessed in the presence/absence of MUFA cosupplementation by determining intracellular ceramide, ROS, neutral lipid accumulation, and cellular respiration. The role for cellular respiration in GLP-1 secretion in response to SFAs/MUFAs was assessed. Results Generation of intracellular ceramide mediate a detrimental increased in ROS production following long term exposure to SFAs in GLP-1-secreting cells. Cosupplementation with MUFAs increases cellular respiration, triglyceride synthesis, and the expression of ceramide kinase, while reducing ceramide synthesis and attenuating ROS production, caspase-3 activity and DNA fragmentation. Further, acute secretory effects of unsaturated FAs are independent of FAO, but mediated by a FFAR1 induced increase in cellular respiration. Conclusion This study demonstrates novel data supporting effects of MUFAs on the ceramide biosynthetic pathway, triglyceride storage respiration and secretion in GLP-1 secreting cells. These findings may be of value for nutritional interventions, as well as for identification of novel targets, to help preserve L-cell mass and potentiate GLP-1 secretion in diabesity.

  • Identification of SEPTIN12 as a novel target of the androgen and estrogen receptors in human testicular cells
    Biochimie (IF 3.188) Pub Date : 2018-12-01
    Pao-Lin Kuo, Jie-Yun Tseng, Hau-Ing Chen, Chia-Yun Wu, Hany A. Omar, Chia-Yih Wang, Han-Yi Cheng, Chao-Chin Hsu, Tzu-Fun Fu, Yen-Ni Teng

    SEPTIN12 (SEPT12) is a testis-enriched gene that is downregulated in the testis of infertile men with severe spermatogenic defects. While SEPT12 is involved in spermatogenic failure and sperm motility disorder, SEPT12 transcriptional regulation is still unknown. Here we report the promoter region of SEPT12 as a 245 bp segment upstream from the transcription start site. One androgen receptor (AR) and two of estrogen receptor α (ERα) binding sites were on this region were identified initially by Bioinformatics prediction and confirmed by chromatin immunoprecipitation assay. Truncated ERα or AR binding sites decreased the promoter activity, which indicated that the ERα and AR are essential for the SEPT12 promoter. On the other hand, the promoter activity was enhanced by the treatment with 17β-estradiol (E2) and 5α-dihydrotestosterone (5α-DHT). Thus, one androgen and two estrogen hormone responsive elements were in the promoter of SEPT12 gene to regulate the SEPT12 expression. Two single nucleotide polymorphisms (SNPs), rs759992 T>C and rs3827527 C>T, were observed in the SEPT12 gene promoter region and were able to decrease the promoter activity. In conclusion, the current work identified the promoter of the human SEPT12 gene and provided key evidence about its transcriptional regulation via E2 and 5α-DHT. Since SEPT12 has an important role in spermatogenesis, SEPT12 expression analysis can be developed as a potential tool for the assessment of environmental or food pollution by hormones or for the evaluation of the risk of endocrine-disrupting chemicals (EDCs) in general.

  • Parametric study of immobilized cellulase-polymethacrylate particle for the hydrolysis of carboxymethyl cellulose
    Biochimie (IF 3.188) Pub Date : 2018-12-01
    Yi Wei Chan, Caleb Acquah, Eugene M. Obeng, Elvina C. Dullah, Jaison Jeevanandam, Clarence M. Ongkudon

    Biocarriers are pivotal in enhancing the reusability of biocatalyst that would otherwise be less economical for industrial application. Ever since the induction of enzymatic technology, varied materials have been assessed for their biocompatibility with enzymes of distinct functionalities. Herein, cellulase was immobilized onto polymethacrylate particles (ICP) as the biocarrier grafted with ethylenediamine (EDA) and glutaraldehyde (GA). Carboxymethyl cellulose (CMC) was used as a model substrate for activity assay. Enzyme immobilization loading was determined by quantifying the dry weight differential of ICP (pre-& post-immobilization). Cellulase was successfully demonstrated to be anchored upon ICP and validated by FTIR spectra analysis. The optimal condition for cellulase immobilization was determined to be pH 6 at 20 oC. The maximum CMCase activity was achieved at pH 5 and 50 oC. Residual activity of ∼50 % was retained after three iterations and dipped to ∼18 % on following cycle. Also, ICP displayed superior pH adaptability as compared to free cellulase. The specific activity of ICP was 65.14 ±1.11 % relative to similar amount of free cellulase.

  • Serotonin in the gut: Blessing or a curse
    Biochimie (IF 3.188) Pub Date : 2018-06-14
    Suhrid Banskota, Jean-Eric Ghia, Waliul I. Khan

    Serotonin (5-hydroxytryptamine or 5-HT) once most extensively studied as a neurotransmitter of the central nervous system, is seen to be predominantly secreted in the gut. About 95% of 5-HT is estimated to be found in gut mainly within the enterochromaffin cells whereas about 5% is found in the brain. 5-HT is an important enteric signaling molecule and is well known for playing a key role in sensory-motor and secretory functions in the gut. In recent times, studies uncovering various new functions of gut-derived 5-HT indicate that many more are yet to be discovered in coming days. Recent studies revealed that 5-HT plays a pivotal role in immune cell activation and generation/perpetuation of inflammation in the gut. In addition to its various roles in the gut, there are now emerging evidences that suggest an important role of gut-derived 5-HT in other biological processes beyond the gut, such as bone remodeling and metabolic homeostasis. This review focuses to briefly summarize the accumulated and newly updated role of 5-HT in the maintenance of normal gut physiology and in the pathogenesis of inflammation in the gut. The collected information about this multifaceted signaling molecule may aid in distinguishing its good and bad effects which may lead to the development of novel strategies to overcome the unwanted effect, such as in inflammatory bowel disease.

  • Serotonin and human cancer: A critical view
    Biochimie (IF 3.188) Pub Date : 2018-06-21
    Denis Sarrouilhe, Marc Mesnil

    Besides its classical functions as a neurotransmitter in the central nervous system, local mediator in the gastrointestinal tract and vasoactive agent in the blood, serotonin has more recently emerged as a growth factor for human tumor cells of different origins (carcinomas, glioma and carcinoids). Several data are also available on serotonin involvement in cancer cell migration, metastatic dissemination and tumor angiogenesis. The serotonin-induced signaling pathways that promote tumor progression are complex and only partly understood in some cancer types. The results of several studies showed that serotonin levels in the tumor played a crucial role in cancer progression. A serotonin production and secretion by neuroendocrine cells have been shown in the progression of several solid tumors and the involvement of a serotoninergic autocrine loop was proposed. Specific receptor subtypes are associated with different fundamental stages of tumor progression and the pattern of receptors expression becomes dysregulated in several human tumors when compared with normal cells or tissues. Serotonin receptors, selective serotonin transporter and serotonin synthesis pathways are potential chemotherapeutic targets for the treatment of several cancers in which therapeutic approaches are limited. Through several asked questions, this critical mini-review discusses the relevance of the involvement of serotonin in human cancer progression.

  • A cross-talk between fat and bitter taste modalities
    Biochimie (IF 3.188) Pub Date : 2018-06-22
    Amira S. Khan, Babar Murtaza, Aziz Hichami, Naim A. Khan

    The choice of food is governed largely by the sense of taste. To date, five basic taste modalities have been described; however, there is an increasing agreement on the existence of a 6th fat taste. The taste modalities might interact with each other and also with other senses. The advancements in cellular and molecular biology have helped the characterization of taste signaling mechanisms, down to the receptor level and beyond. CD36 and GPR120 have been shown to be involved in the detection of fat taste while bitter taste is perceived by a number of receptors that belong to a family of taste-type 2 receptors (T2R or TAS2R). Hence, the most common role is played by TAS2R16 and TAS2R38 in bitter taste perception in humans. Increasing evidences from behavioural studies suggest that fat and bitter taste modalities might interact with each other, and this interaction might be critical in obesity. In the current review, we will discuss the evidence from genetic and behavioural studies and propose the molecular mechanism of a cross-talk between fat and bitter tastes.

  • Experimental human endotoxemia as a model of systemic inflammation
    Biochimie (IF 3.188) Pub Date : 2018-06-22
    Dirk van Lier, Christopher Geven, Guus P. Leijte, Peter Pickkers

    Systemic inflammation plays a pivotal role in a multitude of conditions, including sepsis, trauma, major surgery and burns. However, comprehensive analysis of the pathophysiology underlying this systemic inflammatory response is greatly complicated by variations in the immune response observed in critically ill patients, which is a result of inter-individual differences in comorbidity, comedication, source of infection, causative pathogen, and onset of the inflammatory response. During experimental human endotoxemia, human subjects are challenged with purified endotoxin (lipopolysaccharide) intravenously which induces a short-lived, well-tolerated and controlled systemic inflammatory response, similar to that observed during sepsis. The human endotoxemia model can be conducted in a highly standardized and reproducible manner, using a carefully selected homogenous study population. As such, the experimental human endotoxemia model does not share the aforementioned clinical limitations and enables us to investigate both the mechanisms of systemic inflammation, as well as to evaluate novel (pharmacological) interventions in humans in vivo. The present review provides a detailed overview of the various designs, organ-specific changes, and strengths and limitations of the experimental human endotoxemia model, with the main focus on its use as a translational model for sepsis research.

  • Fatty acids and oxidized lipoproteins contribute to autophagy and innate immunity responses upon the degeneration of retinal pigment epithelium and development of age-related macular degeneration
    Biochimie (IF 3.188) Pub Date : 2018-07-18
    Kai Kaarniranta, Ali Koskela, Szabolcs Felszeghy, Niko Kivinen, Antero Salminen, Anu Kauppinen

    Retinal pigment epithelium (RPE) damage is a primary sign in the development of age-related macular degeneration (AMD) the leading cause of blindness in western countries. RPE cells are exposed to chronic oxidative stress due to constant light exposure, active fatty acid metabolism and high oxygen consumption. RPE cells phagocytosize lipid rich photoreceptor outer segment (POS) which is regulated by circadian rhytmn. Docosahexaenoic acid is present in high quantity in POS and increases oxidative stress, while its metabolites have cytoprotective effects in RPE. During RPE aging, reactive oxygen species and oxidized lipoproteins are considered to be major causes of disturbed autophagy clearance that lead to chronic innate immunity response involved in NOD-Like, Toll-Like, Advanced Glycation End product Receptors (NLRP, TLR, RAGE, respectively), pentraxins and complement systems. We discuss role of fatty acids and lipoproteins in the degeneration of RPE and development of AMD.

  • Cloning, purification and characterization of nigrelysin, a novel actinoporin from the sea anemone Anthopleura nigrescens
    Biochimie (IF 3.188) Pub Date : 2018-07-21
    Javier Alvarado-Mesén, Frank Solano-Campos, Liem Canet, Lohans Pedrera, Yadira P. Hervis, Carmen Soto, Henry Borbón, María E. Lanio, Bruno Lomonte, Aisel Valle, Carlos Alvarez

    Actinoporins constitute a unique class of pore-forming toxins found in sea anemones that being secreted as soluble monomers are able to bind and permeabilize membranes leading to cell death. The interest in these proteins has risen due to their high cytotoxicity that can be properly used to design immunotoxins against tumor cells and antigen-releasing systems to cell cytosol. In this work we describe a novel actinoporin produced by Anthopleura nigrescens, an anemone found in the Central American Pacific Ocean. Here we report the amino acid sequence of an actinoporin as deduced from cDNA obtained from total body RNA. The synthetic DNA sequence encoding for one cytolysin variant was expressed in BL21 Star (DE3) Escherichia coli and the protein purified by chromatography on CM Sephadex C-25 with more than 97% homogeneity as verified by MS-MS and HPLC analyses. This actinoporin comprises 179 amino acid residues, consistent with its observed isotope-averaged molecular mass of 19 661 Da. The toxin lacks Cys and readily permeabilizes erythrocytes, as well as L1210 cells. CD spectroscopy revealed that its secondary structure is dominated by beta structure (58.5%) with 5.5% of α-helix, and 35% of random structure. Moreover, binding experiments to lipidic monolayers and to liposomes, as well as permeabilization studies in vesicles, revealed that the affinity of this toxin for sphingomyelin-containing membranes is quite similar to sticholysin II (StII). Comparison by spectroscopic techniques and modeling the three-dimensional structure of nigrelysin (Ng) showed a high homology with StII but several differences were also detectable. Taken together, these results reinforce the notion that Ng is a novel member of the actinoporin pore-forming toxin (PFT) family with a HA as high as that of StII, the most potent actinoporin so far described, but with peculiar structural characteristics contributing to expand the understanding of the structure-function relationship in this protein family.

  • A stochastic approach to serotonergic fibers in mental disorders
    Biochimie (IF 3.188) Pub Date : 2018-07-26
    Skirmantas Janušonis, Nils Detering

    Virtually all brain circuits are physically embedded in a three-dimensional matrix of fibers that release 5-hydroxytryptamine (5-HT, serotonin). The density of this matrix varies across brain regions and cortical laminae, and it is altered in some mental disorders, including Major Depressive Disorder and Autism Spectrum Disorder. We investigate how the regional structure of the serotonergic matrix depends on the stochastic behavior of individual serotonergic fibers and introduce a new framework for the quantitative analysis of this behavior. In particular, we show that a step-wise random walk, based on the von Mises-Fisher probability distribution, can provide a realistic and mathematically concise description of these fibers. We also consider other stochastic models, including the fractional Brownian motion. The proposed approach seeks to advance the current understanding of the ascending reticular activating system (ARAS) and may also support future theory-guided therapeutic approaches.

  • Sphingolipid metabolism in non-alcoholic fatty liver diseases
    Biochimie (IF 3.188) Pub Date : 2018-07-31
    Marion Régnier, Arnaud Polizzi, Hervé Guillou, Nicolas Loiseau

    Non-alcoholic fatty liver disease (NAFLD) involves a panel of pathologies starting with hepatic steatosis and continuing to irreversible and serious conditions like steatohepatitis (NASH) and hepatocarcinoma. NAFLD is multifactorial in origin and corresponds to abnormal fat deposition in liver. Even if triglycerides are mostly associated with these pathologies, other lipid moieties seem to be involved in the development and severity of NAFLD. That is the case with sphingolipids and more particularly ceramides. In this review, we explore the relationship between NAFLD and sphingolipid metabolism. After providing an analysis of complex sphingolipid metabolism, we focus on the potential involvement of sphingolipids in the different pathologies associated with NAFLD. An unbalanced ratio between ceramides and terminal metabolic products in the liver and plasma promotes weight gain, inflammation, and insulin resistance. In the etiology of NAFLD, some sphingolipid species such as ceramides may be potential biomarkers for NAFLD. We review the clinical relevance of sphingolipids in liver diseases.

  • The multifaceted functions of lipopolysaccharide in plant-bacteria interactions
    Biochimie (IF 3.188) Pub Date : 2018-08-02
    Alexander Kutschera, Stefanie Ranf

    In Gram-negative bacteria, the cell envelope largely consists of lipopolysaccharide (LPS), a class of heterogeneous glycolipids. As a fundamental component of the outer membrane, LPS provides stability to the bacterial cell and forms a protective cover shielding it from hostile environments. LPS is not only fundamental to bacterial viability, but also makes a substantial contribution both directly and indirectly to multiple aspects of inter-organismic interactions. During infection of animal and plant hosts, LPS promotes bacterial virulence but simultaneously betrays bacteria to the host immune system. Moreover, dynamic remodulation of LPS structures allows bacteria to fine-tune OM properties and quickly adapt to diverse and often hostile environments, such as those encountered in host tissues. Here, we summarize recent insights into the multiple functions of LPS in plant-bacteria interactions and discuss what we can learn from the latest advances in the field of animal immunity. We further pinpoint open questions and future challenges to unravel the different roles of LPS in the dynamic interplay between bacteria and plant hosts at the mechanistic level.

  • Serotonin in stem cell based-dental repair and bone formation: A review
    Biochimie (IF 3.188) Pub Date : 2018-08-02
    Anne Baudry, Benoit Schneider, Jean-Marie Launay, Odile Kellermann

    Genetic and pharmacological studies provided evidence that serotonin (5-HT) is an important signaling molecule for the development and the maintenance of mineralized tissues. However, how 5-HT takes part to the homeostasis of teeth and bone remains elusive. In the dental field, a major breakthrough comes from the identification of 5-HT but also dopamine (DA) as "damage" signals necessary for stem cell-based tooth repair. Pulpal stem cells express the overall functions of 5-HT and DA neurons including a definite set of functional 5-HT/DA receptors that render cells responsive for circulating bioamines. Upon tooth injury, activated platelets release bulks of 5-HT/DA that mobilize pulpal stem cells for natural dental repair. The contribution of 5-HT to bone metabolism is more documented with description of both anabolic and resorptive effects. By controlling the tissue-non specific alkaline phosphatase (TNAP), 5-HT2B receptors exert an anabolic function and a pivotal role in mineralization processes. Increasing our understanding of the role of 5-HT receptors in bone metabolism may pave the road for the development of therapeutic strategies towards skeletal-associated pathologies and ectopic calcification.

  • A chronic LPS-induced low-grade inflammation fails to reproduce in lean mice the impairment of preference for oily solution found in diet-induced obese mice
    Biochimie (IF 3.188) Pub Date : 2018-08-10
    Arnaud Bernard, Déborah Ancel, Patricia Passilly-Degrace, Jean-François Landrier, Laurent Lagrost, Philippe Besnard

    Diet-induced obesity (DIO) is associated with a decreased oral fat detection in rodents. This alteration has been explained by an impairment of the lipid-mediated signaling in taste bud cells (TBC). However, factors responsible for this defect remain elusive. Diet rich in saturated fatty acids is known to elicit a metabolic inflammation by promoting intestinal permeation to lipopolysaccharides (LPS), Gram-negative bacteria-derived endotoxins. To determine whether a local inflammation of the gustatory tissue might explain the obese-induced impairment of the oro-sensory detection of lipids, mice were subjected to a DIO protocol. Using a combination of behavioral tests, transcriptomic analyses of gustatory papillae and biochemical assays, we have found that i) DIO elicits a pro-inflammatory genic profile in the circumvallate papillae (CVP), known to house the highest density of lingual taste buds, ii) NFkB, a key player of inflammatory process, might play a role in this transcriptomic pattern, iii) plasma LPS levels are negatively correlated with the preference for oily solution, and iv) a chronic infusion of LPS at a level similar to that found in DIO mice is not sufficient to alter the spontaneous preference for fat in lean mice. Taken together these data bring the demonstration that a saturated high fat diet elicits an inflammatory response at the level of peripheral gustatory pathway and a LPS-induced low-grade endotoxemia alone does not explain the change in the preference for dietary lipids observed in DIO mice.

Some contents have been Reproduced with permission of the American Chemical Society.
Some contents have been Reproduced by permission of The Royal Society of Chemistry.
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